U.S. patent application number 15/875858 was filed with the patent office on 2018-09-06 for treatment of cancer by infusion of oncolytic herpes simplex virus to the blood.
The applicant listed for this patent is VIRTTU BIOLOGICS LIMITED. Invention is credited to Joe Conner.
Application Number | 20180250352 15/875858 |
Document ID | / |
Family ID | 56550907 |
Filed Date | 2018-09-06 |
United States Patent
Application |
20180250352 |
Kind Code |
A1 |
Conner; Joe |
September 6, 2018 |
TREATMENT OF CANCER BY INFUSION OF ONCOLYTIC HERPES SIMPLEX VIRUS
TO THE BLOOD
Abstract
An oncolytic herpes simplex virus is disclosed for use in a
method of treating cancer in a human subject, the method comprising
administering to the human subject at least one dose of oncolytic
herpes simplex virus by infusion to the blood, wherein the
oncolytic herpes simplex virus reaches cells of the cancer in which
it replicates.
Inventors: |
Conner; Joe; (Newhouse,
GB) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
VIRTTU BIOLOGICS LIMITED |
Newhouse |
|
GB |
|
|
Family ID: |
56550907 |
Appl. No.: |
15/875858 |
Filed: |
January 19, 2018 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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PCT/GB2016/052177 |
Jul 19, 2016 |
|
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15875858 |
|
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 39/39566 20130101;
A61K 9/0019 20130101; A61K 9/08 20130101; A61K 39/3955 20130101;
C07K 2317/73 20130101; A61K 45/06 20130101; A61K 35/768 20130101;
A61K 2039/505 20130101; A61K 47/12 20130101; A61P 35/00 20180101;
C07K 16/2818 20130101; C07K 2317/76 20130101; A61K 35/763 20130101;
A61K 2039/57 20130101; A61K 39/3955 20130101; A61K 2300/00
20130101 |
International
Class: |
A61K 35/763 20060101
A61K035/763; A61P 35/00 20060101 A61P035/00; C07K 16/28 20060101
C07K016/28; A61K 39/395 20060101 A61K039/395 |
Foreign Application Data
Date |
Code |
Application Number |
Jul 20, 2015 |
GB |
1512723.6 |
Dec 30, 2015 |
GB |
1523091.5 |
Claims
1. A method of treating cancer in a human subject, the method
comprising administering to the human subject at least one dose of
oncolytic herpes simplex virus by infusion to the blood.
2.-3. (canceled)
4. The method of claim 1, wherein the method comprises determining
whether oncolytic herpes simplex virus DNA is present in a sample
of the subject's blood.
5. (canceled)
6. The method of claim 1, wherein the method comprises determining
whether oncolytic herpes simplex virus is present in a sample of
the subject's tumor tissue.
7. (canceled)
8. The method of claim 1, wherein the oncolytic herpes simplex
virus is a mutant of HSV-1 strain 17 or is HSV1716.
9. The method of claim 1, wherein the cancer is a solid tumor, a
recurrent or metastatic solid tumor, or a non-CNS solid tumor.
10. The method of claim 1, wherein the dose of oncolytic herpes
simplex virus administered is at least 1.times.10.sup.6 iu.
11. The method of claim 1, wherein a dose of oncolytic herpes
simplex virus is administered over a period of 3 hours or less.
12. The method of claim 1, wherein the administered oncolytic
herpes simplex virus is formulated as about 0.5 ml to about 5 ml of
a suspension of virus in about 200 ml to about 300 ml of lactated
Ringer's solution.
13. The method of claim 1, wherein the method comprises
administering to the human subject at least one treatment cycle of
oncolytic herpes simplex virus, wherein a treatment cycle comprises
of at least two doses of oncolytic herpes simplex virus, each dose
administered by infusion to the blood wherein the second and
subsequent doses are each administered within about 17 days of the
preceding dose, each dose of oncolytic herpes simplex virus being
in the range about 1.times.10.sup.6 iu to about 1.times.10.sup.8
iu.
14. The method of claim 13, wherein one dose of oncolytic herpes
simplex virus is administered per week.
15. The method of claim 13, wherein two doses of oncolytic herpes
simplex virus are administered per week.
16. The method of claim 13, wherein each dose of oncolytic herpes
simplex virus is in the range about 1.times.10.sup.7 iu to about
1.times.10.sup.8 iu.
17. (canceled)
18. The method of claim 13, wherein the treatment cycle comprises
of administration of a therapeutically effective amount of an
immune checkpoint inhibitor selected from the group consisting of
an inhibitor of PD-1, PD-L1, CTLA4, TIM-3 or LAG-3.
19. The method of claim 13, wherein the subject receives two or
more treatment cycles.
20. The method of claim 13, wherein the method comprises
determining the presence of a Th1 response in the subject.
21.-59. (canceled)
60. The method of claim 1, wherein the cancer is in a child.
61. The method of claim 1, wherein the oncolytic herpes simplex
virus does not encode or is not further modified to contain nucleic
acid encoding a cytokine or chemokine, an interleukin, an
interferon, a tumor necrosis factor, a colony stimulating factor,
an immune modulator, a member of the CC family, a member of the CXC
family or a member of the CXC family.
62. The method of claim 1, wherein the oncolytic herpes simplex
virus does not express GMCSF.
63. The method of claim 1, wherein the oncolytic herpes simplex
virus encodes a functional ICP47, a functional ICP6 gene, or both a
functional ICP47 gene and a functional ICP6 gene.
64. The method of claim 1, wherein the cancer is not a
melanoma.
65.-72. (canceled)
Description
RELATED APPLICATIONS
[0001] This application is a continuation of International Patent
Application No. PCT/GB2016/052177, filed Jul. 19, 2016, which
claims priority to and the benefit of United Kingdom patent
application No. 1512723.6 filed on 20 Jul. 2015 and United Kingdom
patent application No. 1523091.5 filed on 30 Dec. 2015. The entire
contents of these applications are incorporated herein by
reference.
FIELD OF THE INVENTION
[0002] The present invention relates to methods of treatment of
human cancers involving administration of an oncolytic herpes
simplex virus to a human subject by infusion of the virus to the
blood.
BACKGROUND TO THE INVENTION
[0003] Oncolytic virotherapy concerns the use of lytic viruses
which selectively infect and kill cancer cells. Some oncolytic
viruses are promising therapies as they display exquisite selection
for replication in cancer cells and their self-limiting propagation
within tumors results in fewer toxic side effects. Several
oncolytic viruses have shown great promise in the clinic (Bell, J.,
Oncolytic Viruses: An Approved Product on the Horizon? Mol Ther.
2010; 18(2): 233-234; Russell et al., (Oncolytic Virotherapy.
Nature Biotechnology Vol. 30 No. 7 Jul. 2012).
Talimogene Laherparepvec
[0004] After a long period of development, and question marks over
the validity of the approach, oncolytic virotherapy has
significantly moved forward with the first approval by the US Food
and Drug Administration ("FDA") and European Medicines Agency
("EMA") of an oncolytic virus therapy.
[0005] The oncolytic virus talimogene laherparepvec (IMLYGIC.RTM.)
was approved by FDA on 27.sup.th October 2015 for the "local
treatment of unresectable cutaneous, subcutaneous, and nodal
lesions in patients with melanoma recurrent after initial surgery"
and by EMA on 16 Dec. 2015 for the "treatment of of adults with
unresectable melanoma that is regionally or distantly metastatic
(Stage IIIB, IIIC and IVM1a) with no bone, brain, lung or other
visceral disease".
[0006] Talimogene laherparepvec is an attenuated herpes simplex
virus type-1 (HSV-1) derived by functional deletion of two genes
(ICP34.5 and ICP47) and insertion of coding sequence for human
granulocyte macrophage colony-stimulating factor (GM-CSF) and is
produced in Vero cells by recombinant DNA technology.
[0007] The safety and efficacy of Imlygic.RTM. monotherapy compared
with subcutaneously administered GMCSF were evaluated in a phase 3,
multinational, open-label, and randomised clinical study of
patients with stage IIIB, IIIC, and IV melanoma that was not
considered to be surgically resectable. Previous systemic treatment
for melanoma was allowed but not required. Patients with active
brain metastases, bone metastases, extensive visceral disease,
primary ocular or mucosal melanoma, evidence of immunosuppression,
or receiving treatment with a systemic anti-herpetic agent were
excluded from the study.
[0008] The results of the study are summarised in the EMA's Summary
of Product Characteristics, which states "In an analysis to
evaluate systemic activity of Imlygic, 27 of 79 patients (34.2%)
had a .gtoreq.50% overall decrease in non-visceral lesions that
were not injected with Imlygic, and 8 of 71 patients (11.3%) had a
.gtoreq.50% overall decrease in visceral lesions that were not
injected with Imlygic."
[0009] Exploratory subgroup analyses for durable response rate
(DRR) and overall survival by stage of disease were also carried
out. While the pivotal study was not powered to evaluate efficacy
in these individual subgroups, patients with no visceral disease
derived greater benefit from Imlygic.RTM. treatment than those with
more advanced disease. The secondary endpoint on overall survival
just missed statistical significance.
[0010] The labels approved by both FDA and EMA indicate that
talimogene laherparepvec is approved for injection into cutaneous,
subcutaneous, and/or nodal lesions.
[0011] The FDA approval sets out the dosing regime for talimogene
laherparepvec, as follows: "Your healthcare professional will
inject Imlygic directly into your tumour(s) with a needle and
syringe. Your second injection will be given 3 weeks after the
first injection. After that, you will receive injections every 2
weeks for as long as you have the tumour(s). Your healthcare
professional will decide which tumour(s) to inject and may not
inject every tumour. Your existing tumour(s) may increase in size
and new tumour(s) could appear while you are being treated with
Imlygic. You can expect to be treated with Imlygic for at least 6
months or longer." The EMA approval contains a similar
statement.
[0012] The dosing schedule for Imlygic.RTM. involves an initial
injection of a dose of 10.sup.6 pfu/ml into the largest lesion and
other lesions are prioritised by lesion size for injection until
the maximum injection volume of 4 ml is reached. In the second and
subsequent treatments the dose of virus is 10.sup.8 pfu/ml, new
lesions are injected first and other lesions are prioritised by
lesion size for injection until the maximum injection volume of 4
ml is reached. In subsequent visits new lesions developing since
previous treatments are injected first and other lesions are
prioritised by lesion size for injection until the maximum
injection volume of 4 ml is reached.
[0013] The volume of Imlygic.RTM. to be injected into each lesion
is dependent on the size of the lesion and is determined according
to a table. The total injection volume for each treatment session
is a maximum of 4 ml.
[0014] Patients may experience increase in size of existing
lesion(s) or the appearance of a new lesion prior to achieving a
response. As long as there are injectable lesion(s) remaining,
Imlygic.RTM. should be continued for at least 6 months unless the
physician considers that the patient is not benefitting from
Imlygic.RTM. treatment or that other treatment is required.
[0015] Imlygic.RTM. treatment may be reinitiated if new lesions
appear following a complete response and the physician considers
that the patient will benefit from treatment.
[0016] The dosing regime involves numerous injections in different
locations, which is generally disadvantageous for the patient.
Injection site reactions were very common in the Phase 3 study with
27.2% of patients reporting such incidents and included: injection
site pain, erythema, haemorrhage, swelling, inflammation, secretion
and discharge. In particular, cellulitis and systemic bacterial
infection were reported.
[0017] During preclinical testing in immunocompetent mice, rats,
and dogs, talimogene laherparepvec was administered by
subcutaneous, intravenous or intratumoral injection.
[0018] Talimogene laherparepvec was injected into various xenograft
tumours at doses up to 2.times.10.sup.8 PFU/kg in immunodeficient
mice (nude and SCID). Lethal systemic viral infection was observed
in up to 20% of nude mice (primarily deficient in T lymphocyte
function) and 100% of SCID mice (devoid of both T and B
lymphocytes). Across the studies, fatal disseminated viral
infection was observed in 14% of nude mice following treatment with
talimogene laherparepvec at doses that are 10 to 100-fold higher
than those that result in 100% lethality with wild-type HSV-1.
Imlygic has not been studied in immunocompromised human patients
but the animal data indicates that patients who are severely
immunocompromised may be at an increased risk of disseminated
herpetic infection and the FDA approval indicates that they should
not be treated with Imlygic.RTM..
[0019] In summary, the pre-clinical and clinical studies of
talimogene laherparepvec indicate that it has some measurable
clinical benefit in a sub-group of melanoma patients with local
disease but no significant benefit in patients with visceral
lesions or advanced metastatic disease. It has also failed to
statistically establish an overall survival benefit in any group of
patients and shows limited evidence of a systemic effect,
particularly in late stage and visceral disease. Administration to
human patients is exclusively by injection with notable patient
reactions and the pre-clinical data indicates an unsuitability for
systemic administration.
Experience with HSV1716
[0020] In many animal models, oncolytic viruses (including
oncolytic herpes simplex viruses) work spectacularly well but
repeating the results in human trials has proven challenging. The
difficulties in translating biologic agents with complex (and not
fully understood) structures and complex mechanisms of action will
be apparent and common general knowledge to those skilled in the
art. For example, when it comes to developing treatments for human
patients that involve directed stimulation of parts of the human
immune system studies of the effect of oncolytic virus on the
immune response in mice (and other non-human animals) must be
treated with caution owing to acknowledged differences in immune
response between mouse models and humans (Mestas and Hughes., J
Immunol 2004; 172:2731-2738).
[0021] HSV1716 has been in clinical development for a number of
years. Clinical studies have investigated safety and mechanism of
action of direct intra-tumoral administration for the treatment of
localised brain tumors, melanoma, head and neck cancer. These
studies have demonstrated some proof of safety and mechanism of
action in tumors in which the agent was directly administered by
injection but no evidence of systemic benefit in uninjected lesions
(as seen in animal models) has been observed.
[0022] Local intra-tumoral injection is relatively straight-forward
in certain types of tumor such as some head and neck cancers and
melanomas, which accounts for the preponderance of clinical
investigations with oncolytic viruses in those indications. In
other indications the treatment of local disease requires
considerable resource and skill to place the agent into active
regions of tumor (discussed below). Brain tumors, deep-seated
visceral tumors and multiple active metastases pose considerable
barriers to successful intratumoral administration and attempts to
treat such tumors are often accompanied by significant safety and
technical issues, cost and resource requirements.
[0023] As reported in the Imlygic.RTM. data, the proposed use of
intra-tumoral injection of an augmented oncolytic HSV expressing a
cytokine (GM-CSF) to create a systemic therapy had limited success
in advanced or visceral disease--even in melanoma where multiple
intra-tumoral injections are relatively straight forward to carry
out.
[0024] The optimal dose and route of administration, particularly
for advanced, metastatic or visceral disease, remains a problem in
the art of oncolytic viruses.
[0025] Mace et al (Potential for efficacy of the oncolytic herpes
simplex virus 1716 in patients with oral squamous cell carcinoma.
Head Neck, 2008 August; 30(8):1045-51) utilized direct intratumoral
injection of herpes simplex virus HSV1716 in a Phase I study
designed to investigate safety and mechanism of action of HSV716
when injected up to 14 days prior to planned resection of the
tumor. On page 1050 Mace et al state: "In conclusion, intratumoral
injection of HSV1716 at a dose of 10.sup.5 pfu or 5.times.10.sup.5
pfu into oral SCC is safe and well tolerated but with little
biological activity. As has been seen with other oncolytic viruses,
the effective application is not as straightforward as laboratory
studies might have indicated. The principal problem areas involve
optimizing the dose, delivery and distribution of HSV1716 into a
dense heterogeneous SCC tumor cell matrix. Increasing our knowledge
of the interactions between HSV1716, the HNSCC tumor cell, and the
immune system will help to optimize antitumor efficacy".
[0026] As stated by Mace et al, the question remains--how to
optimise methods of treatment of cancer using oncolytic virus,
particularly oncolytic herpes simplex viruses. These questions and
challenges are also discussed by Russell et al (Oncolytic
Virotherapy. Nature Biotechnology Vol. 30 No. 7 Jul. 2012) and in
Seymour and Fisher (British Journal of Cancer (2016) 114,
357-361).
[0027] Systemic administration of herpes simplex virus has been
seen as problematic outside of model systems in the laboratory. The
main barriers to overcome being considered as (i) non-specificity
of HSV-1 binding and associated losses to non-target tissues; (ii)
the presence of circulating immune cells considered likely to
neutralise the HSV; and (iii) the inability to detect HSV in the
circulation immediately following IV infusion. These problems are
discussed in Russell et al., (Oncolytic virotherapy. Nature
Biotechnology Vol. 30 No. 7 Jul. 2012) which discusses the
challenges presented by administration of virus to the blood in
terms of neutralization of the virus by serum factors,
sequestration of virus by the mononuclear phagocytic system and
lack of extravasation. Seymour and Fisher (British Journal of
Cancer (2016) 114, 357-361) also discuss the challenges of
translating the promise of oncolytic viral therapy to success via
systemic delivery.
[0028] Ongoing clinical studies conducted by the Applicant have for
the first time investigated regional and systemic delivery of
HSV1716 in human patients. These studies in mesothelioma and in
advanced pediatric solid tumors have provided new data and insight
which have led to the invention described herein.
[0029] Clinical trial NCT00931931 is the first clinical study
designed to investigate intravenous infusion of an oncolytic herpes
simplex virus. This is a Phase 1 trial investigating the treatment
of non-central nervous system solid tumors.
[0030] The study was originally established to deliver HSV1716 by
direct injection into deep-seated tumors guided by imaging in an
interventional radiology suite. Patients eligible for the study had
disease refractory to all established treatments and presented with
large tumors and/or extensive metastatic disease. Despite the skill
of the interventional radiologists, intra-tumoral injection had
limited application to patients with such large tumor burden or
advanced metastatic disease. An amendment to the study was sought
in order to investigate, for the first time, whether systemic
delivery of HSV1716 was feasible, safe and had any biological
effect in this patient group.
[0031] Head and Neck Cancer
[0032] Squamous cell carcinoma of the head and neck (hereafter
"head and neck cancer" or "HNSCC") is the most common malignancy
(90%) of the upper aero-digestive tract. Head and neck cancer is
the sixth most common malignancy worldwide and results in approx.
350,000 deaths per year. The incidence of HNSCC has increased over
the past 10 years due to increasing prevalence of of human
papillomavirus (HPV). There are >45,000 estimated new US cases
reported each year (American Cancer Society, 2015) and >375,000
new cases worldwide (Globocan, 2012).
[0033] Intratumoral injection of the herpes simplex virus
talimogene laherparepvec (IMLYGIC.RTM.) was also investigated in a
Phase III study in head and neck cancer in combination with
cisplatin and radiation (Clinicaltrials.gov identifier NCT01161498)
but the study was terminated prior to completion and its results
have not been reported. The agent was subsequently approved for the
treatment of melanoma by the FDA based on a protocol of direct
intratumoral injection.
[0034] Intratumoral injection of herpes simplex viruses in clinical
studies for the treatment of head and neck cancer has therefore led
to disappointing results. The development of an appropriate
strategy for the use of oncolytic herpes simplex virus in order to
provide an effective treatment for Stage III or Stage IV head and
neck cancer is an outstanding problem in the field.
Immune Response
[0035] Cancer cells normally acquire antigenicity that can be
recognised by the adaptive immune system as non-self and lead to
generation of an immune response involving proliferation of
antigen-specific lymphocytes. Tumors are now established to co-opt
certain immune checkpoint pathways as a mechanism of immune
resistance against T-cells specific for tumor antigens (Drew M.
Pardoll., The blockade of immune checkpoints in cancer
immunotherapy. Nature Reviews Cancer Vol. 12 Apr. 2012
252-264).
[0036] Tumor infiltration with M2-phenotype macrophages and myeloid
derived suppressor cells (MDSC) promotes tumor progression whereas
infiltration of memory cytotoxic T cells and T helper 1 (Th1) T
lymphocytes are often associated with a good clinical outcome and
good response to immunotherapy. Therefore, one aim of
immunotherapies is to modify the context of immune, inflammatory
and angiogenic elements to favour a strong Th1 cytotoxic
microenvironment (Giraldo et al., The immune contexture of primary
and metastatic human tumors. Current Opinion in Immunology 2014,
27:8-15).
[0037] Haabeth et al (Inflammation driven by tumor-specific Th1
cells protects against B-cell cancer. Nature Communications. 2:240
15 Mar. 2011 [DOI:10.1038/ncomms1239]) also report that successful
cancer immunosurveillance mediated by tumor-specific CD4.sup.+ T
cells is consistently associated with elevated levels of both
proinflammatory (IL-1.alpha., IL-1.beta. and IL-6) and Th1
associated (interferon-.gamma. (IFN-.gamma.), IL-2 and IL-12)
cytokines. They describe cancer eradication as a collaboration
between tumor specific Th1 cells and tumor infiltrating, antigen
presenting macrophages in which Th1 cells induce secretion of
IL-1.beta. and IL-6 by macrophages and Th1 associated IFN-.gamma.
renders macrophages directly cytotoxic to cancer cells and causes
them to secrete the angiostatic chemokines CXCL9/MIG (monokine
induced by IFN-.gamma.) and CXCL10/IP-10 (IFN-.gamma. inducible
protein 10), and conclude that inflammation driven by tumor
specific Th1 cells may prevent rather than promote cancer. Haabeth
et al also report a panel of nine cytokines consistently associated
with successful cancer immunosurveillance, including
proinflammatory (IL-1.alpha., IL-1.beta. and IL-6) and Th1
associated (IL-2, IL-3, IL-12, IFN-.gamma., CXCL9 and CXCL10)
cytokines.
[0038] Whilst the pattern of cytokine production that characterises
the T helper type 1 (Th1) and type 2 (Th2) responses is reasonably
well understood, it is much less clear how the direction of
differentiation towards Th1 or Th2 is decided.
[0039] Herpes simplex virus type 1 has evolved several strategies
to evade the production and function of interferons (IFNs) and
cytokines generated by the innate immune system (which plays a role
in activation of the adaptive immune response). HSV counteracts the
production of IFN, diminishes IFN-signalling, and blocks the
actions of PKR and activation of the 2'-5' A system through several
viral products, including ICP0, ICP27, ICP34.5 and vhs (Melchjorsen
et al., Activation and Evasion of Innate Antiviral Immunity by
Herpes Simplex Virus. Viruses 2009, 1, 737-759;
doi:10.3390/v1030737; L. Aurelian., Herpes Simplex Virus Type 2
Vaccines: New Ground for Optimism? Clinical and Diagnostic
Laboratory Immunology, May 2004, p. 437-445).
[0040] Young J Kim (Subverting the adaptive immune resistance
mechanism to improve clinical responses to immune checkpoint
blockade therapy. Oncolmmunology 3:12 e954868; December 2014)
describes a mechanistic and clinical rationale for combining
IFN-.gamma. Th1 inducing cancer vaccines with the blockade of the
immune checkpoint protein Programmed cell death 1 (PD-1, also
called CD279), and indicates that strategies to increase
tumor-infiltrating cytotoxic T lymphocyte anticancer activities
with immune checkpoint inhibitors may convert anti-PD-1 blockade
non-responders to responders, thereby circumventing immune
evasion.
[0041] Sagiv-Barfi et al reported that ibrutinib, an inhibitor of
ITK, an essential enzyme in Th2 T cells can shift the balance
between Th1 and Th2 T cells towards Th1 and thereby enhance
anti-tumor responses. They describe the combination of anti-PD-L1
antibody and ibrutinib to suppress tumor growth in mouse models of
lymphoma that are intrinsically insensitive to ibrutinib, as well
as in models of breast and colon cancer (Sagiv-Barfi et al.,
Therapeutic antitumor immunity by checkpoint blockade is enhanced
by ibrutinib, an inhibitor of both BTK and ITK. PNAS E966-972
Published online Feb. 17, 2015).
[0042] Accordingly, there are advantages for treatment of tumors
associated with promoting a Th1 response in the tumor
microenvironment, and further ways of establishing such a
treatment-favourable environment are required.
[0043] WO2014/036412 describes the treatment of stage IIIb to stage
IV melanoma by a method comprising administering to a patient with
stages IIIb to IV melanoma an effective amount of an immune
checkpoint inhibitor and a herpes simplex virus, wherein the herpes
simplex virus lacks functional ICP34.5 genes, lacks a functional
ICP47 gene and comprises a gene encoding human GM-CSF.
SUMMARY OF THE INVENTION
[0044] The inventors have shown that following infusion of
oncolytic herpes simplex virus to the blood of a human subject
having a cancer the virus is undetectable in the blood after about
24 hours, which is consistent with proposed mechanisms of viral
neutralization and sequestration discussed above. However, after
about 72 hours in some subjects viral DNA is detectable again in
the blood. This observation and accompanying tumor imaging
observations indicate that the virus has reached cancer cells in
the subject, has infected those cells and replicated. Lysis of
cancer cells is consistent with release of viral particles and
fragments, including viral DNA, into the blood which may then be
detected.
[0045] These remarkable observations provide the first indication
that cancer cells can be treated in human subjects at a location
distant to the site of administration by a mechanism that involves
the virus reaching those cancer cells, e.g. in which the virus
infects, replicates in and lyses the cancer cells, rather than
relying on induction of an immune response to treat cancer cells
not directly injected with virus.
[0046] These findings open the door to treatment of cancers which
are not suitable for direct injection with virus, such as cancers
occurring in the internal organs, cancers occurring at multiple
sites, and metastatic cancers.
[0047] The treatment of cancer in human subjects may therefore be
effected by infusion of a therapeutically effective amount of
oncolytic herpes simplex virus to the subject's blood. Methods of
treatment may further involve taking of blood or tumor tissue
samples to determine the presence of herpes simplex virus and
provide an indicator that virus has reached cells of the
cancer.
[0048] In one aspect of the present invention an oncolytic herpes
simplex virus for use in a method of treating cancer in a human
subject is provided, the method comprising administering to the
human subject at least one dose of oncolytic herpes simplex virus
by infusion to the blood, wherein the oncolytic herpes simplex
virus reaches cells of the cancer in which it replicates.
[0049] In another aspect of the present invention the use of an
oncolytic herpes simplex virus in the manufacture of a medicament
for use in a method of treating cancer in a human subject is
provided, the method comprising administering to the human subject
at least one dose of oncolytic herpes simplex virus by infusion to
the blood, wherein the oncolytic herpes simplex virus reaches cells
of the cancer in which it replicates.
[0050] In another aspect of the present invention a method of
treating cancer in a human subject in need of treatment is
provided, the method comprising administering to the human subject
at least one dose of oncolytic herpes simplex virus by infusion to
the blood, wherein the oncolytic herpes simplex virus reaches cells
of the cancer in which it replicates.
[0051] In some embodiments, upon infusion oncolytic herpes simplex
is absorbed by cells or is neutralised, but is able to reach cancer
cells where it infects, replicates and lyses cancer cells, lysis of
cancer cells releasing viral DNA which is detectable in the blood.
In some embodiments, oncolytic herpes simplex virus DNA is not
detectable in a sample of the subject's blood within about 24 hours
of infusion, but is detectable in a sample of the subject's blood
taken after one of about 72 hours, about 96 hours, about 120 hours,
about 144 hours, about 168 hours, about 2 weeks, about 3 weeks or
about 4 weeks. In some embodiments, the method comprises
determining whether oncolytic herpes simplex virus DNA is present
in a sample of the subject's blood. The method may comprise taking
a blood sample from the subject, and determining whether herpes
simplex virus DNA is present in the blood sample.
[0052] In some embodiments, the method comprises determining
whether oncolytic herpes simplex virus is present in a sample of
the subject's tumor tissue. The method may comprise taking a sample
of tumor tissue from the subject and determining whether oncolytic
herpes simplex virus is present in a sample of the subject's tumor
tissue. The oncolytic herpes simplex virus may be a mutant of HSV-1
strain 17 or is HSV1716. The cancer may be a solid tumor, a
recurrent or metastatic solid tumor, or a non-CNS solid tumor.
[0053] The dose of oncolytic herpes simplex virus administered may
be at least 1.times.10.sup.6 iu.
[0054] In some embodiments, a dose of oncolytic herpes simplex
virus is administered over a period of 3 hours or less.
[0055] In some embodiments, the administered oncolytic herpes
simplex virus is formulated as about 0.5 ml to about 5 ml of a
suspension of virus in about 200 ml to about 300 ml of lactated
Ringer's solution.
[0056] In some embodiments, the method comprises administering to
the human subject at least one treatment cycle of oncolytic herpes
simplex virus, wherein a treatment cycle comprises, or consists, of
at least two doses of oncolytic herpes simplex virus, each dose
administered by infusion to the blood wherein the second and
subsequent doses are each administered within about 17 days of the
preceding dose, each dose of oncolytic herpes simplex virus being
in the range about 1.times.10.sup.6 iu to about 1.times.10.sup.8
iu. In some embodiments, one dose of oncolytic herpes simplex virus
is administered per week. In some other embodiments, two doses of
oncolytic herpes simplex virus are administered per week. Each dose
of oncolytic herpes simplex virus may be in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu. The treatment
cycle may further comprise, or consist of, administration of a
therapeutically effective amount of an immune checkpoint inhibitor
which may be selected from the group consisting of an inhibitor of
PD-1, PD-L1, CTLA4, TIM-3 or LAG-3. In some embodiments, the
subject may receive two or more treatment cycles.
[0057] In some embodiments, the method comprises determining the
presence of a Th1 response in the subject. In some embodiments, the
level of IL-2 and/or IL-12 and/or IFN-.gamma. in the subject's
blood may be upregulated for more than 7 days after one or more
treatment cycles.
[0058] In another aspect of the present invention an oncolytic
herpes simplex virus for use in a method of treating cancer in a
human subject is provided, the method comprising administering to
the human subject at least one treatment cycle of oncolytic herpes
simplex virus, wherein a treatment cycle comprises, or consists, of
at least two doses of oncolytic herpes simplex virus, each dose
administered by infusion to the blood wherein the second and
subsequent doses are each administered within about 17 days of the
preceding dose, each dose of oncolytic herpes simplex virus being
in the range about 1.times.10.sup.6 iu to about 1.times.10.sup.8
iu.
[0059] In another aspect of the present invention the use of an
oncolytic herpes simplex in the manufacture of a medicament for us
in a method of treating cancer in a human subject is provided, the
method comprising administering to the human subject at least one
treatment cycle of oncolytic herpes simplex virus, wherein a
treatment cycle comprises, or consists, of at least two doses of
oncolytic herpes simplex virus, each dose administered by infusion
to the blood wherein the second and subsequent doses are each
administered within about 17 days of the preceding dose, each dose
of oncolytic herpes simplex virus being in the range about
1.times.10.sup.6 iu to about 1.times.10.sup.8 iu.
[0060] In another aspect of the present invention a method of
treating cancer in a human subject in need of treatment is
provided, the method comprising administering to the human subject
at least one treatment cycle of oncolytic herpes simplex virus,
wherein a treatment cycle comprises, or consists, of at least two
doses of oncolytic herpes simplex virus, each dose administered by
infusion to the blood wherein the second and subsequent doses are
each administered within about 17 days of the preceding dose, each
dose of oncolytic herpes simplex virus being in the range about
1.times.10.sup.6 iu to about 1.times.10.sup.8 iu.
[0061] A treatment cycle may comprise 2 doses, more than 2 doses,
up to 4 doses, 4 doses, up to 6 doses, 6 doses, up to 8 doses, or 8
doses, up to 10 doses, or 10 doses, of oncolytic herpes simplex
virus.
[0062] The second and subsequent doses may be administered within
about 14 (e.g. 14.+-.1, 14.+-.2, 14.+-.3, or 14.+-.4) or about 7
(e.g. 7.+-.1 or 7.+-.2) days of the preceding dose. In some
embodiments one dose of oncolytic herpes simplex virus may be
administered per week. Administration of each weekly dose of
oncolytic herpes simplex virus may be separated by 7.+-.1 or 7.+-.2
days. In some embodiments two doses of oncolytic herpes simplex
virus may be administered per week. Administration of each twice
weekly dose of oncolytic herpes simplex virus may be separated by
4.+-.1 or 4.+-.2 days.
[0063] In some embodiments each dose of oncolytic herpes simplex
virus may be in the range about 1.times.10.sup.7 iu to about
1.times.10.sup.8 iu.
[0064] The subject may receive administration of one or a plurality
(preferably a plurality) of doses of herpes simplex virus. Each
dose of herpes simplex virus is preferably in the range
1.times.10.sup.6 iu to 1.times.10.sup.8 iu, and may be
2.times.10.sup.6 iu or greater than 2.times.10.sup.6 iu. Doses may
be in a range selected from the group consisting of:
2.times.10.sup.6 to 9.times.10.sup.6 iu, 2.times.10.sup.6 to
5.times.10.sup.6 iu, 5.times.10.sup.6 to 9.times.10.sup.6 iu,
2.times.10.sup.6 to 1.times.10.sup.7 iu, 2.times.10.sup.6 to
5.times.10.sup.7 iu, 2.times.10.sup.6 to 1.times.10.sup.8 iu,
2.times.10.sup.6 to 5.times.10.sup.8 iu, 2.times.10.sup.6 to
1.times.10.sup.9 iu, 5.times.10.sup.6 to 1.times.10.sup.7 iu,
5.times.10.sup.6 to 5.times.10.sup.7 iu, 5.times.10.sup.6 to
1.times.10.sup.8 iu, 5.times.10.sup.6 to 5.times.10.sup.8 iu,
5.times.10.sup.6 to 1.times.10.sup.9 iu, 5.times.10.sup.6 to
5.times.10.sup.9 iu, 1.times.10.sup.7 to 9.times.10.sup.7 iu,
1.times.10.sup.7 to 5.times.10.sup.7 iu, 1.times.10.sup.8 to
9.times.10.sup.8 iu, 1.times.10.sup.8 to 5.times.10.sup.8 iu. In
some embodiments suitable doses may be in the range
2.times.10.sup.6 to 9.times.10.sup.6 iu, 1.times.10.sup.7 to
9.times.10.sup.7 iu, or 1.times.10.sup.8 to 9.times.10.sup.8 iu. In
some embodiments suitable doses may be about 1.times.10.sup.7 iu or
about 1.times.10.sup.8 iu. Dosage figures may optionally be +/-half
a log value.
[0065] In some embodiments the treatment cycle further comprises,
or consists, of administration of a therapeutically effective
amount of an immune checkpoint inhibitor.
[0066] In some embodiments the treatment cycle of oncolytic herpes
simplex virus, comprises, or consists, of: [0067] (i) four doses of
oncolytic herpes simplex virus over a period of about four weeks,
one dose administered per week by infusion to the blood, each dose
of oncolytic herpes simplex virus being in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu; or [0068] (ii)
four doses of oncolytic herpes simplex virus over a period of about
two weeks, two doses administered per week by infusion to the
blood, each dose of oncolytic herpes simplex virus being in the
range about 1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0069] As such, in one aspect of the present invention an oncolytic
herpes simplex virus for use in a method of treating cancer in a
human subject is provided, the method comprising administering to
the human subject at least one treatment cycle of oncolytic herpes
simplex virus, wherein a treatment cycle comprises, or consists,
of: [0070] (i) four doses of oncolytic herpes simplex virus over a
period of about four weeks, one dose administered per week by
infusion to the blood, each dose of oncolytic herpes simplex virus
being in the range about 1.times.10.sup.7 iu to about
1.times.10.sup.8 iu; or [0071] (ii) four doses of oncolytic herpes
simplex virus over a period of about two weeks, two doses
administered per week by infusion to the blood, each dose of
oncolytic herpes simplex virus being in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0072] In another aspect of the present invention the use of an
oncolytic herpes simplex in the manufacture of a medicament for us
in a method of treating cancer in a human subject is provided, the
method comprising administering to the human subject at least one
treatment cycle of oncolytic herpes simplex virus, wherein a
treatment cycle comprises, or consists, of: [0073] (i) four doses
of oncolytic herpes simplex virus over a period of about four
weeks, one dose administered per week by infusion to the blood,
each dose of oncolytic herpes simplex virus being in the range
about 1.times.10.sup.7 iu to about 1.times.10.sup.8 iu; or [0074]
(ii) four doses of oncolytic herpes simplex virus over a period of
about two weeks, two doses administered per week by infusion to the
blood, each dose of oncolytic herpes simplex virus being in the
range about 1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0075] In another aspect of the present invention a method of
treating cancer in a human subject in need of treatment is
provided, the method comprising administering to the human subject
at least one treatment cycle of oncolytic herpes simplex virus,
wherein a treatment cycle comprises, or consists, of: [0076] (i)
four doses of oncolytic herpes simplex virus over a period of about
four weeks, one dose administered per week by infusion to the
blood, each dose of oncolytic herpes simplex virus being in the
range about 1.times.10.sup.7 iu to about 1.times.10.sup.8 iu; or
[0077] (ii) four doses of oncolytic herpes simplex virus over a
period of about two weeks, two doses administered per week by
infusion to the blood, each dose of oncolytic herpes simplex virus
being in the range about 1.times.10.sup.7 iu to about
1.times.10.sup.8 iu.
[0078] In another aspect of the present invention an oncolytic
herpes simplex virus for use in a method of treating cancer in a
human subject is provided, the method comprising administering to
the human subject an oncolytic herpes simplex virus and a
therapeutically effective amount of an immune checkpoint inhibitor,
wherein the method comprises at least one treatment cycle of
oncolytic herpes simplex virus comprising, or consisting, of:
[0079] (i) four doses of oncolytic herpes simplex virus over a
period of about four weeks, one dose administered per week by
infusion to the blood, each dose of oncolytic herpes simplex virus
being in the range about 1.times.10.sup.7 iu to about
1.times.10.sup.8 iu; or [0080] (ii) four doses of oncolytic herpes
simplex virus over a period of about two weeks, two doses
administered per week by infusion to the blood, each dose of
oncolytic herpes simplex virus being in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0081] In another aspect of the present invention the use of an
oncolytic herpes simplex in the manufacture of a medicament for us
in a method of treating cancer in a human subject is provided, the
method comprising administering to the human subject an oncolytic
herpes simplex virus and a therapeutically effective amount of an
immune checkpoint inhibitor, wherein the method comprises at least
one treatment cycle of oncolytic herpes simplex virus comprising,
or consisting, of: [0082] (i) four doses of oncolytic herpes
simplex virus over a period of about four weeks, one dose
administered per week by infusion to the blood, each dose of
oncolytic herpes simplex virus being in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu; or [0083] (ii)
four doses of oncolytic herpes simplex virus over a period of about
two weeks, two doses administered per week by infusion to the
blood, each dose of oncolytic herpes simplex virus being in the
range about 1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0084] In another aspect of the present invention a method of
treating cancer in a human subject in need of treatment is
provided, the method comprising administering to the human subject
an oncolytic herpes simplex virus and a therapeutically effective
amount of an immune checkpoint inhibitor, wherein the method
comprises at least one treatment cycle of oncolytic herpes simplex
virus comprising, or consisting, of: [0085] (i) four doses of
oncolytic herpes simplex virus over a period of about four weeks,
one dose administered per week by infusion to the blood, each dose
of oncolytic herpes simplex virus being in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu; or [0086] (ii)
four doses of oncolytic herpes simplex virus over a period of about
two weeks, two doses administered per week by infusion to the
blood, each dose of oncolytic herpes simplex virus being in the
range about 1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0087] In some embodiments administration of each weekly dose of
oncolytic herpes simplex virus in is separated by 7.+-.1 or 7.+-.2
days. In some embodiments administration of each twice weekly dose
of oncolytic herpes simplex virus is separated by 4.+-.1 or 4.+-.2
days.
[0088] In some embodiments the method comprises administering an
immune checkpoint inhibitor according to at least one treatment
cycle of immune checkpoint inhibitor, wherein the periods of time
of the oncolytic herpes simplex treatment cycle and immune
checkpoint inhibitor treatment cycle overlap. In some embodiments
the method comprises at least one treatment cycle of immune
checkpoint inhibitor comprising, or consisting, of 1 or 2 doses of
immune checkpoint inhibitor administered within a period of about 3
or 4 weeks. In some embodiments the method comprises at least one
treatment cycle of immune checkpoint inhibitor comprising, or
consisting, of a period of about 3 weeks in which 1 dose of immune
checkpoint inhibitor is administered. In some embodiments the
method comprises at least one treatment cycle of immune checkpoint
inhibitor comprising, or consisting, of 1 dose of immune checkpoint
inhibitor administered about every 3 weeks. In some embodiments the
method comprises at least two treatment cycles of oncolytic herpes
simplex virus according to (i) and at least two treatment cycles of
immune checkpoint inhibitor each comprising, or consisting, of a
period of about 3 weeks in which 1 dose of immune checkpoint
inhibitor is administered, wherein the periods of time of the first
treatment cycle of oncolytic herpes simplex virus and first
treatment cycle of immune checkpoint inhibitor overlap. In some
embodiments the method comprises at least two treatment cycles of
oncolytic herpes simplex virus according to (ii) and a treatment
cycle of immune checkpoint inhibitor comprising, or consisting, of
a period of about 3 weeks in which 1 dose of immune checkpoint
inhibitor is administered, wherein the periods of time of the
treatment cycles of oncolytic herpes simplex virus and treatment
cycle of immune checkpoint inhibitor overlap.
[0089] The immune checkpoint inhibitor may be an inhibitor of at
least one of PD-1, PD-L1, CTLA4, TIM-3 or LAG-3. The immune
checkpoint inhibitor may be selected from an anti-PD-1 antibody,
anti-PD-L1 antibody, anti-CTLA4 antibody, anti-TIM-3 antibody or
anti-LAG-3 antibody. The immune checkpoint inhibitor may be
selected from the group consisting of pembrolizumab, nivolumab or
ipilumab.
[0090] In another aspect of the present invention an oncolytic
herpes simplex virus for use in a method of treating cancer in a
human subject is provided, the method comprising administering:
[0091] a herpes simplex virus by intravenous infusion at a dose of
greater than 2.times.10.sup.6 iu at day 1 of week 1 followed by a
dose of greater than 2.times.10.sup.6 iu at day 1 of week 2, and
every week thereafter until surgery is scheduled, toxicity or
disease progression, a predetermined maximum number of doses is
reached, a fixed number of doses is reached, complete response,
disease progression, or intolerance of the treatment, wherein the
herpes simplex virus lacks functional ICP34.5 genes; and an immune
checkpoint inhibitor by intravenous infusion every 3 weeks for at
least 3 infusions beginning with or after the second or third dose
of the herpes simplex virus.
[0092] In another aspect of the present invention the use of an
oncolytic herpes simplex in the manufacture of a medicament for us
in a method of treating cancer in a human subject is provided, the
method comprising administering: [0093] a herpes simplex virus by
intravenous infusion at a dose of greater than 2.times.10.sup.6 iu
at day 1 of week 1 followed by a dose of greater than
2.times.10.sup.6 iu at day 1 of week 2, and every week thereafter
until surgery is scheduled, toxicity or disease progression, a
predetermined maximum number of doses is reached, a fixed number of
doses is reached, complete response, disease progression, or
intolerance of the treatment, wherein the herpes simplex virus
lacks functional ICP34.5 genes; and [0094] an immune checkpoint
inhibitor by intravenous infusion every 3 weeks for at least 3
infusions beginning with or after the second or third dose of the
herpes simplex virus.
[0095] In another aspect of the present invention a method for the
treatment of cancer is provided, the method comprising
administering: [0096] a herpes simplex virus by intravenous
infusion at a dose of greater than 2.times.10.sup.6 iu at day 1 of
week 1 followed by a dose of greater than 2.times.10.sup.6 iu at
day 1 of week 2, and every week thereafter until surgery is
scheduled, toxicity or disease progression, a predetermined maximum
number of doses is reached, a fixed number of doses is reached,
complete response, disease progression, or intolerance of the
treatment, wherein the herpes simplex virus lacks functional
ICP34.5 genes; and [0097] an immune checkpoint inhibitor by
intravenous infusion every 3 weeks for at least 3 infusions
beginning with or after the second or third dose of the herpes
simplex virus.
[0098] In some embodiments a dose of oncolytic herpes simplex virus
is administered over a period of 3 hours or less.
[0099] In some embodiments the administered oncolytic herpes
simplex virus is formulated as about 0.5 ml to about 5 ml of a
suspension of virus in about 200 ml to about 300 ml of lactated
Ringer's solution. In some embodiments the administered oncolytic
herpes simplex virus is formulated as about 1.0 ml of a suspension
of virus in about 250 ml of lactated Ringer's solution.
[0100] The subject may receive two or more treatment cycles of
oncolytic herpes simplex virus, which may be consecutive or each
treatment cycle may be separated by a break from treatment. A break
from treatment may be about 1, 2, 3, or 4 weeks.
[0101] The first 1, 2 or 3 treatment cycles of oncolytic herpes
simplex virus may comprise administration of a dose of oncolytic
herpes simplex virus that is lower than the dose administered in
later treatment cycles.
[0102] In some embodiments the method may further comprise
determining the presence of a Th1 response in the subject. The
method may comprise measuring the level of IFN.gamma., IL-2 and/or
IL-12 in a sample obtained from a subject. The level of IL-2 and/or
IL-12 and/or IFN-.gamma. in the subject's blood may be upregulated
for more than 7 days after one or more treatment cycles.
[0103] In some embodiments all copies of the ICP34.5 gene in the
genome of the herpes simplex virus are modified such that the
ICP34.5 gene is incapable of expressing a functional ICP34.5 gene
product. As such the herpes simplex virus may be an ICP34.5 null
mutant.
[0104] In some embodiments one or both of the ICP34.5 genes in the
genome of the herpes simplex virus are modified such that the
ICP34.5 gene is incapable of expressing a functional ICP34.5 gene
product.
[0105] In some embodiments the herpes simplex virus is a mutant of
HSV-1 strain 17. In preferred embodiments the herpes simplex virus
is HSV1716 (ECACC Accession No. V92012803). HSV1716 is also called
SEPREHVIR.RTM.. In some embodiments the herpes simplex virus is a
mutant of HSV-1 strain 17 mutant 1716.
[0106] The cancer to be treated may be a solid tumor, a recurrent
or metastatic tumor, a recurrent or metastatic solid tumor, a
non-CNS tumor, or a non-CNS solid tumor. In some embodiments the
cancer is not a melanoma.
[0107] In some embodiments human subject may be a child.
[0108] In some embodiments the oncolytic herpes simplex virus does
not encode (or is not further modified to contain nucleic acid
encoding) a cytokine or chemokine, an interleukin, an interferon, a
tumor necrosis factor, a colony stimulating factor, an immune
modulator, a member of the CC family, a member of the CXC family or
a member of the CXC family. In some embodiments the oncolytic
herpes simplex virus does not express GMCSF. In some embodiments
the oncolytic herpes simplex virus encodes a functional ICP47
and/or ICP6 gene.
[0109] In another aspect of the present invention a kit comprising
a herpes simplex virus lacking functional ICP34.5 genes, and a
package insert or label with directions to treat cancer by using a
combination of the herpes simplex virus and an immune checkpoint
inhibitor is provided.
[0110] The directions may comprise instructions to administer to a
human subject with cancer at least one treatment cycle of oncolytic
herpes simplex virus, wherein a treatment cycle comprises, or
consists, of at least two doses of oncolytic herpes simplex virus,
each dose administered by infusion to the blood wherein the second
and subsequent doses are each administered within about 17 days of
the preceding dose, each dose of oncolytic herpes simplex virus
being in the range about 1.times.10.sup.6 iu to about
1.times.10.sup.8 iu.
[0111] The directions may comprise instructions to administer to a
human subject with cancer at least one treatment cycle of oncolytic
herpes simplex virus, wherein a treatment cycle comprises, or
consists, of: [0112] (i) four doses of oncolytic herpes simplex
virus over a period of about four weeks, one dose administered per
week by infusion to the blood, each dose of oncolytic herpes
simplex virus being in the range about 1.times.10.sup.7 iu to about
1.times.10.sup.8 iu; or [0113] (ii) four doses of oncolytic herpes
simplex virus over a period of about two weeks, two doses
administered per week by infusion to the blood, each dose of
oncolytic herpes simplex virus being in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0114] The directions may comprise instructions to administer to a
human subject with cancer an oncolytic herpes simplex virus and a
therapeutically effective amount of an immune checkpoint inhibitor,
wherein the instructions for administration of the oncolytic herpes
simplex virus comprise instructions to administer at least one
treatment cycle of oncolytic herpes simplex virus comprising, or
consisting, of: [0115] (i) four doses of oncolytic herpes simplex
virus over a period of about four weeks, one dose administered per
week by infusion to the blood, each dose of oncolytic herpes
simplex virus being in the range about 1.times.10.sup.7 iu to about
1.times.10.sup.8 iu; or [0116] (ii) four doses of oncolytic herpes
simplex virus over a period of about two weeks, two doses
administered per week by infusion to the blood, each dose of
oncolytic herpes simplex virus being in the range about
1.times.10.sup.7 iu to about 1.times.10.sup.8 iu.
[0117] The directions may comprise instructions to administer to a
patient with cancer a herpes simplex virus administered by
intravenous infusion at a dose of greater than 2.times.10.sup.6 iu
at day 1 of week 1 followed by a dose of greater than
2.times.10.sup.6 iu at day 1 of week 2, and every week thereafter
until surgery is scheduled, toxicity or disease progression, a
predetermined maximum number of doses is reached, a fixed number of
doses is reached, complete response, disease progression, or
intolerance of the treatment; and an immune checkpoint inhibitor
administered intravenously every 3 weeks for at least 3 infusions
beginning with or after the second or third dose of the herpes
simplex virus.
[0118] A method of manufacturing the kit is also provided.
[0119] In another aspect of the present invention a method of
promoting a combination treatment comprising a herpes simplex virus
lacking functional ICP34.5 genes and an immune checkpoint
inhibitor, for the treatment of a human patient with cancer, is
provided.
[0120] In some embodiments the promotion is by a package insert,
wherein the package insert provides instructions to receive cancer
treatment with a herpes simplex virus in combination with an immune
checkpoint inhibitor. In some embodiments the promotion is by a
package insert accompanying a formulation comprising the herpes
simplex virus. In some embodiments the promotion is by written
communication to a physician or health care provider. In some
embodiments the promotion is by oral communication to a physician
or health care provider. In some embodiments the promotion is
followed by the treatment of the patient with the herpes simplex
virus.
[0121] In another aspect of the present invention a method of
instructing a human patient with cancer by providing instructions
to receive a combination treatment with a herpes simplex virus
lacking functional ICP34.5 genes and an immune checkpoint inhibitor
to extend survival of the patient, is provided.
[0122] Doses of herpes simplex virus are preferably administered by
intravenous infusion, which may take place over a period of several
hours, e.g. about 30 minutes to about 3 or about 4 hours. For
example, Infusion of a dose of herpes simplex virus may take place
over about 1 hour, about 2 hours or about 3 hours.
[0123] A subject will commonly receive a plurality of doses of
herpes simplex virus as part of a course of treatment, preferably 3
or more doses. The doses may be administered in accordance with a
dosing regime. For example, each dose of herpes simplex virus may
be administered within 1 to 7, 1 to 14, or 1 to 21 days of the
preceding dose.
[0124] Doses of herpes simplex virus may be administered at regular
intervals, e.g. every 7 days, every 14 days, every 21 days, or
every 28 days (+1-1, 2 or 3 days). The number of doses of herpes
simplex virus administered in a course of treatment may be any of
1, 2, 3, 4, 5, 6, 7, 8, 9, 10 doses or more, preferably at least 3
or at least 4 doses, or up to 8 doses.
[0125] A dosing regime for a herpes simplex virus may be designed
to continue dosing until: surgery is scheduled, toxicity or disease
progression, a predetermined maximum number of doses is reached,
e.g. 3, 4, 5, 6, 7, 8, 9 or 10 doses, or a fixed number of doses is
reached, e.g. 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 doses, complete
response, disease progression, or intolerance of the treatment.
[0126] The immune checkpoint inhibitor may be administered
intravenously. The subject may receive a single intravenous
administration of a combined preparation of herpes simplex virus
and immune checkpoint inhibitor, or may receive separate
intravenous administrations of herpes simplex virus and immune
checkpoint inhibitor. Herpes simplex virus and immune checkpoint
inhibitor may be administered on the same day, e.g. during the same
hospital visit, or on separate days.
[0127] A subject may receive a plurality of doses of immune
checkpoint inhibitor during the course of treatment. The doses may
be administered in accordance with a dosing regime. For example,
each dose of immune checkpoint inhibitor may be administered within
1 to 21 or 1 to 28 days of the preceding dose. Doses of immune
checkpoint inhibitor may be administered at regular intervals, e.g.
every 7 days, every 14 days, every 21 days, or every 28 days (+/-1,
2 or 3 days).
[0128] Suitable doses of immune checkpoint inhibitor will vary
depending on the immune checkpoint inhibitor selected and the
prescribing information of the medical practitioner. By way of
example, suitable doses may be one of 100 mg, 200 mg, 300 mg or 400
mg, or 1 mg/kg, 2 mg/kg, 3 mg/kg or 4 mg/kg.
[0129] Doses of immune checkpoint inhibitor may be administered by
intravenous infusion, which may take place over a period of up to
several hours, e.g. about 30 minutes to about 3 or about 4 hours.
For example, Infusion of a dose of immune checkpoint inhibitor may
take place over about 1 hour, about 2 hours or about 3 hours.
[0130] By way of example, pembrolizumab may be administered by
intravenous infusion once every 3 weeks, optionally at a dose of
about 200 mg. In another example, pembrolizumab may be administered
by intravenous infusion once every week, optionally at a dose of
about 200 mg, and optionally in combination with chemotherapy, e.g.
in the form of platinum and/or 5-fluoruracil.
[0131] A combined dosing regime for a herpes simplex virus and
immune checkpoint inhibitor may be designed to continue dosing
until: toxicity or disease progression, a predetermined maximum
number of doses of one or both agents is reached, e.g. 3, 4, 5, 6,
7, 8, 9 or 10 doses, a fixed number of doses of one or both agents
is reached, e.g. 3, 4, 5, 6, 7, 8, 9, or 10 doses, complete
response, disease progression, or intolerance of the treatment.
[0132] In some embodiments herpes simplex virus may be administered
more regularly than the immune checkpoint inhibitor. Administration
of herpes simplex virus may optionally precede, i.e. commence
before, administration of the immune checkpoint inhibitor. For
example, herpes simplex virus may be administered once weekly,
whereas the immune checkpoint inhibitor may be administered once
every 2, 3 or 4 weeks, optionally commencing in the first, second
or third week of treatment. The dosing schedule may be designed
such that administration of herpes simplex virus is on the same day
as administration of the immune checkpoint inhibitor.
[0133] The immune checkpoint inhibitor may be an inhibitor of at
least one of PD-1, PD-L1, CTLA4, TIM-3 or LAG-3. The immune
checkpoint inhibitor may be an anti-PD-1 antibody, anti-PD-L1
antibody, anti-CTLA4 antibody, anti-TIM-3 antibody or anti-LAG-3
antibody. The immune checkpoint inhibitor may be one of
pembrolizumab, nivolumab or ipilimumab.
[0134] In one aspect of the present invention a pharmaceutical
composition comprising a herpes simplex virus and an immune
checkpoint inhibitor is provided. The herpes simplex virus may be a
mutant of HSV-1 strain 17. In preferred embodiments the herpes
simplex virus is HSV1716.
[0135] In one aspect of the present invention a kit is provided,
the kit comprising a predetermined amount of herpes simplex virus
and a predetermined amount of an immune checkpoint inhibitor. The
herpes simplex virus may be a mutant of HSV-1 strain 17. In
preferred embodiments the herpes simplex virus is HSV1716. The kit
may be provided together with instructions for the administration
of the herpes simplex virus, and immune checkpoint inhibitor
sequentially or simultaneously in order to provide a treatment for
cancer.
[0136] In one aspect of the present invention products are provided
containing therapeutically effective amounts of: [0137] (i)
HSV1716, and [0138] (ii) an immune checkpoint inhibitor for
simultaneous or sequential use in a method of medical treatment,
preferably treatment of cancer. The products may be
pharmaceutically acceptable formulations and may optionally be
formulated as a combined preparation for coadministration.
[0139] Optionally, in some embodiments the herpes simplex virus
does not express GMCSF. Optionally, in some embodiments the herpes
simplex virus encodes a functional ICP47 gene. Optionally, in some
embodiments the herpes simplex virus is not a herpes simplex virus
that lacks functional ICP34.5 genes and lacks a functional ICP47
gene and comprises a gene encoding human GM-CSF.
[0140] Optionally, in some embodiments the cancer is not a
melanoma. Optionally, in some embodiments the cancer is not a
primary melanoma. Optionally, in some embodiments the cancer is not
a metastatic (secondary) melanoma. Optionally, in some embodiments
the cancer is not stage IIIb to stage IV melanoma. Optionally, in
some embodiments the cancer is not a head and neck cancer.
DESCRIPTION
[0141] Without being bound to any particular theory of invention,
the inventors have identified that herpes simplex virus oncolytic
immunotherapy works through a multi-modal approach.
Systemic Administration
[0142] Contrary to the disclosure in Mace et al (supra) and the
experience with IMLYGIC.RTM., the inventors have realised that
simply increasing the dosage of herpes simplex virus by
intra-tumoral injection is unlikely to generate a satisfactory
response for a number of reasons: [0143] (i) ever higher doses of
oncolytic treatment risk triggering an uncontrollable cytokine
response raising safety concerns; [0144] (ii) under high
multiplicity of infection, the tumor cells at the site of injection
are unlikely to support virus replication with evidence that the
cells may adopt an apoptotic pathway and commence shut down to
prevent viral spread thus undermining the basis of oncolytic
therapy; [0145] (iii) tumor heterogeneity limits the spread of an
oncolytic virus from the point of injection; [0146] (iv) injection
into accessible tumors--which may even be amenable to surgical
treatment--is unlikely to be effective for disseminated or invasive
disease, e.g. as commonly found in Stage III and IV head and neck
cancer.
[0147] The barriers to effective systemic administration of HSV in
human subjects are common general knowledge in the art (e.g. as
discussed in Russell et al., (Oncolytic virotherapy. Nature
Biotechnology Vol. 30 No. 7 Jul. 2012) and Seymour and Fisher
(British Journal of Cancer (2016) 114, 357-361).
[0148] Despite the teaching in the art, the inventors pursued the
systemic administration of herpes simplex virus, i.e. to the blood,
as a therapeutic strategy in a Phase I trial.
[0149] Surprisingly, even at the relatively low doses used in the
Phase I trial, evidence of virus targeting and replication in the
subjects has emerged and is reported below.
[0150] The inventors have therefore identified systemic
administration of herpes simplex virus, i.e. to the blood, as a
therapeutic strategy. Optionally the systemic administration of
herpes simplex virus may be coupled with monitoring the subject to
determine whether the virus has successfully reached the tumor and
replicated at the site of the tumor. Such monitoring is possible by
taking a sample of blood or tumor tissue and analysing the sample
for the presence of herpes simplex virus, e.g. HSV DNA or envelope
proteins or HSV antigens. Based on the determination the subject
may be selected to receive further treatment with the herpes
simplex virus, optionally in combination with treatment with an
immune checkpoint inhibitor.
[0151] In the context of cancer indicated for surgery, optionally
head and neck cancer, herpes simplex virus may be administered to
the blood prior to surgery and removal of tumor tissue during
surgery permits analysis to determine whether the herpes simplex
virus has been able to reach and replicate in the tumor tissue, and
also provides an option to establish whether a localised immune
response has been induced in the tumor tissue.
Immune Response
[0152] The inventors have identified that loco-regional
administration of oncolytic herpes simplex virus to patients having
cancer induces a strong Th1 immune response. The response appears
to mature and become sustained with time and be stronger in
patients receiving more than one dose of virus.
[0153] Oncolytic herpes simplex virus transfected into tumor cells
will replicate in and selectively lyse the tumor cells bringing
about tumor cell necrosis and the liberation of tumor antigens,
which trigger a Th1 immune response.
[0154] Infiltration of memory cytotoxic T cells and T helper 1
(Th1) T lymphocytes are often associated with a good clinical
outcome and good response to immunotherapy. Therefore, one aim of
immunotherapies is to modify the context of immune, inflammatory
and angiogenic elements to favour a strong Th1 cytotoxic
microenvironment (Giraldo et al., The immune contexture of primary
and metastatic human tumors. Current Opinion in Immunology 2014,
27:8-15).
[0155] Patients treated with oncolytic herpes simplex virus are
therefore well-suited to treatment with an immune checkpoint
inhibitor so as to provide either an enhanced treatment effect
and/or to convert patients having no or sub-optimal response to
immune checkpoint inhibitor therapy to become responders to such
therapy.
[0156] Accordingly, patients may optionally be treated with an
oncolytic herpes simplex virus in order to stimulate and establish
a Th1 immune response thereby priming the subject for treatment
with an immune checkpoint inhibitor.
[0157] Without wishing to be bound by theory of the invention,
treatment with an oncolytic herpes simplex virus may optionally be
used to establish a Th1 immune response in the subject. Having
established a Th1 response, treatment with an immune checkpoint
inhibitor may increase the magnitude of tumor specific T cell
responses as compared to treatment with an immune checkpoint
inhibitor alone. In particular, the response of a subject to
treatment with an immune checkpoint inhibitor may be improved by
use of an oncolytic herpes simplex virus to establish a Th1
response. Subjects who are non-responders or poor responders to
immune checkpoint inhibitor treatment may be converted to
responders.
[0158] As regards "response" to treatment, the mechanism of action
of herpes simplex virus therapy alone, immune checkpoint therapy
alone, and together in combination may result in the appearance of
disease progression on imaging using conventional RECIST criteria.
Response in clinical studies therefore needs to account for the
potential for "pseudo-progression" with these agents and to
recognise that an immunological response may take longer to
manifest itself as compared to chemotherapeutic agents. Imaging
guidelines have been modified to account for these factors and
immune related response criteria ("irRC") should be used to
evaluate imaging response.
[0159] In general, the combination of treatments may enhance the
systemic T-cell activation and the anti-tumor response to tumor
antigens following the lytic replication of oncolytic herpes
simplex virus in the cells of the cancer. This may improve the rate
of overall tumor response and duration of response. Overall, these
effects may provide an extension in overall survival, particularly
when compared to treatment using an immune checkpoint inhibitor
alone.
[0160] T helper (Th) cells play an important role in the adaptive
immune system, regulating the activity of other immune cells by
releasing certain cytokines. They are essential in the activation
and proliferation of cytotoxic T cells and in optimising the
activity of macrophages. Mature Th cells express CD4 (i.e. are
CD4.sup.+ T cells). Proliferating Th cells differentiate from a Th0
state into effector T cells of two main subtypes: Type 1 (Th1) and
Type 2 (Th2) (Kidd P. Altern Med Rev. 2003; 8(3):223-46; Sallusto
et al., Trends in Immunology. Volume 19, Issue 12, p 568-574, 1
Dec. 1998).
[0161] Differentiation towards Th1 is primarily triggered by IL-12
and IL-2. The primary effector cytokine of the Th1 response is
IFN-.gamma., although cytokines secreted by Th1 cells include Tumor
Necrosis Factor (TNF-.alpha.), IFN-.gamma. and interleukins (IL) 2,
12, and 18. IFN-.gamma. promotes production of IL-12 from dendritic
cells and macrophages which further promotes IFN-.gamma. production
in Th cells by a positive feedback mechanism. IFN-.gamma.
production also inhibits production of IL-4, and thereby inhibits
the Th2 response. Th1 immunity is mainly effected through
macrophages, CD8.sup.+ T cells, IgG B cells and IFN-.gamma.
CD4.sup.+ T cells. Th1 cytokines tend to produce pro-inflammatory
cytokines such as IL-6.
[0162] Differentiation towards Th2 is primarily triggered by IL-4
and the effector cytokines of the Th2 response include IL-4, IL-5,
IL-9, IL-10 and IL-13. Th2 immunity is mainly effected through
eosinophils, basophils and mast cells, as well as B cells and
IL-4/IL-5 CD4.sup.+ T cells. IL-10 acts to suppress Th1 cell
differentiation by inhibiting IL-2 and IFN-.gamma. in Th cells and
IL-12 in dendritic cells and macrophages.
[0163] Data presented herein from a trial in human patients shows
that single dose non-intratumoral administration of oncolytic
herpes simplex virus can be sufficient to induce an IFN.gamma.
response (e.g. FIGS. 1 and 22) but the response is not robustly
translated into upregulation of IL-2 and IL-12 (FIGS. 5, 9 and 23).
IFN.gamma. is pleiotropic (Trincheri and Perussia., Immune
interferon: a pleiotropic lymphokine with multiple effects.
Immunology Today, Volume 6, Issue 4, 131-136), and upregulation of
IFN.gamma. alone, without induction of threshold levels of IL-2
and/or IL-12, is not a reliable indicator of induction of a robust
or sustained Th1 response.
[0164] Administration of multiple doses of oncolytic herpes simplex
virus is shown to further upregulate IFN.gamma. and lead to
upregulation of IL-2 and IL-12 (FIG. 23) and expansion of T cells
associated with a Th1 response (FIG. 49). IL-2 and IL-12
upregulation leads to expansion of a Th1 cell population by a
positive feedback mechanism (Busse et al., Competing feedback loops
shape IL-2 signaling between helper and regulatory T lymphocytes in
cellular microenvironments. PNAS 2010 107 (7) 3058-3063; Vignali
and Kuchroo., IL-12 family cytokines: immunological playmakers.
Nature Immunology 13, 722-728 (2012)) which is modulated by
upregulated IL-10 (Taga and Tosato., IL-10 inhibits human T cell
proliferation and IL-2 production. J Immunol. 1992 feb 15;
148(4):1143-8).
[0165] The data obtained (FIGS. 23 and 49) show that multiple dose
administration of oncolytic herpes simplex virus leads to
upregulated IFN.gamma., IL-2, IL-12, IL-10 in human subjects, and
an appropriate T cell population, clearly indicating establishment
of a sustained Th1 response.
[0166] The inventors have identified that administration of
oncolytic herpes simplex virus to human patients having cancer
induces a strong Th1 immune response. The response appears to
mature and become sustained with time and be stronger in patients
receiving more than one dose of virus. Oncolytic herpes simplex
virus transfected into tumor cells will replicate in and
selectively lyse the tumor cells bringing about tumor cell necrosis
and the liberation of tumor antigens, which trigger a Th1 immune
response. Indeed, the inventors have noted that oncolytic herpes
simplex virus remodels the tumor microenvironment away from
immunosuppression by directly interacting with tumor infiltrating
immune cells.
[0167] Human patients treated with oncolytic herpes simplex virus
are therefore well-suited to treatment with an immune checkpoint
inhibitor so as to provide either an enhanced treatment effect
and/or to convert patients having no or sub-optimal response to
immune checkpoint inhibitor therapy to become responders to such
therapy.
[0168] Accordingly, human patients may be treated with an oncolytic
herpes simplex virus in order to stimulate and establish a Th1
immune response thereby priming the subject for treatment with an
immune checkpoint inhibitor.
[0169] Without wishing to be bound by theory of the invention,
treatment with an oncolytic herpes simplex virus may be used to
establish a Th1 immune response in the subject. Having established
a Th1 response treatment with an immune checkpoint inhibitor may
increase the magnitude of tumor specific T cell responses as
compared to treatment with an immune checkpoint inhibitor alone. In
particular, the response of a subject to treatment with an immune
checkpoint inhibitor may be improved by use of an oncolytic herpes
simplex virus to establish a Th1 response. Subjects who are
non-responders or poor responders to immune checkpoint inhibitor
treatment may be converted to responders.
[0170] In general, the combination of treatments may enhance the
systemic T-cell activation and the anti-tumor response to tumor
antigens following the lytic replication of oncolytic herpes
simplex virus in the cells of the cancer. This may lead to enhanced
destruction of tumors to which the virus has been administered,
e.g. by direct injection, but may also enhance the destruction of
tumors to which the virus has not been administered and/or are
distant to the site of administration, e.g. secondary/metastatic
tumors. This may improve the rate of overall tumor response and
duration of response. Overall, these effects may provide an
extension in overall survival, particularly when compared to
treatment using an immune checkpoint inhibitor alone.
[0171] The inventors have noted that mice that receive combination
therapy with oncolytic herpes simplex virus and an immune
checkpoint inhibitor showed more T cell recruitment to the tumor,
and displayed higher immune inflammatory responses and a less
immunosuppressive microenvironment, as measured by increased
proportions of CD4+ and CD8+ T cells relative to CD4+/CD25+/Fox3P+
Treg cells and immunosuppressive cytokines. The combination therapy
did not result in more NK, NKT or B cell recruitment or affect in
vivo virus activity but induced more inflammatory responses with
less immune regulatory/suppressive responses.
[0172] The inventors have identified that induction of a Th1
response through administration of an effective amount of oncolytic
herpes simplex virus is effective to trigger an anti-tumor Th1
response in the subject. They have also identified that the
anti-tumor Th1 response has a memory effect, and may be used to
vaccinate the subject against relapse of the cancer at both primary
and distal sites
Herpes Simplex Virus
[0173] The herpes simplex virus (HSV) genome comprises two
covalently linked segments, designated long (L) and short (S). Each
segment contains a unique sequence flanked by a pair of inverted
terminal repeat sequences. The long repeat (RL or R.sub.L) and the
short repeat (RS or R.sub.S) are distinct.
[0174] The HSV ICP34.5 (also called .gamma.34.5) gene, which has
been extensively studied, has been sequenced in HSV-1 strains F and
syn17+ and in HSV-2 strain HG52. One copy of the ICP34.5 gene is
located within each of the RL repeat regions. Mutants inactivating
one or both copies of the ICP34.5 gene are known to lack
neurovirulence, i.e. be avirulent/non-neurovirulent
(non-neurovirulence is defined by the ability to introduce a high
titre of virus (approx 10.sup.6 plaque forming units (pfu)) to an
animal or patient without causing a lethal encephalitis such that
the LD.sub.50 in animals, e.g. mice, or human patients is in the
approximate range of .gtoreq.10.sup.6 pfu), and be oncolytic.
[0175] An oncolytic virus is a virus that will lyse cancer cells
(oncolysis), preferably in a preferential or selective manner.
Viruses that selectively replicate in dividing cells over
non-dividing cells are often oncolytic. Oncolytic viruses are well
known in the art and are reviewed in Molecular Therapy Vol. 18 No.
2 Feb. 2010 pg 233-234.
[0176] Preferred herpes simplex virus are replication-competent,
being replication-competent at least in the target tumor/cancer
cells.
[0177] HSV that may be used in the present invention include HSV in
which one or both of the .gamma.34.5 (also called ICP34.5) genes
are modified (e.g. by mutation which may be a deletion, insertion,
addition or substitution) such that the respective gene is
incapable of expressing, e.g. encoding, a functional ICP34.5
protein. Preferably, in HSV according to the invention both copies
of the .gamma.34.5 gene are modified such that the modified HSV is
not capable of expressing, e.g. producing, a functional ICP34.5
protein.
[0178] In some embodiments the herpes simplex virus may be an
ICP34.5 null mutant where all copies of the ICP34.5 gene present in
the herpes simplex virus genome (two copies are normally present)
are disrupted such that the herpes simplex virus is incapable of
producing a functional ICP34.5 gene product. In other embodiments
the herpes simplex virus may lack at least one expressible ICP34.5
gene. In some embodiments the herpes simplex virus may lack only
one expressible ICP34.5 gene. In other embodiments the herpes
simplex virus may lack both expressible ICP34.5 genes. In still
other embodiments each ICP34.5 gene present in the herpes simplex
virus may not be expressible. Lack of an expressible ICP34.5 gene
means, for example, that expression of the ICP34.5 gene does not
result in a functional ICP34.5 gene product.
[0179] Herpes simplex virus may be derived from any HSV including
any laboratory strain or clinical isolate (non-laboratory strain)
of HSV. In some preferred embodiments the HSV is a mutant of HSV-1
or HSV-2. Alternatively the HSV may be an intertypic recombinant of
HSV-1 and HSV-2. The mutant may be of one of laboratory strains
HSV-1 strain 17, HSV-1 strain F or HSV-2 strain HG52. The mutant
may be of the non-laboratory strain JS-1. Preferably the mutant is
a mutant of HSV-1 strain 17. The herpes simplex virus may be one of
HSV-1 strain 17 mutant 1716, HSV-1 strain F mutant R3616, HSV-1
strain F mutant G207, HSV-1 mutant NV1020, or a further mutant
thereof in which the HSV genome contains additional mutations
and/or one or more heterologous nucleotide sequences. Additional
mutations may include disabling mutations, which may affect the
virulence of the virus or its ability to replicate. For example,
mutations may be made in any one or more of ICP6, ICP0, ICP4,
ICP27. Preferably, a mutation in one of these genes (optionally in
both copies of the gene where appropriate) leads to an inability
(or reduction of the ability) of the HSV to express the
corresponding functional polypeptide. By way of example, the
additional mutation of the HSV genome may be accomplished by
addition, deletion, insertion or substitution of nucleotides. In
some embodiments the HSV genome does not have a mutation in ICP6,
or in ICP0, ICP4, ICP27.
[0180] A number of oncolytic herpes simplex viruses are known in
the art. Examples include HSV1716, R3616 (e.g. see Chou &
Roizman, Proc. Natl. Acad. Sci. Vol. 89, pp. 3266-3270, April
1992), G207 (Toda et al, Human Gene Therapy 9:2177-2185, Oct. 10,
1995), NV1020 (Geevarghese et al, Human Gene Therapy 2010
September; 21(9):1119-28), RE6 (Thompson et al, Virology 131,
171-179 (1983)), and Oncovex.TM. (Simpson et al, Cancer Res 2006;
66:(9) 4835-4842 May 1, 2006; Liu et al, Gene Therapy (2003): 10,
292-303), dlsptk, hrR3, R4009, MGH-1, MGH-2, G47.DELTA., Myb34.5,
DF3.gamma.34.5, HF10, NV1042, RAMBO, rQNestin34.5, R5111, R-LM113,
CEAICP4, CEA.gamma.34.5, DF3.gamma.34.5, KeM34.5 (Manservigi et al,
The Open Virology Journal 2010; 4:123-156), rRp450, M032
(Campadelli-Fiume et al, Rev Med. Virol 2011; 21:213-226), Baco1
(Fu et al, Int. J. Cancer 2011; 129(6):1503-10) and M032 and C134
(Cassady et al, The Open Virology Journal 2010; 4:103-108).
[0181] In some preferred embodiments the herpes simplex virus is
HSV-1 strain 17 mutant 1716 (HSV1716). HSV 1716 is an oncolytic,
non-neurovirulent HSV and is described in EP 0571410, WO 92/13943,
Brown et al (Journal of General Virology (1994), 75, 2367-2377) and
MacLean et al (Journal of General Virology (1991), 72, 631-639).
HSV 1716 has been deposited on 28 Jan. 1992 at the European
Collection of Animal Cell Cultures, Vaccine Research and Production
Laboratories, Public Health Laboratory Services, Porton Down,
Salisbury, Wiltshire, SP4 0JG, United Kingdom under accession
number V92012803 in accordance with the provisions of the Budapest
Treaty on the International Recognition of the Deposit of
Microorganisms for the Purposes of Patent Procedure (herein
referred to as the `Budapest Treaty`).
[0182] In some embodiments the herpes simplex virus is a mutant of
HSV-1 strain 17 modified such that both ICP34.5 genes do not
express a functional gene product, e.g. by mutation (e.g.
insertion, deletion, addition, substitution) of the ICP34.5 gene,
but otherwise resembling or substantially resembling the genome of
the wild type parent virus HSV-1 strain 17+. That is, the virus may
be a variant of HSV1716, having a genome mutated so as to
inactivate both copies of the ICP34.5 gene of HSV-1 strain 17+ but
not otherwise altered to insert or delete/modify other protein
coding sequences.
[0183] In some embodiments the genome of an Herpes Simplex Virus
according to the present invention may be further modified to
contain nucleic acid encoding at least one copy of a polypeptide
that is heterologous to the virus (i.e. is not normally found in
wild type virus) such that the polypeptide can be expressed from
the nucleic acid. As such, the virus may also be an expression
vector from which the polypeptide may be expressed. Examples of
such viruses are described in WO2005/049846 and WO2005/049845.
[0184] In order to effect expression of the polypeptide, nucleic
acid encoding the polypeptide is preferably operably linked to a
regulatory sequence, e.g. a promoter, capable of effecting
transcription of the nucleic acid encoding the polypeptide. A
regulatory sequence (e.g. promoter) that is operably linked to a
nucleotide sequence may be located adjacent to that sequence or in
close proximity such that the regulatory sequence can effect and/or
control expression of a product of the nucleotide sequence. The
encoded product of the nucleotide sequence may therefore be
expressible from that regulatory sequence.
[0185] In some preferred embodiments, the Herpes Simplex Virus is
not modified to contain nucleic acid encoding at least one copy of
a polypeptide (or other nucleic acid encoded product) that is
heterologous to the virus. That is the virus is not an expression
vector from which a heterologous polypeptide or other nucleic acid
encoded product may be expressed. Such HSV are not suitable for, or
useful in, gene therapy methods and the method of medical treatment
for which they are employed may optionally be one that does not
involve gene therapy.
[0186] In some embodiments the genome of an oncolytic Herpes
Simplex Virus according to the present invention does not encode
(or is not further modified to contain nucleic acid encoding) a
cytokine or chemokine, e.g. a mammalian or human cytokine or
chemokine. For example, the genome of an oncolytic Herpes Simplex
Virus according to the present invention does not encode an
interleukin, e.g. IL-2 and/or IL-12, an interferon, e.g.
IFN-.gamma., a tumor necrosis factor, a colony stimulating factor
(e.g. GM-CSF, G-CSF), an immune modulator, a member of the CC
family, e.g. CCLS, a member of the CXC family or a member of the
CXC family.
[0187] In some embodiments the herpes simplex virus has an intact
ICP0 gene, capable of expressing functional ICP0. In some
embodiments the herpes simplex virus has an intact ICP27 gene,
capable of expressing functional ICP27. In some embodiments the
herpes simplex virus has an intact vhs gene, capable of expressing
functional vhs.
[0188] In some embodiments the herpes simplex virus has an intact
ICP47 gene, capable of expressing functional ICP47. In some
embodiments the oncolytic herpes simplex virus has an intact ICP6
gene, capable of expressing functional ICP6.
[0189] Optionally, in some embodiments the herpes simplex virus
does not encode or express (granulocyte macrophage colony
stimulating factor) GMCSF.
[0190] Optionally, in some embodiments the herpes simplex virus is
not a herpes simplex virus that lacks functional ICP34.5 genes and
lacks a functional ICP47 gene and comprises a gene encoding human
GM-CSF.
[0191] In some optional embodiments the herpes simplex virus is not
Talimogene laherparepvec, HSV-1 [strain JS1]
ICP34.5-/ICP47-/hGM-CSF also known as OncoVEX GM-CSF (Lui et al.,
Gene Therapy, 10:292-303, 2003; U.S. Pat. No. 7,223,593 and U.S.
Pat. No. 7,537,924)]. In talimogene laherparepvec, the HSV-1 viral
genes encoding ICP34.5 are functionally deleted, the ICP47 is
functionally deleted, the coding sequence for human GM-CSF is
inserted into the viral genome such that it replaces nearly all of
the ICP34.5 gene and the HSV thymidine kinase (TK) gene remains
intact.
[0192] In some optional embodiments the herpes simplex virus is not
a herpes simplex virus that lacks only one of the two functional
copies of the .gamma..sub.134.5 gene. For example, in some optional
embodiments the herpes simplex virus is not NV1020 or a variant
thereof.
[0193] Herpes simplex viruses may be formulated as medicaments and
pharmaceutical compositions for clinical use and in such
formulations may be combined with a pharmaceutically acceptable
carrier, diluent or adjuvant. The composition is preferably
formulated for intravenous or intra-arterial routes of
administration which may include infusion or injection. Suitable
formulations may comprise the virus in a sterile or isotonic
medium. Medicaments and pharmaceutical compositions may be
formulated in fluid (including gel) or solid (e.g. tablet) form.
Fluid formulations may be formulated for administration by
injection or via catheter to a selected region of the human or
animal body.
[0194] Administration is preferably in a "therapeutically effective
amount", this being sufficient to show benefit to the individual.
The actual amount administered, and rate and time-course of
administration, will depend on the nature and severity of the
disease being treated. Prescription of treatment, e.g. decisions on
dosage etc, is within the responsibility of general practitioners
and other medical doctors, and typically takes account of the
disorder to be treated, the condition of the individual patient,
the site of delivery, the method of administration and other
factors known to practitioners. Examples of the techniques and
protocols mentioned above can be found in Remington's
Pharmaceutical Sciences, 20th Edition, 2000, pub. Lippincott,
Williams & Wilkins.
[0195] Targeting therapies may be used to deliver the virus to
certain types of cell, e.g. by the use of targeting systems such as
antibody or cell specific ligands. Targeting may be desirable for a
variety of reasons; for example if the virus is unacceptably toxic
in high dose, or if it would otherwise require too high a dosage,
or if it would not otherwise be able to enter the target cells.
[0196] HSV capable of targeting cells and tissues are described in
(PCT/GB2003/000603; WO 03/068809), hereby incorporated in its
entirety by reference.
[0197] An HSV may be administered alone or in combination with
other treatments, either simultaneously or sequentially dependent
upon the condition to be treated. Such other treatments may include
chemotherapy (including either systemic treatment with a
chemotherapeutic agent or targeted therapy using small molecule or
biological molecule (e.g. antibody) based agents that target key
pathways in tumor development, maintenance or progression) or
radiotherapy provided to the subject as a standard of care for
treatment of the cancer.
[0198] In addition to direct action of oncolytic herpes simplex
virus (oHSV) on tumors, there is growing evidence that the host
immune response plays an important role in establishing the
efficacy of the anti-tumor response through innate immune
effectors, adaptive antiviral immune responses and adaptive
antitumor immune responses (e.g. see Prestwich et al., Oncolytic
viruses: a novel form of immunotherapy. Expert Rev Anticancer Ther.
October 2008; 8(10): 1581-1588).
[0199] Several studies have shown that oHSV is capable of inducing
an anti-tumor immune response. This can manifest as tumor growth
reduction in lesions treated with oHSV and in untreated lesions in
the same animal, efficacy of oHSV requiring an intact immune
response, induction of anti-tumor cytokine response, reversal of
tumor immune dysfunction and facilitation of tumor antigen
presentation. Induction of an anti-tumor immune response can reduce
establishment of metastases, or contribute to their elimination,
and protect from re-occurrence of tumor.
[0200] For example, in Benencia et al., ((2008) Herpes virus
oncolytic therapy reverses tumor immune dysfunction and facilitates
tumor antigen presentation. Cancer Biol. Ther. 7, 1194-1205) growth
reduction in treated and untreated lesions was reported. In Miller
and Fraser ((2003) Requirement of an integrated immune response for
successful neuroattenuated HSV-1 therapy in an intracranial
metastatic melanoma model. Mol. Ther. 7(6):741-747) efficacy of
HSV176 required an intact immune response which was mediated by a
tumor-specific proliferative T cell response.
Administration of Herpes Simplex Virus
[0201] Administration of herpes simplex virus is preferably for a
period of time sufficient to allow virus to reach the site of the
tumor tissue, and preferably to begin to replicate in the tumor
tissue. It may also be for a period of time sufficient to induce or
elicit a Th1 response in the subject.
[0202] This may involve administration at regular intervals, e.g.
weekly or fortnightly, of doses of herpes simplex virus sufficient
to allow accumulation of virus at the tumor site and/or to induce a
sustained Th1 response over a period of time. For example, doses
may be given at regular, defined, intervals over a period of one of
at least 1, 2, 3, 4, 5, 6, 7, 8, weeks or 1, 2, 3, 4, 5 or 6
months.
[0203] As such, multiple doses of herpes simplex virus may be
administered. For example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more
doses of herpes simplex virus may be administered to a subject as
part of a course of treatment. In some embodiments one of at least
1, 2, 3, or 4 doses of herpes simplex virus are administered to the
subject, preferably at regular intervals (e.g. weekly). In some
embodiments this may prime the subject for treatment with an immune
checkpoint inhibitor.
[0204] Doses of herpes simplex virus may be separated by a
predetermined time interval, which may be selected to be one of 1,
2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or 31 days, or 1, 2, 3, 4,
5, or 6 months. By way of example, doses may be given once every 7,
14, 21 or 28 days (plus or minus 3, 2, or 1 days). The dose of
herpes simplex virus given at each dosing point may be the same,
but this is not essential. For example, it may be appropriate to
give a higher priming dose at the first, second and/or third dosing
points.
[0205] Administration of oncolytic herpes simplex virus may be of
one or more treatment cycles, e.g. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or
more treatment cycles. A subject receiving multiple treatment
cycles may be given subsequent treatment cycles consecutively,
without a break from treatment, or may separate all or selected
treatment cycles by a break from treatment, e.g. a break of 1, 2,
3, 4, 5, 6, 7, 8 or 9 days or about 1, 2, 3, or 4 weeks.
[0206] In some embodiments a treatment cycle may comprise, or
consist of, 4 doses of oncolytic herpes simplex virus, one dose per
week over a period of 4 weeks. In some embodiments a treatment
cycle may comprise, or consist of, 8 doses of oncolytic herpes
simplex virus, one dose per week over a period of 8 weeks. Weekly
doses may be separated by 7.+-.1 or 7.+-.2 days. For example,
weekly doses may be given on days 1, 8, 15 and 22.
[0207] In some embodiments a treatment cycle may comprise, or
consist of, 4 doses of oncolytic herpes simplex virus, two doses
per week over a period of 2 weeks. In some embodiments a treatment
cycle may comprise, or consist of, 8 doses of oncolytic herpes
simplex virus, two doses per week over a period of 4 weeks. Twice
weekly doses may be separated by 4.+-.1 or 4.+-.2 days. For
example, weekly doses may be given on days 1, 5, 8, 13 or 1, 5, 8,
12.
[0208] Subjects may receive the same dosage at each administration
within a given treatment cycle, e.g. a dosage of 1.times.10.sup.7
iu or 1.times.10.sup.8 iu, or between 1.times.10.sup.6 and
1.times.10.sup.8 iu or between 1.times.10.sup.7 iu and
1.times.10.sup.8 iu. In some embodiments the first 1, 2 or 3
treatment cycles may comprise administration of a lower dosage
amount at each administration, e.g. 1.times.10.sup.7 iu, and later
treatment cycles may comprise administration of a higher dosage
amount at each administration, e.g. 1.times.10.sup.8 iu.
[0209] Administration of oncolytic herpes simplex virus may be by
infusion to the blood (intravenous or intra-arterial) and subjects
will preferably attend clinic on the scheduled administration days
for administration of oncolytic herpes simplex virus.
Administration of oncolytic herpes simplex virus may continue until
development of severe toxicity or withdrawal of consent.
[0210] A tumour biopsy may be taken in the period commencing 14
days before the first dose of oncolytic herpes simplex virus (Day
1). A tumour biopsy or surgical resection sample may be taken after
completion of a cycle of treatment, e.g. within 14 days of the last
dose of oncolytic herpes simplex virus in a given treatment cycle.
Samples obtained from tumour biopsy or surgical resection may be
used to determine the presence and/or maintenance of a Th1
response.
[0211] Blood or serum samples may be taken at the stage of initial
subject assessment (before treatment with oncolytic herpes simplex
virus), and during a or each treatment cycle, e.g. on days 1, 8,
15, 22, for weekly administration, days 1, 5, 8, 13, or days 1, 5,
8, 12 for twice weekly administration. Blood or serum samples may
be used to determine the presence and/or maintenance of a Th1
response.
[0212] Where a Th1 response has been induced the subject may
continue with dosing of oncolytic herpes simplex virus on the same
dosing schedule in order to maintain the response. Alternatively,
dosing frequency may be reduced and the subject may receive either
no further doses of oncolytic herpes simplex virus, e.g. in cases
where the Th1 response is self-sustaining following induction, or
the status of the Th1 response in the subject may be monitored and
the subject may be given booster administrations as and when
considered appropriate, e.g. by a medical practitioner, in order to
maintain the sustained Th1 response.
[0213] Suitable dosage amounts of herpes simplex virus may be in
the range 10.sup.6 to 10.sup.9 iu or 2.times.10.sup.6 to 10.sup.9
iu. Doses of herpes simplex virus in this range may be particularly
required for systemic, e.g. intravenous, administration where the
viral dose is diluted by administration to the blood. Each dose of
herpes simplex virus is preferably of greater than 2.times.10.sup.6
iu. Each dose of virus may be in a range selected from the group
consisting of: 2.times.10.sup.6 to 9.times.10.sup.6 iu,
2.times.10.sup.6 to 5.times.10.sup.6 iu, 5.times.10.sup.6 to
9.times.10.sup.6 iu, 2.times.10.sup.6 to 1.times.10.sup.7 iu,
2.times.10.sup.6 to 5.times.10.sup.7 iu, 2.times.10.sup.6 to
1.times.10.sup.8 iu, 2.times.10.sup.6 to 5.times.10.sup.8 iu,
2.times.10.sup.6 to 1.times.10.sup.9 iu, 5.times.10.sup.6 to
1.times.10.sup.7 iu, 5.times.10.sup.6 to 5.times.10.sup.7 iu,
5.times.10.sup.6 to 1.times.10.sup.8 iu, 5.times.10.sup.6 to
5.times.10.sup.8 iu, 5.times.10.sup.6 to 1.times.10.sup.9 iu,
5.times.10.sup.6 to 5.times.10.sup.9 iu, 1.times.10.sup.7 to
9.times.10.sup.7 iu, 1.times.10.sup.7 to 5.times.10.sup.7 iu,
1.times.10.sup.8 to 9.times.10.sup.8 iu, 1.times.10.sup.8 to
5.times.10.sup.8 iu. In some embodiments suitable doses may be in
the range 2.times.10.sup.6 to 9.times.10.sup.6 iu, 1.times.10.sup.7
to 9.times.10.sup.7 iu, or 1.times.10.sup.8 to 9.times.10.sup.8 iu.
In some embodiments suitable doses may be about 1.times.10.sup.7 iu
or 1.times.10.sup.8 iu. Dosage figures may optionally be +/-half a
log value.
[0214] The term `infectious units` is used to refer to virus
concentrations derived using the TCID50 method and `plaque forming
units (pfus)` to refer to plaque-based assay results. As 1 iu will
form a single plaque in a titration assay, 1 iu is equivalent to 1
pfu.
[0215] In general, administration is preferably in a "effective
amount", this optionally being sufficient to induce a Th1 response
in the individual and/or for the virus to have an independent
treatment effect on the cancer. The actual amount administered, and
rate and time-course of administration, will depend on the nature
and severity of the disease being treated. Prescription of
treatment, e.g. decisions on dosage etc, is within the
responsibility of general practitioners and other medical doctors,
and typically takes account of the disorder to be treated, the
condition of the individual patient, the site of delivery, the
method of administration and other factors known to practitioners.
Examples of the techniques and protocols mentioned above can be
found in Remington's Pharmaceutical Sciences, 20th Edition, 2000,
pub. Lippincott, Williams & Wilkins.
[0216] In some embodiments herpes simplex virus is administered in
the form of monotherapy, i.e. not part of a combination treatment,
although subjects may also receive, or continue to receive,
standard of care chemotherapy or radiation therapy. In other
embodiments herpes simplex virus may be administered as part of a
programme of treatment in which an immune checkpoint inhibitor is
also administered, the two agents providing a combination therapy
treatment.
[0217] Administration of herpes simplex virus may be carried out
for a period of time prior to treatment with an immune checkpoint
inhibitor in which period the subject receives herpes simplex virus
but does not receive an immune checkpoint inhibitor. Treatment is
preferably for a period of time suitable, or sufficient, to allow
virus to reach the site of the tumor tissue, and preferably to
begin to replicate in the tumor tissue so as to be detectable in a
tumor tissue sample, e.g. biopsy, and/or to induce or elicit a Th1
immune response in the subject. This may be referred to as
"pre-treatment" with herpes simplex virus.
[0218] In some preferred embodiments pre-treatment may involve a
period of time in which the subject is administered herpes simplex
virus but is not administered an immune checkpoint inhibitor
(called "herpes simplex virus monotherapy" herein). The period of
herpes simplex virus monotherapy may be sufficient to allow virus
to reach the site of the tumor tissue and/or to induce or elicit a
Th1 response in the subject. During a period of herpes simplex
virus monotherapy the subject may also receive treatment in the
form of simultaneous, sequential or separate administration of
other chemotherapy or radiation therapy, e.g. which may be part of
the standard of care for the cancer being treated, but in that time
period the patient will not receive a therapeutically effective
dose of an immune checkpoint inhibitor.
[0219] As such, methods according to the present invention may
comprise administration of an herpes simplex virus for a period of
time in which the subject receives herpes simplex virus but does
not receive an immune checkpoint inhibitor. The period of time may
be suitable, or sufficient, to allow virus to reach the site of the
tumor tissue and/or to induce a Th1 response in the subject.
[0220] Following the pre-treatment a determination may be made as
to whether virus has reached the site of the tumor tissue and/or
whether a Th1 response has been induced or elicited in the subject.
The determination may involve analysis of a tumor tissue sample
obtained during surgery and following herpes simplex virus
monotherapy. A selection of subject(s) suitable for treatment with
an immune checkpoint inhibitor may then be made.
[0221] A subject may then begin treatment with an immune checkpoint
inhibitor. That is, the method may then further comprises the
administration of an immune checkpoint inhibitor to the
subject.
[0222] Accordingly, at a selected time point the period of
pre-treatment may end and the subject may then be administered an
immune checkpoint inhibitor. Optionally the subject will continue
to be administered herpes simplex virus simultaneously,
sequentially or separately such that the subject receives
co-therapy with immune checkpoint inhibitor and herpes simplex
virus. The subject may also receive, or continue to receive,
treatment in the form of simultaneous, sequential or separate
administration of other chemotherapy or radiation therapy, e.g.
which may be part of the standard of care for the cancer being
treated.
[0223] For example, pre-treatment may occur for one of at least 1,
2, 3, 4, or 5 weeks in which the subject receives herpes simplex
virus but does not receive an immune checkpoint inhibitor.
Preferably the period of time is sufficient to allow virus to reach
the site of the tumor tissue and/or to induce or elicit a Th1
response in the subject. By way of example, a subject may receive
herpes simplex virus monotherapy in the form of weekly doses of
herpes simplex virus for one of at least 1, 2, 3, 4, 5, 6, 7, 8,
weeks or 1, 2, 3, 4, 5 or 6 months.
[0224] In other embodiments a subject may receive herpes simplex
virus monotherapy as described above and may discontinue treatment
with herpes simplex virus and begin receiving treatment with an
immune checkpoint inhibitor. In such embodiments there may be no
day on which a subject is receiving co-therapy, i.e. no day on
which an ongoing scheduled programme of treatment with herpes
simplex virus and immune checkpoint inhibitor overlaps.
[0225] In other embodiments there may a substantial overlap of
treatment with herpes simplex virus. In one arrangement, co-therapy
with herpes simplex virus and immune checkpoint inhibitor may
commence at the start of treatment, or during a period in which the
subject is receiving an herpes simplex virus, e.g. in order to
induce or elicit a Th1 response, preferably a sustained Th1
response. In other arrangements a short period of herpes simplex
virus monotherapy may be provided, not necessarily suitable or
sufficient to induce or elicit a Th1 response, after which the
subject begins to also receive treatment with an immune checkpoint
inhibitor, i.e. co-therapy. During co-therapy the herpes simplex
virus and immune checkpoint inhibitor may be administered on the
same day or on different days.
[0226] Co-therapy may comprises simultaneous or sequential
administration of herpes simplex virus and immune checkpoint
inhibitor.
[0227] Simultaneous administration refers to administration of the
herpes simplex virus and immune checkpoint inhibitor together, for
example as a pharmaceutical composition containing both agents, or
immediately after each other and optionally via the same route of
administration, e.g. to the same artery, vein or other blood
vessel.
[0228] Sequential administration refers to administration of one of
the herpes simplex virus or immune checkpoint inhibitor followed
after a given time interval by separate administration of the other
agent. It is not required that the two agents are administered by
the same route, although this is the case in some embodiments. The
time interval may be any time interval.
[0229] Whilst simultaneous or sequential administration may be
intended such that both the herpes simplex virus and immune
checkpoint inhibitor are delivered to the same tumor tissue to
effect treatment it is not essential for both agents to be present
in the tumor tissue in active form at the same time.
[0230] However, in some embodiments of sequential administration
the time interval is selected such that the herpes simplex virus
and immune checkpoint inhibitor are expected to be present in the
tumor tissue in active form at the same time, thereby allowing for
a combined, additive or synergistic effect of the two agents in
treating the tumor. In such embodiments the time interval selected
may be any one of 5 minutes or less, 10 minutes or less, 15 minutes
or less, 20 minutes or less, 25 minutes or less, 30 minutes or
less, 45 minutes or less, 60 minutes or less, 90 minutes or less,
120 minutes or less, 180 minutes or less, 240 minutes or less, 300
minutes or less, 360 minutes or less, or 720 minutes or less, or 1
day or less, or 2 days or less.
[0231] In some embodiments, a subject will receive oncolytic herpes
simplex virus before treatment with the immune checkpoint inhibitor
that is intended to take advantage of the Th1 response which the
oncolytic herpes simplex virus may induce or elicit when providing
a treatment effect.
[0232] Where co-therapy with an herpes virus occurs it may continue
for as long as desired or prescribed. In some embodiments,
treatment with herpes simplex virus may be discontinued in favour
of continued treatment with the immune checkpoint inhibitor.
[0233] Doses of immune checkpoint inhibitor may also be separated
by a predetermined time interval, which may be selected to be one
of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,
19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or 31 days, or 1,
2, 3, 4, 5, or 6 months. Administration is preferably in a
"therapeutically effective amount", this being sufficient to show
benefit to the individual. The actual amount administered, and rate
and time-course of administration, will depend on the nature and
severity of the disease being treated. Prescription of treatment,
e.g. decisions on dosage etc, is within the responsibility of
general practitioners and other medical doctors, and typically
takes account of the disorder to be treated, the condition of the
individual patient, the site of delivery, the method of
administration and other factors known to practitioners. Examples
of the techniques and protocols mentioned above can be found in
Remington's Pharmaceutical Sciences, 20th Edition, 2000, pub.
Lippincott, Williams & Wilkins.
[0234] During co-therapy the subject may also receive, or continue
to receive, treatment in the form of simultaneous, sequential or
separate administration of other chemotherapy or radiation therapy,
e.g. which may be part of the standard of care for the cancer being
treated.
[0235] A Th1 response is preferably a sustained Th1 response. By
"sustained Th1 response" we mean that the level of one or more
relevant cytokines and/or relevant T cell population is upregulated
for more than 7 days, and preferably for one of at least 2, 3, 4,
5, 6, 7, or 8 weeks, or for one of at least 1, 2, 3, 4, 5, or 6
months.
Immune Checkpoint Proteins and Inhibitors
[0236] The term "immune checkpoint inhibitor" refers to molecules
that totally or partially reduce, inhibit, interfere with or
modulate one or more immune checkpoint proteins. An inhibitor may
inhibit or block the interaction of an immune checkpoint protein
with one of its ligands or receptors.
[0237] Immune checkpoint proteins negatively regulate T-cell
activation or function. Numerous immune checkpoint proteins are
known, such as CTLA-4 (Cytotoxic T-Lymphocyte-Associated protein 4)
and its ligands CD80 and CD86; and PD-1 (Programmed Death 1) with
its ligands PD-L1 and PD-L2 (Pardoll, Nature Reviews Cancer 12:
252-264, 2012), TIM-3 (T-cell Immunoglobulin domain and Mucin
domain 3), LAG-3 (Lymphocyte Activation Gene-3), BTLA (CD272 or B
and T Lymphocyte Attenuator), KIR (Killer-cell Immunoglobulin-like
Receptor), VISTA (V-domain immunoglobulin suppressor of T-cell
activation), and A2aR (Adenosine A2A receptor). These proteins are
responsible for down-regulating T-cell responses. Immune checkpoint
proteins regulate and maintain self-tolerance and the duration and
amplitude of physiological immune responses. Immune checkpoint
inhibitors include antibodies and small molecule inhibitors.
[0238] Cytotoxic T-lymphocyte associated antigen 4 (CTLA-4) is an
immune checkpoint protein that down-regulates pathways of T-cell
activation (Fong et al., Cancer Res. 69(2):609-5 615, 2009; Weber
Cancer Immunol. Immunother, 58:823-830, 2009). CTLA-4 is a negative
regulator of T-cell activation. Blockade of CTLA-4 has been shown
to augment T-cell activation and proliferation. Inhibitors of
CTLA-4 include anti-CTLA-4 antibodies. Anti-CTLA-4 antibodies bind
to CTLA-4 and block the interaction of CTLA-4 with its ligands
CD80/CD86 expressed on antigen presenting cells and thereby
blocking the negative down regulation of the immune responses
elicited by the interaction of these molecules. Examples of
anti-CTLA-4 antibodies are described in U.S. Pat. Nos. 5,811,097;
5,811,097; 5,855,887; 6,051,227; 6,207,157; 6,682,736; 6,984,720;
and 7,605,238.
[0239] Anti-CTLA-4 antibodies include tremelimumab, (ticilimumab,
CP-675,206), ipilimumab (also known as IODI, MDX-DOIO; marketed
under the name Yervoy.TM. and) a fully human monoclonal IgG
antibody that binds to CTLA-4 approved for the treatment of
unresectable or metastatic melanoma.
[0240] Another immune checkpoint protein is programmed cell death 1
(PD-1). PD-1, also called CD279, is a type I membrane protein
encoded in humans by the PDCD1 gene. It has two ligands, PD-L1 and
PD-L2. The PD-1 pathway is a key immune-inhibitory mediator of
T-cell exhaustion. Blockade of this pathway can lead to T-cell
activation, expansion, and enhanced effector functions. As such,
PD-1 negatively regulates T cell responses. PD-1 has been
identified as a marker of exhausted T cells in chronic disease
states, and blockade of PD-1:PD-1L interactions has been shown to
partially restore T cell function. (Sakuishi et al., JEM Vol. 207,
Sep. 27, 2010, pp 2187-2194). PD-1 limits the activity of T cells
in peripheral tissues at the time of an inflammatory response to
infection and to limit autoimmunity. PD-1 blockade in vitro
enhances T-cell proliferation and cytokine production in response
to a challenge by specific antigen targets or by allogeneic cells
in mixed lymphocyte reactions. A strong correlation between PD-1
expression and response was shown with blockade of PD-1 (Pardoll,
Nature Reviews Cancer, 12: 252-264, 2012). PD-1 blockade can be
accomplished by a variety of mechanisms including antibodies that
bind PD-1 or its ligand, PD-L1, or soluble PD-1 decoy receptors
(e.g. sPD-1, see Pan et al., Oncology Letters 5: 90-96, 2013).
Examples of PD-1 and PD-L1 blockers are described in U.S. Pat. Nos.
7,488,802; 7,943,743; 8,008,449; 8,168,757; 8,217,149, and PCT
Published Patent Application Nos: WO03042402, WO2008156712,
WO2010089411, WO2010036959, WO2011066342, WO2011159877,
WO2011082400, and WO2011161699.
[0241] PD-1 blockers include anti-PD-1 and anti-PD-L1 antibodies
and proteinaceous binding agents. Nivolumab (BMS-936558) is an
anti-PD-1 antibody that was approved for the treatment of melanoma
in Japan in July 2014. It is a fully human IgG4 antibody that binds
to and blocks the activation of PD-1 by its ligands PD-L1 and
PD-L2. Other anti-PD-1 antibodies include pembrolizumab
(lambrolizumab; MK-3475 or SCH 900475), a humanized monoclonal IgG4
antibody against PD-1; CT-011 a humanized antibody that binds PD-1.
AMP-224 is a fusion protein of B7-DC; an antibody Fc portion;
BMS-936559 (MDX-1105-01) for PD-L1 (B7-HI) blockade. Other
anti-PD-1 antibodies are described in WO 2010/077634, WO
2006/121168, WO2008/156712 and WO2012/135408. AUNP-12 (Aurigene) is
a branched 29 amino acid peptide antagonist of the interaction of
PD-1 with PD-L1 or PD-L2 and has been shown to inhibit tumor growth
and metastasis in preclinical models of cancer.
[0242] T cell immunoglobulin mucin 3 (TIM-3) is an immune regulator
identified as being upregulated on exhausted CD8.sup.+ T cells
(Sakuishi et al., JEM Vol. 207, Sep. 27, 2010, pp 2187-2194 and
Fourcade et al., 2010, J. Exp. Med. 207:2175-86). TIM-3 was
originally identified as being selectively expressed on
IFN-.gamma.-secreting Th1 and Tc1 cells. Interaction of TIM-3 with
its ligand, galectin-9, triggers cell death in TIM-3.sup.+ T cells.
Anti-TIM-3 antibodies are described in Ngiow et al (Cancer Res.
2011 May 15; 71(10):3540-51), and in U.S. Pat. No. 8,552,156
[0243] Other immune-checkpoint inhibitors include lymphocyte
activation gene-3 (LAG-3) inhibitors, such as IMP321, a soluble Ig
fusion protein (Brignone et al., 2007, J. Immunol. 179:4202-4211).
Other immune-checkpoint inhibitors include B7 inhibitors, such as
B7-H3 and B7-H4 inhibitors. In particular, the anti-B7-H3 antibody
MGA271 (Loo et al., 2012, 5 Clin. Cancer Res. July 15 (18)
3834).
[0244] Reference to an "antibody" includes a fragment or derivative
thereof, or a synthetic antibody or synthetic antibody fragment.
Antibodies may be provided in isolated form or may be formulated as
a medicament or pharmaceutical composition, e.g. combined with a
pharmaceutically acceptable adjuvant, carrier, diluent or
excipient.
[0245] In view of today's techniques in relation to monoclonal
antibody technology, antibodies can be prepared to most antigens.
The antigen-binding portion may be a part of an antibody (for
example a Fab fragment) or a synthetic antibody fragment (for
example a single chain Fv fragment [ScFv]). Suitable monoclonal
antibodies to selected antigens may be prepared by known
techniques, for example those disclosed in "Monoclonal Antibodies:
A manual of techniques", H Zola (CRC Press, 1988) and in
"Monoclonal Hybridoma Antibodies: Techniques and Applications", J G
R Hurrell (CRC Press, 1982). Chimaeric antibodies are discussed by
Neuberger et al (1988, 8th International Biotechnology Symposium
Part 2, 792-799).
[0246] Monoclonal antibodies (mAbs) are useful in the methods of
the invention and are a homogenous population of antibodies
specifically targeting a single epitope on an antigen.
[0247] Polyclonal antibodies may also be useful in the methods of
the invention. Monospecific polyclonal antibodies are preferred.
Suitable polyclonal antibodies can be prepared using methods well
known in the art.
[0248] Fragments of antibodies, such as Fab and Fab.sub.2 fragments
may also be provided as can genetically engineered antibodies and
antibody fragments. The variable heavy (V.sub.H) and variable light
(V.sub.L) domains of the antibody are involved in antigen
recognition, a fact first recognised by early protease digestion
experiments. Further confirmation was found by "humanisation" of
rodent antibodies. Variable domains of rodent origin may be fused
to constant domains of human origin such that the resultant
antibody retains the antigenic specificity of the rodent parented
antibody (Morrison et al (1984) Proc. Natl. Acad. Sd. USA 81,
6851-6855).
[0249] That antigenic specificity is conferred by variable domains
and is independent of the constant domains is known from
experiments involving the bacterial expression of antibody
fragments, all containing one or more variable domains. These
molecules include Fab-like molecules (Better et al (1988) Science
240, 1041); Fv molecules (Skerra et al (1988) Science 240, 1038);
single-chain Fv (ScFv) molecules where the V.sub.H and V.sub.L
partner domains are linked via a flexible oligopeptide (Bird et al
(1988) Science 242, 423; Huston et al (1988) Proc. Natl. Acad. Sd.
USA 85, 5879) and single domain antibodies (dAbs) comprising
isolated V domains (Ward et al (1989) Nature 341, 544). A general
review of the techniques involved in the synthesis of antibody
fragments which retain their specific binding sites is to be found
in Winter & Milstein (1991) Nature 349, 293-299.
[0250] By "ScFv molecules" we mean molecules wherein the V.sub.H
and V.sub.L partner domains are covalently linked, e.g. by a
flexible oligopeptide.
[0251] Fab, Fv, ScFv and dAb antibody fragments can all be
expressed in and secreted from E. coli, thus allowing the facile
production of large amounts of the said fragments.
[0252] Whole antibodies, and F(ab').sub.2 fragments are "bivalent".
By "bivalent" we mean that the said antibodies and F(ab').sub.2
fragments have two antigen combining sites. In contrast, Fab, Fv,
ScFv and dAb fragments are monovalent, having only one antigen
combining site. Synthetic antibodies which bind to an immune
checkpoint protein may also be made using phage display technology
as is well known in the art.
Administration of Immune Checkpoint Inhibitor
[0253] Administration of immune checkpoint inhibitor may involve
administration at regular intervals, e.g. weekly, fortnightly, or
once every three or four weeks. For example, doses may be given at
regular, defined, intervals over a period of one of at least 1, 2,
3, 4, 5, 6, 7, 8, weeks or 1, 2, 3, 4, 5 or 6 months.
[0254] As such, multiple doses of immune checkpoint inhibitor may
be administered. For example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more
doses of immune checkpoint inhibitor may be administered to a
subject as part of a course of treatment. In some preferred
embodiments 1 or 2 doses, optionally 3 or more doses, of immune
checkpoint inhibitor are administered to the subject, preferably at
regular intervals (e.g. weekly, fortnightly, or once every three or
four weeks). Each dose is preferably administered within a single
day, e.g. over a period of 1, 2, 3, 4, 5, or 6 hours, and
optionally at the same time as a dose of oncolytic herpes simplex
virus.
[0255] Doses of immune checkpoint inhibitor may be separated by a
predetermined time interval, which may be selected to be one of 1,
2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or 31 days, or 1, 2, 3, 4,
5, or 6 months. By way of example, doses may be given once every 7,
14, 21 or 28 days (plus or minus 3, 2, or 1 days). The dose of
immune checkpoint inhibitor given at each dosing point may be the
same, but this is not essential. For example, it may be appropriate
to give a higher priming dose at the first, second and/or third
dosing points.
[0256] Administration of immune checkpoint inhibitor may be of one
or more treatment cycles, e.g. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or
more treatment cycles. A subject receiving multiple treatment
cycles may be given subsequent treatment cycles consecutively,
without a break from treatment, or may separate all or selected
treatment cycles by a break from treatment, e.g. a break of 1, 2,
3, 4, 5, 6, 7, 8 or 9 days or about 1, 2, 3, or 4 weeks.
[0257] In some embodiments a treatment cycle may comprise, or
consist of, 1 or 2 doses of immune checkpoint inhibitor
administered per period of about 3 or about 4 weeks. In some
embodiments a treatment cycle may comprise, or consist of, one dose
of immune checkpoint inhibitor administered per three week period.
Doses may be separated by 21.+-.3, 21.+-.2 or 21.+-.1 days. For
example, doses may be given on days 1, 22, 43 etc.
[0258] A treatment cycle of immune checkpoint inhibitor may be
given in conjunction with a treatment cycle of oncolytic herpes
simplex virus to provide a combined treatment. The treatment cycles
are not required to commence on the same day, although they may.
For example, a treatment cycle of oncolytic herpes simplex virus
may commence on day 1 and a treatment cycle of immune checkpoint
inhibitor may commence on day 8, or on any of days 1, 2, 3, 4, 5,
6, 7, 8, 9, 10, 11, 12, 13, or 14. As such a treatment cycle of
immune checkpoint inhibitor may commence after a treatment cycle of
oncolytic herpes simplex virus (e.g. about a week after) or before
a treatment cycle of oncolytic herpes simplex virus (e.g. about a
week before). Preferably both treatment cycles will have a duration
that causes them to overlap, e.g. by at least one day or more
preferably by about one or about two weeks.
[0259] Subjects may receive the same dosage of immune checkpoint
inhibitor at each administration within a given treatment cycle,
e.g. a dosage of 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350
mg or 400 mg. In some embodiments the first 1, 2 or 3 treatment
cycles may comprise administration of a lower dosage amount at each
administration, e.g. 150 mg, and later treatment cycles may
comprise administration of a higher dosage amount at each
administration, e.g. 200 mg.
[0260] Administration of immune checkpoint inhibitor may be by
infusion to the blood (intravenous or intra-arterial) and subjects
will preferably attend clinic on the scheduled administration days
for administration of immune checkpoint inhibitor. Administration
of immune checkpoint inhibitor may continue until development of
severe toxicity or withdrawal of consent.
[0261] In general, administration is preferably in a "effective
amount", this being sufficient to induce a treatment effect in the
individual. The actual amount administered, and rate and
time-course of administration, will depend on the nature and
severity of the disease being treated. Prescription of treatment,
e.g. decisions on dosage etc, is within the responsibility of
general practitioners and other medical doctors, and typically
takes account of the disorder to be treated, the condition of the
individual patient, the site of delivery, the method of
administration and other factors known to practitioners. Examples
of the techniques and protocols mentioned above can be found in
Remington's Pharmaceutical Sciences, 20th Edition, 2000, pub.
Lippincott, Williams & Wilkins.
Two Stage Programme of Treatment
[0262] In some aspects and embodiments of the present invention a
subject may initially receive one dose, preferably several doses,
of herpes simplex virus, preferably by intravenous administration,
without receiving an immune checkpoint inhibitor. The subject's
response to treatment with herpes simplex virus is monitored and
based on the determined response the subject may be selected for
treatment with an immune checkpoint inhibitor, preferably in
addition to continued treatment with herpes simplex virus--the
subject thereby progressing to a further stage of treatment in
which they receive combined treatment with herpes simplex virus and
an immune checkpoint inhibitor.
[0263] The response to initial treatment with herpes simplex virus
may involve determining whether the herpes simplex virus has
reached the cancer. This may involve detecting herpes simplex
virus, or fragments of herpes simplex virus such as HSV DNA, HSV
proteins of HSV antigens, in a blood sample or in a tumor tissue
sample.
[0264] Additionally, or alternatively, the response to initial
treatment with herpes simplex virus may involve determining whether
a Th1 response has been induced in the cancer. Detection of a Th1
response may be indicative of the herpes simplex virus successfully
reaching the tumor and initiating an anti-cancer response, which
may involve lysis of tumor cells by the herpes simplex virus and/or
induction of a host anti-cancer immune response. This may involve
detecting a Th1 response in a blood sample or in a tumor tissue
sample.
[0265] In cases where a determination is made that the herpes
simplex virus has reached the cancer or has induced a Th1 response,
the subject may be selected for a further stage of treatment with
an immune checkpoint inhibitor. In this stage, the subject
preferably continues treatment with herpes simplex virus and
treatment with immune checkpoint inhibitor is provided as an
additional treatment. The subject thereby receiving combined
treatment with herpes simplex virus and immune checkpoint
inhibitor.
[0266] Subjects selected for the initial stage of treatment with
herpes simplex virus may be indicated for surgical removal of tumor
tissue (referred to herein as `tumor resection`). For example, they
may have a cancer considered, by a medical practitioner, operable
to remove some or all of the tumor tissue. Treatment with herpes
simplex virus may commence prior to tumor resection.
[0267] Tumor tissue removed during tumor resection may be analysed
to determine whether herpes simplex virus has reached the tumor,
e.g. by detecting in vitro the presence of HSV DNA in excised tumor
tissue and/or to determine whether a Th1 response is present in the
cancer, e.g. by detecting in vitro the presence of a Th1 response
in excised tumor tissue.
[0268] A determination that herpes simplex virus has reached the
cancer and/or that a Th1 response has been induced in the cancer
may be indicative of the herpes simplex virus successfully reaching
the tumor and initiating an anti-cancer response, which may involve
lysis of tumor cells by the herpes simplex virus and/or induction
of a host anti-cancer immune response. In such cases, the subject
may be selected for a further stage of treatment with an immune
checkpoint inhibitor. In this stage, the subject preferably
continues treatment with herpes simplex virus and treatment with
immune checkpoint inhibitor is provided as an additional treatment.
The subject thereby receiving combined treatment with herpes
simplex virus and immune checkpoint inhibitor.
[0269] Accordingly, in one aspect of the present invention a herpes
simplex virus or immune checkpoint inhibitor is provided for use in
a method for the treatment of cancer, the method comprising: [0270]
(a) intravenous administration to a subject having cancer of one or
more doses of a herpes simplex virus, wherein the herpes simplex
virus lacks functional ICP34.5 genes; [0271] (b) determining,
preferably by in vitro analysis of a blood sample or of a tumor
tissue sample, one or both of: (i) whether the herpes simplex virus
has reached the tumor; and/or (ii) the presence of a Th1 response;
and [0272] (c) administering to a subject in which one or both of
(i) herpes simplex virus has reached the tumor; and/or (ii) a Th1
response is present, a herpes simplex virus and an immune
checkpoint inhibitor.
[0273] In another aspect of the present invention the use of a
herpes simplex virus or an immune checkpoint inhibitor in the
manufacture of a medicament is provided for use in a method for the
treatment of cancer, the method comprising: [0274] (a) intravenous
administration to a subject having cancer of one or more doses of a
herpes simplex virus, wherein the herpes simplex virus lacks
functional ICP34.5 genes; [0275] (b) determining, preferably by in
vitro analysis of a blood sample or tumor tissue sample, one or
both of: (i) whether the herpes simplex virus has reached the
tumor; and/or (ii) the presence of a Th1 response; and [0276] (c)
administering to a subject in which one or both of (i) herpes
simplex virus has reached the tumor; and/or (ii) a Th1 response is
present, a herpes simplex virus and an immune checkpoint
inhibitor.
[0277] In another aspect of the present invention a method for the
treatment of cancer is provided, the method comprising: [0278] (a)
intravenous administration to a subject having cancer of one or
more doses of a herpes simplex virus, wherein the herpes simplex
virus lacks functional ICP34.5 genes; [0279] (b) determining,
preferably by in vitro analysis of a blood sample or tumor tissue
sample, one or both of: (i) whether the herpes simplex virus has
reached the tumor; and/or (ii) the presence of a Th1 response; and
[0280] (c) administering to a subject in which one or both of (i)
herpes simplex virus has reached the tumor; and/or (ii) a Th1
response is present, a herpes simplex virus and an immune
checkpoint inhibitor.
[0281] In (a) the subject may be a subject indicated for surgical
removal of some or all of the tumor, and administration of the
herpes simplex virus is prior to said surgery. In (b) the tumor
tissue sample may be a sample of tumor tissue removed during
surgery following intravenous administration of a herpes simplex
virus according to (a).
[0282] In some embodiments methods according to the present
invention comprise administering one or more doses of herpes
simplex virus to the subject, determining the presence of a Th1
immune response in the subject, administering an immune checkpoint
inhibitor to a subject in which a Th1 immune response is induced or
elicited in combination with said herpes simplex virus.
[0283] In some embodiments methods according to the present
invention comprise administering one or more doses of herpes
simplex virus to the subject, determining whether herpes simplex
virus has reached the tumor, administering an immune checkpoint
inhibitor to a subject in which herpes simplex virus has reached
the tumor in combination with said herpes simplex virus. In some
embodiments determining whether herpes simplex virus has reached
the tumor may comprise determining the presence of HSV DNA in
tissue of the cancer, e.g. in a tumor tissue sample which may have
been obtained during surgical removal of some or all of the tumor
tissue.
[0284] In some embodiments the subject may receive administration
of a plurality of doses of herpes simplex virus. The doses of
herpes simplex virus may be administered in two stages, each stage
may involve administration of one or a plurality of doses.
[0285] Where a subject has undergone surgery to remove some or all
of the tumor tissue the timing of the next dose of herpes simplex
virus may be varied to accommodate the condition of the subject and
may, for example, be within 14 days of the day of surgery.
[0286] Where surgery is indicated, administration of herpes simplex
virus may commence prior to the subject undergoing surgery for
removal of tumor tissue, and preferably prior to receiving
treatment with immune checkpoint inhibitor. The subject may not
receive treatment with the immune checkpoint inhibitor, in
particular as a combined treatment with herpes simplex virus, until
after surgery for removal, and preferably analysis, of tumor
tissue.
[0287] In a first stage, the subject does not receive treatment
with an immune checkpoint inhibitor, although the subject may
receive concomitant chemotherapy or radiation therapy, e.g. as part
of standard of care treatment. Preferably, this stage takes place
prior to determining, whether the herpes simplex virus has reached
the tumor, and/or whether a Th1 response has been induced.
[0288] The subject response to treatment of herpes simplex virus in
the first stage is thereby monitored to determine whether treatment
with an immune checkpoint inhibitor as an additional agent will be
helpful. Surgical removal of tumor tissue or the taking of a blood
sample, followed by determination of the subject response to herpes
simplex virus monotherapy may form a decision point in terms of
selecting a subject for entry into a second stage of treatment.
[0289] Where a subject is selected for treatment with an immune
checkpoint inhibitor in a second stage of treatment the subject
will preferably continue to receive treatment with herpes simplex
virus, preferably receiving the herpes simplex virus by intravenous
administration.
[0290] The dosing schedule and amount of herpes simplex virus may
continue unchanged from the first to second stage, or may be
changed as considered necessary by the responsible medical
practitioner.
[0291] In another aspect of the present invention a method for
selecting patients with cancer for treatment with an immune
checkpoint inhibitor and a herpes simplex virus is provided, the
method comprising: [0292] intravenous administration of a herpes
simplex virus to a subject having cancer, wherein the herpes
simplex virus lacks functional ICP34.5 genes; [0293] determining,
preferably in vitro in a sample of blood or tumor tissue obtained
from the subject, one or both of: (i) whether the herpes simplex
virus has reached the tumor; and/or (ii) the presence of a Th1
response; and [0294] selecting a subject in which one or both of
(i) herpes simplex virus has reached the tumor; and/or (ii) a Th1
response is present, for treatment with an immune checkpoint
inhibitor in combination with said herpes simplex virus.
[0295] In another aspect of the present invention a method for
selecting patients with cancer for treatment with an immune
checkpoint inhibitor and a herpes simplex virus is provided, the
method comprising: [0296] intravenous administration of a herpes
simplex virus to a subject having cancer, the subject being
indicated for surgical removal of some or all of the tumor and the
intravenous administration of herpes simplex virus being prior to
the surgery, wherein the herpes simplex virus lacks functional
ICP34.5 genes; [0297] determining, preferably in vitro, one or both
of: (i) whether the herpes simplex virus has reached the tumor;
and/or (ii) the presence of a Th1 response in tumor tissue removed
during the surgery; and [0298] selecting a subject in which one or
both of (i) herpes simplex virus has reached the tumor; and/or (ii)
a Th1 response is present in tumor tissue removed during the
surgery, for treatment with an immune checkpoint inhibitor in
combination with said herpes simplex virus.
[0299] In some embodiments determining whether herpes simplex virus
has reached the tumor may comprise determining, in vitro, the
presence of HSV DNA in a blood sample or tumor tissue sample, e.g.
tumor tissue sample removed during surgery.
[0300] In some embodiments the subject has been treated with herpes
simplex virus and is, or has been, selected for treatment with the
immune checkpoint inhibitor as having a Th1 immune response.
[0301] In some embodiments methods according to the present
invention may comprise administering one or more doses of the
herpes simplex virus effective to induce a Th1 immune response.
[0302] Accordingly, in some embodiments, the method comprises
administering one or more doses of a herpes simplex virus effective
to induce a Th1 immune response in the subject, and administering a
therapeutically effective amount of an immune checkpoint
inhibitor.
[0303] In some aspects of the present invention a method of
treating cancer in a subject in need of treatment is provided, the
method comprising administering one or more doses of a herpes
simplex virus to a subject, and administering to a subject in which
a Th1 immune response is induced or elicited a therapeutically
effective amount of an immune checkpoint inhibitor.
[0304] In some embodiments the method may comprise the step of
determining the presence of a Th1 immune response in the subject,
optionally prior to administration of the immune checkpoint
inhibitor.
[0305] In some embodiments the method may comprise the step of
selecting a subject in which a Th1 immune response is induced or
elicited by the herpes simplex virus for treatment with the immune
checkpoint inhibitor, said selection preferably being made prior to
administration of the immune checkpoint inhibitor.
[0306] In some embodiments the method comprises administering one
or more doses of a herpes simplex virus to the subject, determining
the presence of a Th1 immune response in the subject, administering
to a subject in which a Th1 immune response is induced or elicited
a therapeutically effective amount of an immune checkpoint
inhibitor.
[0307] In some embodiments the method comprises administering one
or more doses of a herpes simplex virus to the subject, selecting a
subject in which a Th1 immune response is induced or elicited for
treatment with an immune checkpoint inhibitor, and administering to
a selected subject a therapeutically effective amount of an immune
checkpoint inhibitor.
[0308] In some embodiments the method comprises selecting a subject
in which a Th1 immune response is induced or elicited by the herpes
simplex virus for treatment with the immune checkpoint inhibitor.
In some embodiments administration of the herpes simplex virus is
for a period of time sufficient to induce or elicit a Th1 response
in the subject.
[0309] In some embodiments the method comprises selecting a subject
in which herpes simplex virus administered in a first stage of
treatment has reached the tumor for treatment with the immune
checkpoint inhibitor. In some embodiments administration of the
herpes simplex virus is for a period of time sufficient to allow
the herpes simplex virus to reach the tumor.
[0310] In some embodiments administration of the herpes simplex
virus is for a period of time prior to treatment with an immune
checkpoint inhibitor in which period the subject is administered
herpes simplex virus but is not administered an immune checkpoint
inhibitor.
Determining Whether Herpes Simplex Virus has Reached the Tumor
[0311] In some embodiments a determination is made as to whether
herpes simplex virus administered to the subject has reached the
site of a tumor. This is of particular relevance where the herpes
simplex virus is administered by intravenous administration as the
site of administration will be distal from the tumor and the herpes
simplex virus must reach the tumor and preferably be able to
replicate in the tumor.
[0312] The determination may be made by detecting intact herpes
simplex virus, or components or fragments of herpes simplex virus
(e.g. HSV DNA, HSV proteins or HSV antigens), in a sample obtained
from the subject, preferably in either a blood sample or tumor
tissue sample.
[0313] Intact herpes simplex virus may be detected by viral culture
of a sample, e.g. shell vial culture (e.g. see Tse et al., J Clin
Microbiol. 1989 Jan. 27(1): 199-200).
[0314] HSV DNA may be detected by routine techniques such as qPCR
or Real-Time PCR (e.g. see Kessler et al., J Clin Microbiol. 2000
July; 38(7):2638-2642; Pandori et al BMC Infectious Diseases 2006
6:104; Hong et al., BioMed Research International Vol 2014 (2014)
Article ID 261947, 5 pages).
[0315] In some embodiments samples may be collected at selected
intervals, e.g. before treatment with herpes simplex virus and at
selected intervals after treatment has commenced, to monitor the
detection of herpes simplex virus, components or fragments of
herpes simplex virus.
[0316] For example, samples, e.g. blood samples, may be collected
after treatment with herpes simplex virus has commenced at one of
about 24 hours, about 48 hours, about 72 hours, about 96 hours,
about 120 hours, about 144 hours, about one week, about two weeks,
about three weeks or about four weeks. The level of HSV DNA in the
sample may be detected.
[0317] Such monitoring may show an inability to detect, or
detection of only low levels of, herpes simplex virus, components
or fragments thereof, immediately after commencement of treatment
and/or during initial stages of treatment followed by ability to
detect herpes simplex virus, components or fragments thereof as
treatment progresses. This re-emergence of detection of herpes
simplex virus, components or fragments thereof may indicate that
the herpes simplex virus has reached the tumor, and optionally that
it is replicating in the tumor.
[0318] Accordingly, in some embodiments the determination may be
made after passage of a period of time from administration of
herpes simplex virus sufficient for the virus or viral fragments or
components to be detectable in a sample, e.g. a blood or tumor
tissue sample.
Detecting a Th1 Immune Response
[0319] A Th1 immune response may be characterised by upregulation
of one or more relevant cytokines selected from the group
consisting of IFN-.gamma., IL-2, IL-12, IP-10, MIG, TNF-.alpha.,
IL-18, IL-27, TNF-.beta.. In some embodiments a Th1 response may be
characterised by upregulation of IFN-.gamma., IL-2, or IL-12.
[0320] Not all subjects respond to administration of oncolytic HSV
by induction of a Th1 response and subjects may therefore be
monitored to determine their response to treatment with the
oncolytic HSV. Subjects who respond may continue treatment with
oncolytic HSV at a dose and/or dosage regime designed to maintain
the sustained Th1 response. Subjects who do not respond to
treatment by generation of a Th1 response may discontinue treatment
with oncolytic HSV, or may transition to combination treatment with
oncolytic HSV and one or more additional chemotherapeutic
agents.
[0321] Accordingly, the presence of a Th1 response may be
determined by measuring the level of a Th1 cytokine in a sample
obtained from a subject.
[0322] In addition to the cytokines secreted by Th1 cells, the
expression of specific cell surface proteins or receptors,
including IL-12 R beta 2, IL-27 R alpha/VVSX-1, IFN-gamma R2, CCR5,
and CXCR3, can be used to distinguish Th1 cells from other T cell
subtypes. T cells associated with a Th1 response can also be
partially characterised as CD4.sup.+ or CD8.sup.+. Accordingly, the
presence of a Th1 response may also be determined or partially
determined by measuring the expression of Th1 cell surface proteins
or receptors by T cells in a sample obtained from a subject and/or
by measuring the proportion of T cells that are CD4.sup.+ or
CD8.sup.+ in a sample obtained from a subject.
[0323] In some preferred embodiments a Th1 response may be
characterised by upregulation of IFN-.gamma.. In some preferred
embodiments a Th1 response may be characterised by upregulation of
IL-2. In some preferred embodiments a Th1 response may be
characterised by upregulation of IL-12. In some preferred
embodiments a Th1 response may be characterised by upregulation of
IFN-.gamma. and determination of T cell CD8.sup.+ status. In some
preferred embodiments a Th1 response may be characterised by
upregulation of IL-2 and determination of T cell CD8.sup.+ status.
In some preferred embodiments a Th1 response may be characterised
by upregulation of IL-12 and determination of T cell CD8.sup.+
status. In some preferred embodiments a Th1 response may be
characterised by upregulation of IFN-.gamma. and one or both of
IL-2 and IL-12, and optionally determination of T cell CD8.sup.+
status. In some preferred embodiments a Th1 response may be
characterised by upregulation of IL-2 and one or both of
IFN-.gamma. and IL-12, and optionally determination of T cell
CD8.sup.+ status. In some preferred embodiments a Th1 response may
be characterised by upregulation of IL-12 and one or both of
IFN-.gamma. and IL-2, and optionally determination of T cell
CD8.sup.+ status. In some preferred embodiments a Th1 response may
be characterised by upregulation of IL-2 and/or IL-12.
[0324] The detection and characterisation of a Th1 immune response
is a routine procedure for one of ordinary skill in the art.
Numerous cytokine and cell detection assays are available including
single and multiplexed ELISAs, reverse-transcription-PCR, Taqman
real-time PCR, immunohistochemistry and flow cytometry (e.g. see
Whiteside T L. Cytokine assays. Biotechniques 2002; Suppl:4-8, 10,
12-5; Pala P, Hussell T, Openshaw P J. Flow cytometric measurement
of intracellular cytokines. J Immunol Methods 2000; 243:107-24;
Jason J, Lamed J. Single-cell cytokine profiles in normal humans:
comparison of flow cytometric reagents and stimulation protocols. J
Immunol Methods 1997; 207:13-22; Farrell A M et al. A rapid flow
cytometric assay to detect CD4+ and CD8+ T-helper (Th) 0, Th1 and
Th2 cells in whole blood and its application to study cytokine
levels in atopic dermatitis before and after cyclosporin therapy.
Br J Dermatol. 2001 January; 144(1):24-33).
[0325] The induction of a Th1 response and maintenance of the
response over time may therefore be monitored at desired intervals
by analysing the level of cytokine expression in a sample taken
from a subject. Such monitoring may be conducted, daily, weekly,
fortnightly, monthly or yearly as desired and/or consider
appropriate by a medical practitioner.
[0326] In some embodiments detection of a Th1 response involves
detection of an upregulation of a Th1 cell population or of Th1
cytokines.
[0327] Upregulation may be determined by comparing the level of a
cell population or cytokine before and after a treatment, e.g.
before and then after a period of treatment (optionally
pre-treatment) with an herpes simplex virus. Levels of a cell
population or of a cytokine may be quantitated for absolute
comparison, or relative comparisons may be made.
[0328] In some embodiments upregulation may be considered to be
present when the level of a cell population or cytokine in the test
sample is at least 1.1 times that in the control sample (e.g. a
sample obtained before treatment with herpes simplex virus). More
preferably, the level may be selected from one of at least 1.2, at
least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7,
at least 1.8, at least 1.9, at least 2.0, at least 2.1, at least
2.2, at least 2.3, at least 2.4 at least 2.5, at least 2.6, at
least 2.7, at least 2.8, at least 2.9, at least 3.0, at least 3.5,
at least 4.0, at least 5.0, at least 6.0, at least 7.0, at least
8.0, at least 9.0, at least 10.0, at least 11.0, at least 12.0, at
least 13.0, at least 14.0, or at least 15.0 times that in the
control sample.
[0329] Detection of a Th1 response may be further confirmed by
detection of an anti-tumor IgG response. This may be determined,
for example, by immunoassay to detect the presence of IgG in a
sample from a subject having received treatment with oncolytic
herpes simplex virus. Comparison may be made to the absence of such
a response in a patient not having received treatment with herpes
simplex virus.
[0330] In addition to detection of a Th1 response, the presence of
HSV DNA may be detected in a sample. HSV DNA may be detected by
routine techniques such as qPCR or Real-Time PCR (e.g. see Kessler
et al., J Clin Microbiol. 2000 July; 38(7):2638-2642; Pandori et al
BMC Infectious Diseases 2006 6:104; Hong et al., BioMed Research
International Vol 2014 (2014) Article ID 261947, 5 pages).
Sample
[0331] A sample may be obtained from a subject. Samples may be
obtained before, during and/or after treatment with an herpes
simplex virus. A sample obtained before treatment has commenced may
provide reference values, e.g. for cytokine levels, cell surface
proteins or receptors or proportion of CD4.sup.+ or CD8+ T cells,
which may allow for comparison with levels determined during or
after treatment, the comparison enabling a determination of whether
herpes simplex virus has reached the tumor or a Th1 response has
been induced or elicited in the subject.
[0332] A sample may be taken or derived from any tissue, e.g. tumor
tissue, or bodily fluid, and may be processed to isolate a cell
population of interest, e.g. white blood cells, lymphocytes or T
cells.
[0333] A sample may comprise or may be derived from: a quantity of
blood; a quantity of serum derived from the individual's blood
which may comprise the fluid portion of the blood obtained after
removal of the fibrin clot and blood cells; saliva; other bodily
fluids, pleural fluid, effusion fluid, ascites, or a fluid produced
by a cancer.
[0334] In preferred arrangements the sample is taken from a bodily
fluid, more preferably one that circulates through the body.
Accordingly, the sample may be a blood sample or lymph sample.
[0335] In some embodiments the sample is a blood sample or
blood-derived sample. The blood derived sample may be a selected
fraction of a patient's blood, e.g. a selected cell-containing
fraction or a plasma or serum fraction. A selected cell-containing
fraction may contain cell types of interest which may include white
blood cells (WBC), lymphocytes, peripheral blood mononuclear cells
(PBC) and/or granulocytes, and/or red blood cells (RBC).
[0336] In some embodiments the sample is a biopsy, or is derived
from a biopsy. In some embodiments the sample is a tumor tissue
sample, e.g. obtained during surgery to remove tumor tissue. In
some embodiments the sample may be obtained during surgical
resection of a tumor. Solid tumors are particularly suitable for
obtaining samples by biopsy or during surgical resection.
[0337] Subjects selected for treatment with herpes simplex virus
may be indicated for surgical removal of tumor tissue (referred to
herein as `tumor resection`). For example, they may have a cancer
considered, by a medical practitioner, operable to remove some or
all of the tumor tissue. Treatment with herpes simplex virus may
commence prior to tumor resection.
[0338] Tumor tissue removed during tumor resection may be analysed
to determine whether herpes simplex virus has reached the tumor,
e.g. by detecting in vitro the presence of HSV DNA in excised tumor
tissue and/or the expression of HSV antigens by
immunohistochemistry and/or to determine whether a Th1 response is
present in the cancer, e.g. by detecting in vitro the presence of a
Th1 response in excised tumor tissue.
[0339] In some embodiments the sample is a sample of effusion
fluid, e.g. pleural fluid or ascites. Effusion fluid refers to an
excess of fluid produced by a subject in direct or indirect
response to the presence of a cancer in the subject. Effusion fluid
may collect in a body cavity such that accumulation of effusion
fluid may occur where the rate of production of the effusion fluid
exceeds the rate of reabsorption. Pleural effusions (sometimes
called malignant pleural effusions) lead to accumulation of fluid
in the pleural cavity and occur in some lung cancers, e.g.
mesothelioma. Effusion fluid collecting in the peritoneal cavity is
commonly referred to as ascites, and can be a symptom of a number
of types of cancer including cancer of the breast, lung, colon,
stomach, pancreas, ovary, endometrium as well as lymphoma.
Pericardial effusion is the abnormal accumulation of fluid in the
pericardial cavity. The effusion fluid is preferably exudative.
[0340] Effusion fluid can be drained from a respective body cavity
by well-known aseptic procedures (e.g. see Warren et al. Ann Thorac
Surg 2008; 85:1049-55; Warren et al. European Journal of
Cardio-thoracic Surgery 33 (2008) 89-94). In some instances, a tube
or catheter is inserted in the body cavity in order to drain
effusion fluid. Drainage of effusion fluid is a common part of the
diagnosis, treatment and management of many forms of cancer.
Drainage of effusion fluid provides a means of obtaining a sample
of a subject's effusion fluid for analysis.
Cancer
[0341] A cancer may be any unwanted cell proliferation (or any
disease manifesting itself by unwanted cell proliferation),
neoplasm or tumor or increased risk of or predisposition to the
unwanted cell proliferation, neoplasm or tumor. The cancer may be
benign or malignant and may be primary or secondary (metastatic). A
neoplasm or tumor may be any abnormal growth or proliferation of
cells and may be located in any tissue. Examples of tissues include
the adrenal gland, adrenal medulla, anus, appendix, bladder, blood,
bone, bone marrow, brain, breast, cecum, central nervous system
(including or excluding the brain) cerebellum, cervix, colon,
duodenum, endometrium, epithelial cells (e.g. renal epithelia),
gallbladder, oesophagus, glial cells, heart, ileum, jejunum,
kidney, lacrimal glad, larynx, liver, lung, lymph, lymph node,
lymphoblast, maxilla, mediastinum, mesentery, myometrium,
nasopharynx, omentume, oral cavity, ovary, pancreas, parotid gland,
peripheral nervous system, peritoneum, pleura, prostate, salivary
gland, sigmoid colon, skin, small intestine, soft tissues, spleen,
stomach, testis, thymus, thyroid gland, tongue, tonsil, trachea,
uterus, vulva, white blood cells.
[0342] Tumors to be treated may be nervous or non-nervous system
tumors. Nervous system tumors may originate either in the central
or peripheral nervous system, e.g. glioma, medulloblastoma,
meningioma, neurofibroma, ependymoma, Schwannoma,
neurofibrosarcoma, astrocytoma and oligodendroglioma. Non-nervous
system cancers/tumors may originate in any other non-nervous
tissue, examples include melanoma, mesothelioma, lymphoma, myeloma,
leukemia, Non-Hodgkin's lymphoma (NHL), Hodgkin's lymphoma, chronic
myelogenous leukemia (CML), acute myeloid leukemia (AML),
myelodysplastic syndrome (MDS), cutaneous T-cell lymphoma (CTCL),
chronic lymphocytic leukemia (CLL), hepatoma, epidermoid carcinoma,
prostate carcinoma, breast cancer, lung cancer, colon cancer,
ovarian cancer, pancreatic cancer, thymic carcinoma, NSCLC,
haematologic cancer and sarcoma.
[0343] In the context of treatment of a metastatic cancer,
treatment may be of cancers or tumors of a given cell type. The
treatment may involve eliciting a systemic anti-tumor Th1 immune
response in the subject, who may be at risk of developing single or
multiple metastatic cancers or tumors of the given cell type.
Administration of oncolytic herpes simplex virus may therefore
induce a Th1 immune response that is specific for the tumor cell
type and that kills cells of inoculated tumor and non-inoculated
tumors.
[0344] In some embodiments the cancer may be a solid tumor. Solid
tumors may, for example, be in bladder, bone, breast, eye, stomach,
head and neck, germ cell, kidney, liver, lung, nervous tissue,
ovary, pancreas, prostate skin, soft-tissues, adrenal gland,
nasopharynx, thyroid, retina, and uterus. Solid tumors may include
melanoma, rhabdomyosarcoma, Ewing sarcoma, and neuroblastoma.
[0345] The cancer may be a pediatric solid tumor, i.e. solid tumor
in a child, for example osteosarcoma, chondroblastoma,
chondrosarcoma, Ewing sarcoma, malignant germ cell tumor, Wilms
tumor, malignant rhabdoid tumor, hepatoblastoma, hepatocellular
carcinoma, neuroblastoma, melanoma, adrenocorticoid carcinoma,
nasopharyngeal carcinoma, thyroid carcinoma, retinoblastoma,
soft-tissue sarcoma, rhabdomyosarcoma, desmoid tumor, fibrosarcoma,
liposarcoma, malignant fibrous histiocytoma, neurofibrosarcoma.
[0346] The cancer may be in a location that would require surgery
or an invasive procedure in order to administer the virus by
intratumoral injection. For example, the cancer may be a visceral
cancer or visceral tumor, e.g. a cancer of the internal organs of
the body such as within the chest (e.g. heart, lungs), abdomen,
(e.g. liver, pancreas, stomach, intestines), body cavities (e.g.
pleura, peritoneum) or brain. Cancers to be treated may be visceral
lesions, e.g. metastatic visceral lesions.
[0347] In some embodiments the cancer is a head and neck cancer. In
some optional embodiments the cancer is not a head and neck
cancer.
[0348] In some preferred embodiments the cancer may be a
mesothelioma, e.g. a malignant pleural mesothelioma.
[0349] The cancer may be one that is associated with effusion
fluid. Such association may involve production of effusion fluid by
the cancerous tissue, e.g. by cancer cells, or by normal cells near
to or contained in the cancerous tissue, or it may involve
overproduction of effusion fluid by other tissues (e.g. the
lymphatic system) as a direct or indirect response to the presence
of the cancer in the subject.
[0350] The cancer may be characterised by the collection of
effusion fluid in one or more locations in the subject's body. Such
locations may include one or more body cavities or tissue spaces.
Body cavities (or serous cavities) may be formed by a serous
membrane surrounding an organ or tissue and forming a sac in which
fluid may collect.
[0351] For example, effusion fluid may collect in one or each
(right or left) pleural cavity (the space between the visceral and
parietal pleura). In another example, effusion fluid may accumulate
in the peritoneal cavity (the space between the parietal peritoneum
and visceral peritoneum). In another example, fluid may accumulate
in the pericardial cavity surrounding the heart (formed by the
parietal and visceral pericardium). In another example, fluid may
accumulate in the perimetrium surrounding the uterus.
[0352] Thus, in some embodiments the cancer is one in which pleural
effusion, peritoneal effusion (ascites), pericardial effusion or
perimetrial effusion occurs.
[0353] All types of cancer may be associated with production of
effusion fluid, partly because all types of cancer can metastasize
to any of the body's serous cavities and result in malignant
effusion (Olopade CA-A Cancer Journal For Clinicians Vol. 41, No. 3
May/June 1991). Cancers in which production of effusion fluid is
known to occur include cancers of the following type or tissues:
lung cancers, pleural cancers, mesothelioma, malignant pleural
mesothelioma, non-small cell lung cancer (NSCLC), small cell lung
cancer (SCLC), ovarian cancers, ovarian carcinoma, uterine cancer,
endometrial cancer, heart cancer, breast cancer, colon cancer,
stomach cancer, gastric cancer, pancreatic cancer, kidney cancer,
liver cancer, lymphatic cancer (e.g. lymphoma, non-Hodgkin
lymphoma), soft tissue sarcoma, osteosarcoma, adenocarcinoma,
parotid cancer (e.g. parotid adenocarcinoma), thymic carcinoma,
cancers of the reproductive tract (including cervical, fallopian
tube, endometrium), gastrointestinal tract, or genitourinary tract,
leukemia, larynx, prostate, bile duct, hypernephroma, sinus
piriformis carcinoma, thyroid cancer, melanoma and cancers of
unknown primary (CUP) origin.
[0354] The development of a malignant pleural effusion is a common
complication of advanced malignancies of many types of cancer,
especially breast, lung (including NSCLC and SCLC) and ovarian
carcinoma (Warren et al. European Journal of Cardio-thoracic
Surgery 33 (2008) 89-94). Pleural effusions are at least known to
be associated with cancers of the following type or tissue: lung,
breast, lymphoma, uterus, ovarian, female reproductive tract (e.g.
cervical, fallopian tube, endometrium), leukemia, pancreas, kidney,
colon, stomach (gastric), mesothelioma, sarcoma, larynx, prostate,
bile duct, hypernephroma, sinus piriformis carcinoma, thyroid
cancer, non-Hodgkin lymphoma, malignant melanoma, reproductive
tract, gastrointestinal tract, genitourinary tract, (Warren et al.
Ann Thorac Surg 2008; 85:1049-55; Warren et al. European Journal of
Cardio-thoracic Surgery 33 (2008) 89-94; Schulze et al. Ann Thorac
Surg 2001; 71:1809-12; Olopade CA-A Cancer Journal For Clinicians
Vol. 41, No. 3 May/June 1991).
[0355] Peritoneal effusions (ascites) are at least known to be
associated with cancers of the following type or tissue: ovarian,
epithelial related ovarian, uterus, breast, colon, gastric,
pancreatic, hepatic, colon, lymphoma, mesothelioma, and cancers of
unknown primary (CUP) origin (Olopade CA-A Cancer Journal For
Clinicians Vol. 41, No. 3 May/June 1991) Pericardial effusions are
at least known to be associated with cancers of the following type
or tissue: lung, breast, leukemia, lymphoma, sarcoma, melanoma
(Olopade CA-A Cancer Journal For Clinicians Vol. 41, No. 3 May/June
1991).
[0356] Optionally, in some preferred embodiments the cancer is not
a melanoma. Optionally, in some preferred embodiments the cancer is
not a cancer occurring in the skin. Optionally, in some preferred
embodiments the cancer is not a primary melanoma. Optionally, in
some preferred embodiments the cancer is not metastatic (secondary
melanoma). Optionally, in some embodiments the cancer is not stage
IIIb to stage IV melanoma.
Head and Neck Cancer
[0357] Some aspects and embodiments of the present invention
concern the use of herpes simplex virus to treat head and neck
cancer.
[0358] In some embodiments, the subject may receive a herpes
simplex virus and an immune checkpoint inhibitor as part of the
programme of treatment. During combined treatment, the herpes
simplex virus and immune checkpoint inhibitor may be administered
simultaneously, e.g. as a combined preparation or as separate
preparations one administered immediately after the other.
Alternatively, they may be administered separately and
sequentially, where one agent is administered and then the other
administered later after a predetermined time interval.
[0359] In one aspect of the present invention a herpes simplex
virus is provided for use in a method of treating head and neck
cancer, the method comprising administering to a subject having
head and neck cancer said herpes simplex virus and an immune
checkpoint inhibitor, wherein the herpes simplex virus lacks
functional ICP34.5 genes.
[0360] In another aspect of the present invention an immune
checkpoint inhibitor is provided for use in a method of treating
head and neck cancer, the method comprising administering to a
subject having head and neck cancer said immune checkpoint
inhibitor and a herpes simplex virus, wherein the herpes simplex
virus lacks functional ICP34.5 genes.
[0361] In another aspect of the present invention the use of a
herpes simplex virus in the manufacture of a medicament for use in
a method of treating head and neck cancer is provided, the method
comprising administering to a subject having head and neck cancer
said herpes simplex virus and an immune checkpoint inhibitor,
wherein the herpes simplex virus lacks functional ICP34.5
genes.
[0362] In another aspect of the present invention the use of an
immune checkpoint inhibitor in the manufacture of a medicament for
use in a method of treating head and neck cancer is provided, the
method comprising administering to a subject having head and neck
cancer said immune checkpoint inhibitor and a herpes simplex virus,
wherein the herpes simplex virus lacks functional ICP34.5
genes.
[0363] In another aspect of the present invention a method for the
treatment of head and neck cancer is provided, the method
comprising administering to a subject having head and neck cancer
an immune checkpoint inhibitor and a herpes simplex virus, wherein
the herpes simplex virus lacks functional ICP34.5 genes.
[0364] In preferred embodiments the herpes simplex virus is
administered to the subject systemically, preferably to the blood.
In a preferred embodiment administration is by intravenous
administration, e.g. by intravenous infusion.
[0365] In preferred embodiments the head and neck cancer is stage
III or stage IV.
[0366] In one aspect of the present invention method comprises
administering to a patient with stage III or stage IV head and neck
cancer a therapeutically effective amount of an immune checkpoint
inhibitor and an herpes simplex virus, wherein the herpes simplex
virus lacks functional ICP34.5 genes and the herpes simplex virus
is administered to the patient systemically by intravenous infusion
on one or more occasions.
[0367] In some embodiments the subject has a head and neck cancer
which is suitable for surgical removal of some or all of the tumor
tissue.
[0368] In some embodiments the subject may receive administration
of one or a plurality (preferably a plurality) of doses of herpes
simplex virus. Each dose of herpes simplex virus is preferably of
greater than 2.times.10.sup.6 iu. Doses may be in a range selected
from the group consisting of: 2.times.10.sup.6 to 9.times.10.sup.6
iu, 2.times.10.sup.6 to 5.times.10.sup.6 iu, 5.times.10.sup.6 to
9.times.10.sup.6 iu, 2.times.10.sup.6 to 1.times.10.sup.7 iu,
2.times.10.sup.6 to 5.times.10.sup.7 iu, 2.times.10.sup.6 to
1.times.10.sup.8 iu, 2.times.10.sup.6 to 5.times.10.sup.8 iu,
2.times.10.sup.6 to 1.times.10.sup.9 iu, 5.times.10.sup.6 to
1.times.10.sup.7 iu, 5.times.10.sup.6 to 5.times.10.sup.7 iu,
5.times.10.sup.6 to 1.times.10.sup.8 iu, 5.times.10.sup.6 to
5.times.10.sup.8 iu, 5.times.10.sup.6 to 1.times.10.sup.9 iu,
5.times.10.sup.6 to 5.times.10.sup.9 iu, 1.times.10.sup.7 to
9.times.10.sup.7 iu, 1.times.10.sup.7 to 5.times.10.sup.7 iu,
1.times.10.sup.8 to 9.times.10.sup.8 iu, 1.times.10.sup.8 to
5.times.10.sup.8 iu. In some embodiments suitable doses may be in
the range 2.times.10.sup.6 to 9.times.10.sup.6 iu, 1.times.10.sup.7
to 9.times.10.sup.7 iu, or 1.times.10.sup.8 to 9.times.10.sup.8 iu.
In some embodiments suitable doses may be about 1.times.10.sup.7 iu
or about 1.times.10.sup.8 iu. Dosage figures may optionally be +/-
half a log value.
[0369] Doses of herpes simplex virus are preferably administered by
intravenous infusion, which may take place over a period of several
hours, e.g. about 30 minutes to about 4 hours. A subject will
commonly receive a plurality of doses of herpes simplex virus as
part of a course of treatment, preferably 3 or more doses. The
doses may be administered in accordance with a dosing regime. For
example, each dose of herpes simplex virus may be administered
within 1 to 7, 1 to 14, or 1 to 21 days of the preceding dose.
[0370] Doses of herpes simplex virus may be administered at regular
intervals, e.g. every 7 days, every 14 days, every 21 days, or
every 28 days (+/-1, 2 or 3 days). The number of doses of herpes
simplex virus administered in a course of treatment may be any of
1, 2, 3, 4, 5, 6, 7, 8, 9, 10 doses or more, preferably at least 3
or at least 4 doses, or up to 8 doses.
[0371] A dosing regime for a herpes simplex virus may be designed
to continue dosing until: surgery is scheduled, toxicity or disease
progression, a predetermined maximum number of doses is reached,
e.g. 3, 4, 5, 6, 7, 8, 9 or 10 doses, or a fixed number of doses is
reached, e.g. 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 doses, complete
response, disease progression, or intolerance of the treatment.
[0372] The immune checkpoint inhibitor may be administered
intravenously. The subject may receive a single intravenous
administration of a combined preparation of herpes simplex virus
and immune checkpoint inhibitor, or may receive separate
intravenous administrations of herpes simplex virus and immune
checkpoint inhibitor. Herpes simplex virus and immune checkpoint
inhibitor may be administered on the same day, e.g. during the same
hospital visit, or on separate days.
[0373] A subject will preferably receive a plurality of doses of
immune checkpoint inhibitor during the course of treatment. The
doses may be administered in accordance with a dosing regime. For
example, each dose of immune checkpoint inhibitor may be
administered within 1 to 21 or 1 to 28 days of the preceding dose.
Doses of immune checkpoint inhibitor may be administered at regular
intervals, e.g. every 7 days, every 14 days, every 21 days, or
every 28 days (+/-1, 2 or 3 days).
[0374] Suitable doses of immune checkpoint inhibitor will vary
depending on the immune checkpoint inhibitor selected and the
prescribing information of the medical practitioner. By way of
example, suitable doses may be one of 100 mg, 200 mg, 300 mg or 400
mg, or 1 mg/kg, 2 mg/kg, 3 mg/kg or 4 mg/kg.
[0375] Doses of immune checkpoint inhibitor may be administered by
intravenous infusion, which may take place over a period of up to
several hours, e.g. about 30 minutes to about 4 hours.
[0376] By way of example, pembrolizumab may be administered by
intravenous infusion once every 3 weeks, optionally at a dose of
about 200 mg. In another example, pembrolizumab may be administered
by intravenous infusion once every week, optionally at a dose of
about 200 mg, and optionally in combination with chemotherapy, e.g.
in the form of platinum and/or 5-fluoruracil.
[0377] A combined dosing regime for a herpes simplex virus and
immune checkpoint inhibitor may be designed to continue dosing
until: toxicity or disease progression, a predetermined maximum
number of doses of one or both agents is reached, e.g. 3, 4, 5, 6,
7, 8, 9 or 10 doses, a fixed number of doses of one or both agents
is reached, e.g. 3, 4, 5, 6, 7, 8, 9, or 10 doses, complete
response, disease progression, or intolerance of the treatment.
[0378] In some embodiments herpes simplex virus may be administered
more regularly than the immune checkpoint inhibitor. Administration
of herpes simplex virus may optionally precede, i.e. commence
before, administration of the immune checkpoint inhibitor. For
example, herpes simplex virus may be administered once weekly,
whereas the immune checkpoint inhibitor may be administered once
every 2, 3 or 4 weeks, optionally commencing in the first, second
or third week of treatment. The dosing schedule may be designed
such that administration of herpes simplex virus is on the same day
as administration of the immune checkpoint inhibitor.
[0379] In another aspect of the present invention a herpes simplex
virus is provided for use in a method of treating head and neck
cancer, the method comprising administering: a herpes simplex virus
by intravenous infusion at a dose of greater than 2.times.10.sup.6
iu at day 1 of week 1 followed by a dose of greater than
2.times.10.sup.6 iu at day 1 of week 2, and every week thereafter
until surgery is scheduled, toxicity or disease progression, a
predetermined maximum number of doses is reached or a fixed number
of doses is reached, complete response, disease progression, or
intolerance of the treatment, wherein the herpes simplex virus
lacks functional ICP34.5 genes; and an immune checkpoint inhibitor
by intravenous infusion every 3 weeks for at least 3 infusions
beginning with or after the second or third dose of the herpes
simplex virus.
[0380] In another aspect of the present invention an immune
checkpoint inhibitor is provided for use in a method of treating
head and neck cancer, the method comprising administering: a herpes
simplex virus by intravenous infusion at a dose of greater than
2.times.10.sup.6 iu at day 1 of week 1 followed by a dose of
greater than 2.times.10.sup.6 iu at day 1 of week 2, and every week
thereafter until surgery is scheduled, toxicity or disease
progression, a predetermined maximum number of doses is reached or
a fixed number of doses is reached, complete response, disease
progression, or intolerance of the treatment, wherein the herpes
simplex virus lacks functional ICP34.5 genes; and an immune
checkpoint inhibitor by intravenous infusion every 3 weeks for at
least 3 infusions beginning with or after the second or third dose
of the herpes simplex virus.
[0381] In another aspect of the present invention the use of a
herpes simplex virus in the manufacture of a medicament for use in
a method of treating head and neck cancer is provided, the method
comprising administering: a herpes simplex virus by intravenous
infusion at a dose of greater than 2.times.10.sup.6 iu at day 1 of
week 1 followed by a dose of greater than 2.times.10.sup.6 iu at
day 1 of week 2, and every week thereafter until surgery is
scheduled, toxicity or disease progression, a predetermined maximum
number of doses is reached or a fixed number of doses is reached,
complete response, disease progression, or intolerance of the
treatment, wherein the herpes simplex virus lacks functional
ICP34.5 genes; and an immune checkpoint inhibitor by intravenous
infusion every 3 weeks for at least 3 infusions beginning with or
after the second or third dose of the herpes simplex virus.
[0382] In another aspect of the present invention the use of an
immune checkpoint inhibitor in the manufacture of a medicament for
use in a method of treating head and neck cancer is provided, the
method comprising administering: a herpes simplex virus by
intravenous infusion at a dose of greater than 2.times.10.sup.6 iu
at day 1 of week 1 followed by a dose of greater than
2.times.10.sup.6 iu at day 1 of week 2, and every week thereafter
until surgery is scheduled, toxicity or disease progression, a
predetermined maximum number of doses is reached or a fixed number
of doses is reached, complete response, disease progression, or
intolerance of the treatment, wherein the herpes simplex virus
lacks functional ICP34.5 genes; and an immune checkpoint inhibitor
by intravenous infusion every 3 weeks for at least 3 infusions
beginning with or after the second or third dose of the herpes
simplex virus.
[0383] In another aspect of the present invention a method for the
treatment of head and neck cancer is provided, the method
comprising administering: a herpes simplex virus by intravenous
infusion at a dose of greater than 2.times.10.sup.6 iu at day 1 of
week 1 followed by a dose of greater than 2.times.10.sup.6 iu at
day 1 of week 2, and every week thereafter until surgery is
scheduled, toxicity or disease progression, a predetermined maximum
number of doses is reached or a fixed number of doses is reached,
complete response, disease progression, or intolerance of the
treatment, wherein the herpes simplex virus lacks functional
ICP34.5 genes; and an immune checkpoint inhibitor by intravenous
infusion every 3 weeks for at least 3 infusions beginning with or
after the second or third dose of the herpes simplex virus.
[0384] The immune checkpoint inhibitor may be an inhibitor of at
least one of PD-1, PD-L1, CTLA4, TIM-3 or LAG-3. The immune
checkpoint inhibitor may be an anti-PD-1 antibody, anti-PD-L1
antibody, anti-CTLA4 antibody, anti-TIM-3 antibody or anti-LAG-3
antibody. The immune checkpoint inhibitor may be one of
pembrolizumab, nivolumab or ipilimumab.
[0385] In some embodiments the head and neck cancer is selected
from cancer of the larynx, thyroid, pharynx (e.g. throat,
nasopharynx, oropharynx or hypopharynx), salivary glands, oral
cavity (e.g. mouth, lips, tongue, gums, lining inside of the
cheeks, floor of the mouth under the tongue, hard palate), nasal
cavity, nose, paranasal sinuses, or skin cancers of the head and
neck.
[0386] The head and neck cancer may be human papilloma virus
(HPV)-positive and/or Epstein Barr Virus (EBV)-positive, or may be
HPV-negative and/or EBV-negative.
[0387] In one aspect of the present invention a kit of parts is
provided, the kit comprising a herpes simplex virus lacking
functional ICP34.5 genes, and a package insert or label with
directions to treat head and neck cancer by using a combination of
the herpes simplex virus and an immune checkpoint inhibitor.
[0388] The directions may comprise instructions to administer to a
patient with head and neck cancer: a herpes simplex virus
administered by intravenous infusion at a dose of greater than
2.times.10.sup.6 iu at day 1 of week 1 followed by a dose of
greater than 2.times.10.sup.6 iu at day 1 of week 2, and every week
thereafter until surgery is scheduled, toxicity or disease
progression, a predetermined maximum number of doses is reached or
a fixed number of doses is reached, complete response, disease
progression, or intolerance of the treatment; and an immune
checkpoint inhibitor administered intravenously every 3 weeks for
at least 3 infusions beginning with or after the second or third
dose of the herpes simplex virus.
[0389] A method of manufacturing the kit is also provided.
[0390] In another aspect of the present invention a method of
promoting a combination treatment comprising a herpes simplex virus
lacking functional ICP34.5 genes and an immune checkpoint
inhibitor, for the treatment of a patient with head and neck cancer
is provided.
[0391] In some embodiments the promotion is by a package insert,
wherein the package insert provides instructions to receive cancer
treatment with a herpes simplex virus in combination with an immune
checkpoint inhibitor. In some embodiments the promotion is by a
package insert accompanying a formulation comprising the herpes
simplex virus. In some embodiments the promotion is by written
communication to a physician or health care provider. In some
embodiments the promotion is by oral communication to a physician
or health care provider. In some embodiments the promotion is
followed by the treatment of the patient with the herpes simplex
virus.
[0392] In another aspect of the present invention a method of
instructing a patient with head and neck cancer by providing
instructions to receive a combination treatment with a herpes
simplex virus lacking functional ICP34.5 genes and an immune
checkpoint inhibitor to extend survival of the patient is
provided.
[0393] In some embodiments all copies of the ICP34.5 gene in the
genome of the herpes simplex virus are modified such that the
ICP34.5 gene is incapable of expressing a functional ICP34.5 gene
product. As such the herpes simplex virus may be an ICP34.5 null
mutant.
[0394] In some embodiments one or both of the ICP34.5 genes in the
genome of the herpes simplex virus are modified such that the
ICP34.5 gene is incapable of expressing a functional ICP34.5 gene
product.
[0395] The herpes simplex virus may be an oncolytic herpes simplex
virus.
[0396] In some embodiments the herpes simplex virus is a mutant of
HSV-1 strain 17. In preferred embodiments the herpes simplex virus
is HSV1716 (ECACC Accession No. V92012803). HSV1716 is also called
SEPREHVIR.RTM.. In some embodiments the herpes simplex virus is a
mutant of HSV-1 strain 17 mutant 1716.
[0397] In one aspect of the present invention a pharmaceutical
composition comprising a herpes simplex virus and an immune
checkpoint inhibitor is provided. The herpes simplex virus may be a
mutant of HSV-1 strain 17. In preferred embodiments the herpes
simplex virus is HSV1716.
[0398] In one aspect of the present invention a kit is provided,
the kit comprising a predetermined amount of herpes simplex virus
and a predetermined amount of an immune checkpoint inhibitor. The
herpes simplex virus may be a mutant of HSV-1 strain 17. In
preferred embodiments the herpes simplex virus is HSV1716. The kit
may be provided together with instructions for the administration
of the herpes simplex virus, and immune checkpoint inhibitor
sequentially or simultaneously in order to provide a treatment for
head and neck cancer.
[0399] In one aspect of the present invention products are provided
containing therapeutically effective amounts of: [0400] (i)
HSV1716, and [0401] (ii) an immune checkpoint inhibitor for
simultaneous or sequential use in a method of medical treatment,
preferably treatment of head and neck cancer. The products may be
pharmaceutically acceptable formulations and may optionally be
formulated as a combined preparation for coadministration.
[0402] Optionally, in some embodiments the herpes simplex virus
does not express GMCSF. Optionally, in some embodiments the herpes
simplex virus encodes a functional ICP47 gene. Optionally, in some
embodiments the herpes simplex virus is not a herpes simplex virus
that lacks functional ICP34.5 genes and lacks a functional ICP47
gene and comprises a gene encoding human GM-CSF.
[0403] Optionally, in some embodiments the cancer is not a
melanoma. Optionally, in some embodiments the cancer is not a
primary melanoma. Optionally, in some embodiments the cancer is not
a metastatic (secondary) melanoma. Optionally, in some embodiments
the cancer is not stage IIIb to stage IV melanoma.
[0404] A cancer may be any unwanted cell proliferation (or any
disease manifesting itself by unwanted cell proliferation),
neoplasm or tumor or increased risk of or predisposition to the
unwanted cell proliferation, neoplasm or tumor. A cancer may be
benign or malignant and may be primary or secondary (metastatic). A
neoplasm or tumor may be any abnormal growth or proliferation of
cells and may be located in any tissue.
[0405] Head and neck cancers include cancers of the larynx,
thyroid, pharynx (e.g. throat, nasopharynx, oropharynx or
hypopharynx), salivary glands, oral cavity (e.g. mouth, lips,
tongue, gums, lining inside of the cheeks, floor of the mouth under
the tongue, hard palate), nasal cavity, nose, paranasal sinuses, or
skin cancers of the head and neck.
[0406] Head and neck cancer may be oral cancer, e.g. of the tongue,
the lip, the floor of the mouth, the salivary glands, or the gums;
laryngeal cancer, e.g. of the larynx or trachea; pharyngeal cancer,
e.g. cancer of the nasopharynx, the oropharynx, or the hypopharynx;
nasopharyngeal cancer; oropharyngeal cancer, e.g. of the base of
the tongue, the soft palate, and the area around the tonsils;
hypopharyngeal cancer, e.g. of the uppermost portion of the
oesophagus and surrounding the larynx; cancer of the nasal cavity
or cancer of the paranasal sinus.
[0407] In some embodiments head and neck cancer may be squamous
cell carcinoma of the head and neck (SCCHN or HNSCC); primary skin
tumour (squamous cell carcinomas, basal cell carcinomas, melanomas,
and Merkel cell carcinomas), lymphoma, or a malignant tumour of the
salivary glands, soft tissues, and bones. In some embodiments head
and neck cancer may be a metastasis from tumours elsewhere in the
body to the head and neck area.
[0408] A head and neck cancer may be at any stage, e.g. one of
stages I, II, III, IV (i.e. one of IVA, IVB or IVC). In some
preferred embodiments the head and neck cancer is stage III or IV.
In some embodiments the head and neck cancer may be stage III or a
higher stage. In some embodiments the head and neck cancer may be
stage IV. In some embodiments the head and neck cancer may be stage
IVA or a higher stage. In some embodiments the head and neck cancer
may be stage IVB or a higher stage. In some embodiments the head
and neck cancer may be stage IVC or a higher stage.
[0409] In some embodiments a subject having head and neck cancer
may be treated with herpes simplex virus prior to surgery, e.g.
surgical resection of tumor tissue. In some preferred embodiments a
subject having head and neck cancer may be treated with herpes
simplex virus during surgery of the head and neck cancer. For
example, herpes simplex virus may be administered to the site of
surgical resection during or following removal/ablation of tissue.
In some embodiments a subject having head and neck cancer may be
treated with oncolytic virus after surgery of the head and neck
cancer, e.g. during a separate surgical procedure.
[0410] Optionally, the head and neck cancer may not be a cancer of
one or more of the brain, central nervous system, oesophagus, glial
cells, lacrimal gland, larynx, nasopharynx, oral cavity, parotid
gland, salivary gland, skin, thyroid gland, tongue, tonsil, or
trachea.
Types of Head & Neck Cancer
[0411] Cancer can develop in several different parts of the head
and neck, including thyroid, salivary, and skin cancers, but by far
the most common type of cancer is squamous cell carcinoma of the
head and neck (SCCHN or HNSCC). Most HNSCC begins in the layer of
flat cells (the epithelium) which line the structures of the upper
aerodigestive tract, including the mouth (oral cavity), throat
(pharynx), and voice box (larynx). Other malignancies in the head
and neck area include primary skin tumours (squamous cell
carcinomas, basal cell carcinomas, melanomas, and Merkel cell
carcinomas), lymphomas, and malignant tumours of the salivary
glands, soft tissues, and bones. Furthermore, metastases from
tumours from elsewhere in the body may reveal in the head and neck
area.
[0412] Some of the most common head and neck cancers include:
[0413] Oral Cancer: Cancer of the oral cavity is the most common
type of head and neck cancer. Nearly 30,000 new cases of oral
cancer are diagnosed in the US each year. Most oral cancers arise
in the tongue, the lip, the floor of the mouth, and the minor
salivary glands. The rest are found in the gums and other
sites.
[0414] The main risk factors for oral cancer are smoking or chewing
tobacco and excessive alcohol use. People who both smoke and drink
heavily may be as much as 100 times more likely to develop oral
cancer than those who neither smoke nor drink. Additional risk
factors for oral cancer include infection with the human
papillomavirus (HPV), although this risk is not as high as it is
for pharyngeal cancer, and prolonged exposure to sunlight.
[0415] Many oral cancers are found incidentally during a routine
dental examination. Most of these cancers can be cured if
discovered early. The most common symptoms include a sore or lump
on the lip or in the mouth that does not heal; a white and/or red
patch on the gums, tongue, or cheeks (these white or red areas may
also be a precancerous condition called dysplasia); unusual or
persistent bleeding, pain, or numbness in the mouth; and swelling
that causes dentures to fit poorly or become uncomfortable.
[0416] Laryngeal Cancer: Laryngeal cancer is the second most common
type of head and neck cancer. An estimated 12,000 new cases of
laryngeal cancer are diagnosed in the US each year. The vast
majority of laryngeal cancers occur in men.
[0417] The larynx is located at the top of the trachea (windpipe)
and is surrounded by the hypopharynx (the lower part of the throat
where swallowing takes place). The larynx is visible on most men's
throats as the Adam's apple. The larynx contains two bands of
muscle called vocal cords, which vibrate as air passes through to
make speech. The larynx also prevents food from entering the lungs.
Tobacco and alcohol use, and especially the combination of the two,
are the most common risk factors for laryngeal cancer. Additional
risk factors include exposure in the workplace to wood and metal
dusts, asbestos, paint fumes, and other chemical inhalants; a diet
low in vitamins A and E; gastroesophageal reflux disease (GERD),
which chronically exposes the throat to stomach acid; and infection
with the human papillomavirus (HPV). People with aplastic anaemia,
a blood disorder associated with certain hereditary conditions,
also have a higher risk of developing laryngeal cancer.
[0418] The most common symptoms of laryngeal cancer include
hoarseness, a lump in the neck (due to an enlarged lymph node), ear
pain, and difficulty swallowing.
[0419] Pharyngeal (Throat) Cancer: Pharyngeal cancer arises in the
hollow tube inside the neck that starts behind the nose and ends at
the top of the oesophagus. Tumours in this region include cancer of
the nasopharynx (the upper part of the throat behind the nose), the
oropharynx (the middle part of the pharynx), and the hypopharynx
(the bottom part of the pharynx). Each year in the US, an estimated
11,800 people develop pharyngeal cancers.
[0420] Nasopharyngeal Cancer: The nasopharynx, located behind the
nose, includes two openings that lead to the ears. Nasopharyngeal
cancer is much more common in Asia, especially southeast China, the
Mediterranean area, and Africa than in the US, and is less commonly
associated with tobacco and alcohol use than other head and neck
cancers. Risk factors for this type of cancer include a diet high
in salt-cured fish and infection with Epstein-Barr virus, a member
of the herpesvirus family and one of the most common human viruses.
The most common sign of nasopharyngeal cancer is a lump in the
neck, caused by the spread of cancer to the lymph nodes. Other
symptoms may include nasal congestion, pain or ringing in the ears,
a persistent sore throat, or frequent nosebleeds.
[0421] Oropharyngeal Cancer: The oropharynx is located behind the
mouth and includes the base of the tongue, the soft palate, and the
area around the tonsils. Smoking and chewing tobacco and heavy
alcohol use are the most common risk factors for oropharyngeal
cancer, but there is evidence that a diet low in fruits and
vegetables is linked to this form of head and neck cancer. Prior
infection with HPV is also a particularly strong risk factor for
this cancer site. Symptoms of oropharyngeal cancer may include a
lump in the neck or throat, persistent sore throat, hoarseness,
difficulty swallowing, and ear and/or jaw pain.
[0422] Hypopharyngeal Cancer: The hypopharynx is the uppermost
portion of the oesophagus and surrounds the larynx (voice box). As
with most other head and neck cancers, tobacco use and heavy
alcohol consumption are the most common risk factors. Other risk
factors for hypopharyngeal cancer may include a diet low in
vitamins A and E; exposure in the work place to asbestos, wood
dust, paint fumes, and other inhalants. Symptoms of hypopharyngeal
cancer may include a lump in the neck, hoarseness, difficulty
swallowing, and ear pain.
[0423] Nasal Cavity & Paranasal Sinus Cancers: Each year,
approx 2,000 people in the US are diagnosed with cancer in the
mucus-producing tissues that line the nasal cavity (the space
behind the nose through which air passes to the throat) and the
paranasal sinuses (hollow areas in the facial bones near the nose).
More than half of nasal cavity and paranasal sinus cancers occur in
the maxillary sinuses (hollow spaces on either side of the nose and
below the eyes); fewer cancers develop in the nasal cavity and in
the ethmoid sinuses (sieve-like spaces made of thin bone and mucous
tissues behind the bridge of the nose). These cancers arise more
frequently in people who are exposed to wood and metal dusts,
asbestos, paint fumes, and air pollution. Symptoms may include
persistent nasal congestion, chronic sinus infections that do not
respond to antibiotic treatment, frequent headaches or sinus pain,
swelling of the eyes, and reduced sense of smell.
Staging
[0424] The tumor, node, metastasis (TNM) staging system allows
clinicians to categorize tumors of the head and neck region in a
specific manner to assist with the assessment of disease status,
prognosis, and management. [4.sup.th edition of the Quick Reference
Guide to TNM Staging of Head and Neck Cancer and Neck Dissection
Classification, the American Academy of Otolaryngology--Head and
Neck Surgery Foundation and American Head and Neck Society
(2014)].
[0425] All available clinical information may be used in staging:
physical exam, radiographic, intraoperative, and pathologic
findings. Other than histopathologic analysis, biomarkers and
molecular studies are not yet included in the staging of head and
neck cancers.
[0426] Three categories comprise the system: [0427] T: the
characteristics of the tumor at the primary site (this may be based
on size, location, or both); [0428] N: the degree of regional lymph
node involvement; and [0429] M: the absence or presence of distant
metastases.
[0430] The specific TNM status of each patient is then tabulated to
give a numerical status of Stage I, II, III, or IV. Specific
subdivisions may exist for each stage and may be denoted with an a,
b, or c status.
Stage 0 Head and Neck Cancer
[0431] A stage 0 head and neck cancer tumor means the cancer is
only growing in the part of the head and neck where it started. No
cancer cells are present in deeper layers of tissue, nearby
structures, lymph nodes, or distant sites (Example: Tis, N0, M0
carcinoma in situ).
Stage I Head and Neck Cancer
[0432] A stage I head and neck cancer tumor means the primary tumor
is 2 cm across or smaller, and no cancer cells are present in
nearby structures, lymph nodes, or distant sites (Example: T1, N0,
M0).
Stage II Head and Neck Cancer
[0433] A stage II head and neck tumor measures 2-4 cm across, and
no cancer cells are present in nearby structures, lymph nodes, or
distant sites (Example: T2, N0, M0).
Stage III Head and Neck Cancer
[0434] A stage III head and neck tumor means one of the following:
[0435] The head and neck tumor is >4 cm across, and no cancer
cells are present in nearby structures, lymph nodes, or distant
sites (Example: T3, N0, M0). [0436] The head and neck tumor is any
size but has not grown into nearby structures or distant sites.
However, cancer cells are present in one lymph node, which is
located on the same side of the head or neck as the primary tumor
and is <3 cm across (Example: T1-3, N1, M0).
Stage IV Head and Neck Cancer
[0437] Stage IVA: One of the following applies-- [0438] T4a, N0 or
N1, M0: the head and neck cancer tumor is any size and is growing
into nearby structures. Cancer cells may not be present in the
lymph nodes, or they may have spread to one lymph node, which is
located on the same side of the head or neck as the primary tumor
and is <3 cm across. Cancer has not spread to distant sites; or
[0439] T1-4a, N2, M0: the tumor is any size and may or may not have
invaded nearby structures, it has not spread to distant sites, and
one of the following is true: [0440] cancer present in one lymph
node, located on the same side of the head or neck as the primary
tumor and measuring 3-6 cm across (N2a); [0441] cancer present in
one lymph node on the opposite side of the head or neck and
measuring <6 cm across (N2b); [0442] cancer present in 2 or more
lymph nodes, all <6 cm across and located on either side of the
head or neck (N2c);
[0443] Stage IVB: One of the following applies-- [0444] T4b, any N,
M0: the tumor has invaded deeper areas and/or tissues. It may or
may not have spread to lymph nodes and has not spread to distant
sites. or [0445] Any T, N3, M0: the tumor is any size and may or
may not have grown into other structures. It has spread to one or
[0446] more lymph nodes >6 cm across, but has not spread to
distant sites.
[0447] Stage IVC: [0448] Any T, Any N, M1: The head and neck cancer
tumor is any size and may or may not have spread to lymph nodes.
[0449] Cancer cells have spread to distant sites.
[0450] T4a disease indicates moderately advanced disease and is
specific by subsite, but is still considered resectable. T4b
disease is very advanced disease with findings--such as carotid
artery encasement, prevertebral involvement, and skullbase
involvement--that previously determined the disease to be
unresectable. In general, early-stage disease is denoted as Stage I
or II disease, and advanced stage disease as Stage III or IV
disease. Of importance is that any positive metastatic disease to
the neck will classify the disease as advanced, except in select
nasopharynx and thyroid cancers. T4a disease is staged as IVa. T4b
disease is staged as IVb, and any distant metastasis is staged as
IVc.
Risk Factors
[0451] Risk factors for head and neck cancers include: tobacco use,
heavy alcohol consumption, prolonged sun exposure, and certain
viruses, including human papillomavirus (HPV) and Epstein-Barr
virus (EBV). In particular, HPV infection is a risk factor for
oropharyngeal cancer (cancer of the middle of the throat, including
the tonsils and base of tongue). The overall incidence of
HPV-positive head and neck cancers is rapidly increasing in the US,
while the incidence of HPV-negative (primarily tobacco- and
alcohol-related) cancer is decreasing. While a strong causal
relationship has been established between HPV type 16 and the
development of oropharyngeal cancer, other HPV types have also been
associated with oropharyngeal cancer. Human papillomavirus (HPV) is
the most common sexually transmitted disease in the US, infecting
79 million Americans. HPV is known to play a major role in the
development of head and neck cancers, which include cancers of the
oral cavity, oropharynx, nose/nasal passages and larynx. Head and
neck cancers associated with HPV account for nearly 3% of all
cancers in the US and are twice as prevalent in men as in women.
Incidence rates of HPV-caused head and neck cancers have been on
the rise, especially HPV-associated oropharyngeal cancer in men,
and are expected to continue growing. By 2025, researchers believe
that HPV will be the causative factor of 90% of all head and neck
cancers. HPV-related head and neck cancer has a unique risk factor
profile, and a more favourable prognosis than tobacco or alcohol
induced HNSCC. Unlike HPV-negative SCCHN, which is driven by
stepwise mutations in the squamous epithelium, HPV-positive SCCHN
is caused by two viral oncogenes that inactivate tumour suppressor
genes and lead to malignant transformation of the squamous
epithelium.
Treatment
[0452] Many cancers of the head and neck can be cured, especially
if they are found early. Treatment varies according to the type,
location, and extent of the cancer. In addition to treatment with
hereps simplex virus and immune checkpoint inhibitors, accompanying
treatment(s) may include a combination of surgery, radiation
therapy, and chemotherapy.
Surgery
[0453] Surgery is the primary treatment for most cancers of the
head and neck. Improvements in surgical techniques allow removal of
many more tumours while preserving nearby structures involved in
sensory and physical functioning. Some patients may require
surgical examination of the lymph nodes in the neck to determine if
any cancer cells have spread beyond their original site. New
techniques allow surgeons to remove these lymph nodes while sparing
nerves that are important for shoulder function. Complex surgery to
remove tumours at the base of the skull, once considered nearly
impossible, can now be safely performed. Reconstruction of bones
and other structures is often possible immediately following
surgery.
[0454] Minimally invasive surgical techniques are used when
possible to remove tumours that are located near structures
involved in sensory and physical functioning. In many cases,
patients can recover more quickly when treated with minimally
invasive surgery compared with traditional, open surgery.
[0455] Endoscopic Laser Surgery may be used to remove tumours in
the larynx or pharynx while preserving the structures involved in
speech and swallowing.
[0456] Robotic Surgery can be used for tumours of the tongue and
tonsils can be removed with the aid of small robotic arms that are
placed in the mouth, avoiding the need to make a large incision or
to split the jawbone.
Radiation Therapy
[0457] Radiation therapy alone or in combination with chemotherapy
is standard curative treatment for many patients with head and neck
cancers. Which approach is used depends on the extent of the
tumour; radiation and chemotherapy are used in combination when
treating more advanced disease. In select situations, such as oral
cavity tumours, the patient undergoes surgery followed by radiation
therapy and/or chemotherapy. Radiation therapy, or a combination of
radiation and chemotherapy, may be used to treat patients who would
develop significant side effects from surgery, those with
inoperable cancers, or those who have a poor prognosis after
surgery.
[0458] Patients may be treated with one or both of the following
types of radiation therapy: External-beam radiation therapy called
intensity-modulated radiation therapy (IMRT), can be used which
uses 3-D images from CT scans to deliver radiation to tumours with
greater precision than conventional radiation therapy.
[0459] Brachytherapy uses tiny, radioactive seeds which are
implanted into the tumour site, where they deliver the highest dose
of radiation possible with minimal effect on nearby healthy
tissue.
[0460] Proton Therapy: For some cases of head and neck cancer,
proton therapy can be used to deliver high doses of radiation to
tumours that may be resistant to conventional forms while
minimising exposure to the surrounding healthy tissues thereby
reducing the risk of treatment-related side effects.
Chemotherapy
[0461] Increasingly, chemotherapy, in combination with radiation
therapy, is used to treat head and neck cancers that are difficult
to reach surgically or that cannot be cured by surgery alone. This
approach is also used to treat patients for whom surgery would
cause significant functional or cosmetic disability, such as loss
of the larynx, with its associated loss of natural voice and the
need for a permanent stoma in the front of the neck. Chemotherapy
has been shown to enhance the effectiveness of radiation therapy,
improving cure rates compared to radiation therapy alone for
advanced cancers such as those originating in the nasopharynx.
[0462] Chemotherapy is also used for patients with incurable
disease, in an attempt to improve survival and decrease
cancer-related symptoms. The most commonly used chemotherapy drugs
include cisplatin, fluorouracil, methotrexate, carboplatin,
paclitaxel, docetaxel, and, more recently, cetuximab.
[0463] There are many investigational agents currently in clinical
trials for head and neck cancer. Early stage tumours are often
cured by radiation or surgery alone. However, up to 60% of patients
present as locally advanced disease (stages III and IVA/B)
requiring multi-modality therapy.
[0464] Based on a pivotal phase III trial published in 2003,
concurrent chemo-radiation using cisplatin has become the standard
of care for patients with unresectable locally advanced disease.
Cisplatin causes crosslinking of DNA followed by inhibiting
mitosis, and also has anti-tumour immunomodulatory effects. These
effects are induced by four distinguishing mechanisms, for example,
upregulation of MHC class I, recruitment and proliferation of
effector T cells and macrophages, enhancement of lytic activity in
cytotoxic effector cells, and downregulation of MDSCs and Tregs.
The three year overall survival was 23% in the radiation alone arm
compared to 37% in the chemo-radiation arm (p value 0.014)
[Adelstein D J, Li Y, Adams G L, Wagner H, Jr, Kish J A, Ensley J
F, et al. An intergroup phase III comparison of standard radiation
therapy and two schedules of concurrent chemoradiotherapy in
patients with unresectable squamous cell head and neck cancer. J
Clin Oncol. 2003 Jan. 1; 21(1):92-8]. However, despite current
therapy, many of these patients are faced with high rates of
recurrence and progression necessitating the development of novel
agents.
[0465] In patients that develop unresectable local recurrences
and/or metastatic disease, the prognosis is poor. The 1-year
survival rate is approximately 10-15% [Patel P R, Salama J K.
Reirradiation for recurrent head and neck cancer. Expert Rev
Anticancer Ther. 2012 September; 12(9):1177-89]. Overall survivals
(OS) were in the range of 5-6 months in these patients when treated
with single agent chemotherapy [Jacobs C, Lyman G, Velez-Garcia E,
Sridhar K S, Knight W, Hochster H, et al. A phase III randomized
study comparing cisplatin and fluorouracil as single agents and in
combination for advanced squamous cell carcinoma of the head and
neck. J Clin Oncol. 1992 February; 10(2):257-63 and Forastiere A A,
Metch B, Schuller D E, Ensley J F, Hutchins L F, Triozzi P, et al.
Randomized comparison of cisplatin plus fluorouracil and
carboplatin plus fluorouracil versus methotrexate in advanced
squamous-cell carcinoma of the head and neck: A southwest oncology
group study. J Clin Oncol. 1992 August; 10(8):1245-51] and up until
recently, combination regimens did not significantly increase the
OS over single agents.
[0466] In 2005, it was showed for the first time in a phase III
study that adding the EGFR antibody, cetuximab, to cisplatin
increased OS over cisplatin alone in the recurrent and metastatic
disease setting [Burtness B, Goldwasser M A, Flood W, Mattar B,
Forastiere A A, Eastern Cooperative Oncology Group. Phase III
randomized trial of cisplatin plus placebo compared with cisplatin
plus cetuximab in metastatic/recurrent head and neck cancer: An
eastern cooperative oncology group study. J Clin Oncol. 2005 Dec.
1; 23(34):8646-54]. The response rate in the combination arm in the
latter study was 26%. In the XTREME study that combined cisplatin
plus 5 FU chemotherapy with cetuximab showed a response rate of 36%
that equated to an OS in these patients to 10.1 months [Vermorken J
B, Mesia R, Rivera F, Remenar E, Kawecki A, Rottey S, et al.
Platinum-based chemotherapy plus cetuximab in head and neck cancer.
N Engl J Med. 2008 Sep. 11; 359(11):1116-27]. In another study, the
alternate anti-EGFR monoclonal antibody, panitumumab in addition to
cisplatin/5 FU also showed an improvement in OS to 11.1 over
control, but this benefit was limited to p16-negative tumours and
was associated with an increase in serious adverse events
[Vermorken J B, Stohlmacher-Williams J, Davidenko I, Licitra L,
Winquist E, Villanueva C, et al. Cisplatin and fluorouracil with or
without panitumumab in patients with recurrent or metastatic
squamous-cell carcinoma of the head and neck (SPECTRUM): An
open-label phase 3 randomised trial. Lancet Oncol. 2013 July;
14(8):697-710].
Subjects
[0467] The subject to be treated may be any animal or human. The
subject is preferably mammalian, more preferably human. The subject
may be a non-human mammal, but is more preferably human. The
subject may be male or female. The subject may be a patient. A
subject may have been diagnosed with a cancer, or be suspected of
having a cancer.
[0468] The subject may be a child, i.e. a human subject of age less
than 18 years, or of age less than 16 years, or of age less than 14
years, or of age less than 12 years. The age may be determined at
the point of first dose with oncolytic herpes simplex virus.
[0469] Subjects may optionally be indicated for surgical removal of
tumor tissue (referred to herein as `tumor resection`). For
example, they may have a cancer considered, by a medical
practitioner, operable to remove some or all of the tumor
tissue.
[0470] Subjects may be selected for treatment as being subjects who
have not mounted a clinical response to previous treatment with an
immune checkpoint inhibitor as monotherapy.
[0471] A subject may be immunocompetent or immunocompromised.
Other Chemotherapeutic Agents
[0472] In addition to treating a cancer by using an oncolytic
herpes simplex virus with or without an immune checkpoint
inhibitor, subjects being treated may also receive treatment with
other chemotherapeutic agents. For example, other chemotherapeutic
agents may be selected from: [0473] (i) alkylating agents such as
cisplatin, carboplatin, mechlorethamine, cyclophosphamide,
chlorambucil, ifosfamide; [0474] (ii) purine or pyrimidine
anti-metabolites such as azathiopurine or mercaptopurine; [0475]
(iii) alkaloids and terpenoids, such as vinca alkaloids (e.g.
vincristine, vinblastine, vinorelbine, vindesine), podophyllotoxin,
etoposide, teniposide, taxanes such as paclitaxel (Taxol.TM.),
docetaxel; [0476] (iv) topoisomerase inhibitors such as the type I
topoisomerase inhibitors camptothecins irinotecan and topotecan, or
the type II topoisomerase inhibitors amsacrine, etoposide,
etoposide phosphate, teniposide; [0477] (v) antitumor antibiotics
(e.g. anthracyline antibiotics) such as dactinomycin, doxorubicin
(Adriamycin.TM.), epirubicin, bleomycin, rapamycin; [0478] (vi)
antibody based agents, such as anti-VEGF, anti-TNF.alpha.,
anti-IL-2, antiGpIIb/IIIa, anti-CD-52, anti-CD20, anti-RSV,
anti-HER2/neu(erbB2), anti-TNF receptor, anti-EGFR antibodies,
monoclonal antibodies or antibody fragments, examples include:
cetuximab, panitumumab, infliximab, basiliximab, bevacizumab
(Avastin.RTM.), abciximab, daclizumab, gemtuzumab, alemtuzumab,
rituximab (Mabthera.RTM.), palivizumab, trastuzumab, etanercept,
adalimumab, nimotuzumab, [0479] (vii) EGFR inihibitors such as
erlotinib, cetuximab and gefitinib [0480] (viii) anti-angiogenic
agents such as bevacizumab (Avastin.RTM.).
Routes of Administration
[0481] Immune checkpoint inhibitors, chemotherapeutic agents,
medicaments and pharmaceutical compositions according to aspects of
the present invention may be formulated for administration by a
number of routes, including but not limited to, parenteral,
intravenous, intra-arterial, intramuscular, intratumoral and oral.
Immune checkpoint inhibitors, chemotherapeutic agents, medicaments
and pharmaceutical compositions may be formulated in fluid or solid
form. Fluid formulations may be formulated for administration by
injection to a selected region of the human or animal body.
[0482] In the present invention herpes simplex virus is
administered to the blood, e.g. by intravenous administration
(intravenous infusion) or intra-arterial administration, and is
formulated for such administration.
[0483] In preferred embodiments the immune checkpoint inhibitor is
administered to the blood, e.g. by intravenous administration
(intravenous infusion), and is formulated for such
administration.
[0484] Herpes simplex virus may be formulated as a suspension of
virus in lactated Ringer's or in Hartmann;s solution. One litre of
lactated Ringer's solution typically contains about 130 mEq of
sodium ion (130 mmol/L), 109 mEq of chloride ion (109 mmol/L), 28
mEq of lactate (28 mmol/L), 4 mEq of potassium ion (4 mmol/L), and
3 mEq of calcium ion (1.5 mmol/L). One litre of Hartmann's solution
typically contains about 131 mEq of sodium ion (131 mmol/L), 111
mEq of chloride ion (111 mmol/L), 29 mEq of lactate (29 mmol/L), 5
mEq of potassium ion (5 mmol/L), and 4 mEq of calcium ion (2
mmol/L).
[0485] Virus may be formulated for delivery in the clinic by mixing
a small aliquot of virus with a specified volume of the chosen
fluid carrier, e.g. lactated Ringer's or Hartmann's solution. Virus
is supplied in fluid suspension at the specified dosage
concentration, e.g. 1.times.10.sup.7 iu/ml and a small aliquot in
the range 0.5 to 5 ml, e.g. one of about 0.5 ml, about 1 ml, about
2 ml, about 3 ml, about 4 ml or about ml, preferably about 1 ml of
virus, is mixed with the fluid carrier. The volume of fluid carrier
to which the aliquot of virus is added may be one of about 100 ml,
about 150 ml, about 200 ml, about 250 ml, about 300 ml, about 350
ml, about 400 ml, about 450 ml, about 500 ml. In some preferred
embodiments the volume of fluid carrier is about 250 ml. The fluid
carrier may be provided in a bag suitable for use in intravenous or
intra-arterial infusion. The viral suspension, fluid carrier and
bag are all preferably sterile and the virus formulation is
prepared in sterile conditions.
[0486] Infusion of the formulated viral composition to the blood
may take between about 30 minutes and about 3 hours, for example
about 1 hour, about 2 hours or about 3 hours.
[0487] Intravenous administration may comprise infusion into the
venous system in close proximity to the location or locations of
the cancer, e.g. head and neck cancer.
[0488] Infusion to the blood is preferably at a peripheral site,
e.g. to a vein or artery near the surface of the skin and not
within deep tissue. Examples of suitable peripheral locations are
veins in the arm or leg. In some related embodiments,
administration may be via a central venous line. Administration is
preferably non-invasive, e.g. does not require a surgical, invasive
or interventional radiological procedure in order to locate a
specific vein or artery within deep tissue or proximal to internal
organs. For example, administration is optionally not to the
hepatic artery. The subject may have a peripheral venous device,
catheter or cannula fitted in order to facilitate the
administration. As such, administration can be performed in an
out-patient setting in which the patient is connected to a
drip.
[0489] Administration of oncolytic herpes simplex virus may be
locoregional administration, e.g. to a localised region of the body
in which the tumor is present. Locoregional administration may be
achieved by use of chemoembolization in which administration of an
oncolytic herpes simplex virus may be combined with other
embolization (e.g. chemical embolization) of the tumor.
[0490] An example of a less preferred, and non-peripheral, route of
administration, developed in the context of treatment of primary
liver cancer, is trans-arterial chemoembolization (TACE).
[0491] TACE is normally performed by an interventional radiologist
and involves accessing the hepatic artery with a catheter, which is
possible by puncturing the common femoral artery in the right groin
and passing a catheter through the abdominal aorta, through the
celiac trunk and common hepatic artery, into the proper hepatic
artery.
[0492] An arteriogram is performed to identify the branches of the
hepatic artery supplying the tumor(s). Smaller catheters may then
be threaded into these branches (so-called superselective
positioning). This allows precision delivery of the active agents
to the tumor tissue.
[0493] Once the catheter is in position, doses of the active agent
(e.g. oncolytic herpes simplex virus, and/or chemotherapeutic agent
and/or embolisation agent and/or contrast agent) are injected
through the catheter. The total dose may be given to a single
vessel, or if there are several tumor foci may be divided among
several vessels supplying the tumors.
[0494] Because most liver tumors are supplied by the hepatic
artery, arterial embolization interrupts the blood supply to the
tumor and delays tumor growth. The focused nature of the
administration of active agents enables delivery of a high
therapeutic dose to the tissue requiring treatment whilst reducing
systemic exposure and therefore toxicity. Embolization of the
vessel assists this process in that the active agent(s) is not
washed out from the tumor bed and the supply of nutrients to the
tumor is decreased thereby promoting tumor necrosis.
[0495] TACE is widely used as a palliative treatment for surgically
unresectable primary or metastatic HCC tumors.
[0496] In some optional embodiments, administration is not
intraperitoneal.
Kits
[0497] In some aspects of the present invention a kit of parts is
provided. In some embodiments the kit may have at least one
container having a predetermined quantity of herpes simplex virus,
e.g. predetermined viral dose or number/quantity/concentration of
viral particles. The herpes simplex virus may be formulated so as
to be suitable for injection or infusion to a tumor or to the
blood. In some embodiments the kit may further comprise at least
one container having a predetermined quantity of immune checkpoint
inhibitor. The immune checkpoint inhibitor may also be formulated
so as to be suitable for injection or infusion to the tumor or to
the blood, or alternatively may be formulated for oral
administration. In some embodiments a container having a mixture of
a predetermined quantity of herpes simplex virus and predetermined
quantity of immune checkpoint inhibitor is provided, which may
optionally be formulated so as to be suitable for injection or
infusion to the tumor or to the blood.
[0498] In some embodiments the kit may also contain apparatus
suitable to administer one or more doses of the herpes simplex
virus and/or immune checkpoint inhibitor. Such apparatus may
include one or more of a catheter and/or needle and/or syringe,
such apparatus preferably being provided in sterile form.
[0499] The kit may further comprise instructions for the
administration of a therapeutically effective dose of the herpes
simplex virus and/or immune checkpoint inhibitor.
[0500] The invention includes the combination of the aspects and
preferred features described except where such a combination is
clearly impermissible or expressly avoided.
[0501] The section headings used herein are for organizational
purposes only and are not to be construed as limiting the subject
matter described.
[0502] Aspects and embodiments of the present invention will now be
illustrated, by way of example, with reference to the accompanying
figures. Further aspects and embodiments will be apparent to those
skilled in the art. All documents mentioned in this text are
incorporated herein by reference.
[0503] Throughout this specification, including the claims which
follow, unless the context requires otherwise, the word "comprise,"
and variations such as "comprises" and "comprising," will be
understood to imply the inclusion of a stated integer or step or
group of integers or steps but not the exclusion of any other
integer or step or group of integers or steps.
[0504] It must be noted that, as used in the specification and the
appended claims, the singular forms "a," "an," and "the" include
plural referents unless the context clearly dictates otherwise.
Ranges may be expressed herein as from "about" one particular
value, and/or to "about" another particular value. When such a
range is expressed, another embodiment includes from the one
particular value and/or to the other particular value. Similarly,
when values are expressed as approximations, by the use of the
antecedent "about," it will be understood that the particular value
forms another embodiment.
BRIEF DESCRIPTION OF THE FIGURES
[0505] Embodiments and experiments illustrating the principles of
the invention will now be discussed with reference to the
accompanying figures in which:
[0506] FIG. 1. Table showing changes in cytokine production in
pleural fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
Changes are relative to patient levels of cytokine production prior
to treatment with SEPREHVIR.RTM.. *No samples were available for
testing from patient 06. +.fwdarw.+++++=small to large increase
over baseline level; -=no change; .dwnarw.=decrease; tba=sample yet
to be analysed.
[0507] FIG. 2. Table showing changes in IFN-.gamma. production in
pleural fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0508] FIG. 3. Table showing changes in IFN-.alpha. production in
pleural fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0509] FIG. 4. Table showing changes in IL-1.alpha. production in
pleural fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0510] FIG. 5. Table showing changes in IL-2 production in pleural
fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0511] FIG. 6. Table showing changes in IL-4 production in pleural
fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0512] FIG. 7. Table showing changes in IL-6 production in pleural
fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0513] FIG. 8. Table showing changes in IL-10 production in pleural
fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0514] FIG. 9. Table showing changes in IL-12 production in pleural
fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0515] FIG. 10. Table showing changes in IL-21 production in
pleural fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0516] FIG. 11. Table showing changes in TNF-.alpha. production in
pleural fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0517] FIG. 12. Table showing changes in IP-10 production in
pleural fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0518] FIG. 13. Table showing changes in MIG production in pleural
fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0519] FIG. 14. Table showing changes in VEGF production in pleural
fluid samples in human patients having malignant pleural
mesothelioma in response to treatment with 1, 2 or 4 doses (each of
1.times.10.sup.7 iu) of SEPREHVIR.RTM. given at weekly intervals.
No samples were available for testing from patient 06.
+.fwdarw.+++++=small to large increase over baseline level; -=no
change; .dwnarw.=decrease; tba=sample yet to be analysed.
[0520] FIG. 15. Diagram illustrating Western Blotting procedure
used to probe cell extracts for anti-tumor IgG response induced by
intrapleural administration of SEPREHVIR.RTM..
[0521] FIG. 16. Western Blot showing results for sera taken from
patients 01, 02, 03, 04 and 05 against MSTO-211H cells. Arrows
indicates novel IgG anti-tumor response.
[0522] FIG. 17. Western Blot showing results for sera taken from
patients 06 and 07 against MSTO-211H cells. Arrows indicates novel
IgG anti-tumor response.
[0523] FIG. 18. Western Blot showing results for sera taken from
patients 08 and 09 against MSTO-211H cells. Arrows indicates novel
IgG anti-tumor response.
[0524] FIG. 19. Table showing Th1 response and patient survival for
one year or less or for greater than one year. Median survival is
12 months from diagnosis. *Only 7/9 patients were evaluated. No
samples available from patient 06 and patient 09 is yet to reach 12
months from treatment. .sup.#10 patients, **patients 08 and 09
still alive, ***oldest treated patient was 84 yrs. Historical
median survival for all malignant pleural mesothelioma patients is
9.5 mths from diagnosis (Beckett et al (2015) Lung Cancer 88,
344).
[0525] FIG. 20. Table showing cytokines and chemokines detected in
pleural fluid samples at high levels (ng/ml) and low levels (pg/ml)
and cytokines and chemokines not detected.
[0526] FIG. 21. Seprehvir replicates and persists but patient 02
fails to mount a Th1 response. Charts show: HSV DNA detectable in
pleural fluid samples taken from patient 02 at intervals post
administration of Seprehvir; IFN.gamma. response, MIG (CXCL9)
response and IL-12 response.
[0527] FIG. 22. Seprehvir is undetectable but patient 03 mounts a
robust IFN.gamma. response. Charts show: HSV DNA detectable in
pleural fluid samples taken from patient 03 at intervals post
administration of Seprehvir; IFN.gamma. response, MIG (CXCL9)
response and IL-12 response.
[0528] FIG. 23. Table showing expression of pleural fluid cytokines
and chemokines in nine human patients in response to treatment with
Seprehvir.
[0529] FIG. 24. Table showing cytokines and chemokines showing low
or no response to treatment with Seprehvir in pleural fluid samples
from nine human patients.
[0530] FIG. 25. Table showing summary of patient Th1 cytokine and
chomokine responses in nine human patients.
[0531] FIG. 26. Charts showing immune cell recruitment
post-Seprehvir is associated with Th1 cytokine response in two
patients receiving two doses of Seprehvir (on days 1 and 8) and
three patients receiving four doses of Seprehvir (on days 1, 8, 15
and 22). Day 0 data point represents cell count prior to treatment
with Seprehvir. Th1 responses were observed in patients 04, 07, 08
and 09. Pt=patient.
[0532] FIG. 27. Charts showing Granzyme B is associated with Th1
cytokine response post Seprehvir treatment in three patients
receiving one dose of Seprehvir, two patients receiving two doses
of Seprehvir and three patients receiving four doses of Seprehvir.
Th1 responses were observed in patients 01, 03, 04, 07, 08 and 09.
Pt=patient. Granzyme B and perforin have been shown to induce
CTL-mediated target cell DNA fragmentation and apoptosis. (Lord et
al., Granzyme B: a natural born killer. Immun Rev. 2003 Jun.
193:31-8).
[0533] FIG. 28. C57BL/6 mice were injected with 5.times.10.sup.6
M3-9-M cells subcutaneously. Tumors were treated intra-tumorally
(i.tu.) with Seprehvir when sizes reached 200.about.400 mm.sup.3.
Intra-peritoneal (i.p.) injection of anti-PD-1 antibody were given
twice a week after last dose of virus treatment. Tumor growth was
monitored twice a week. Mice were sacrificed when tumors reached
2,500 mm.sup.3 in volume or grew over 2 cm in length. pfu=Plaque
Forming Unit.
[0534] FIG. 29. Combination of Seprehvir with anti-PD-1 antibody
significantly prolongs survival with several complete responses in
male to female M3-9-M rhabdomyosarcoma tumor model. Female C57BL/6
mice were injected with 5.times.10.sup.6 M3-9-M cells
subcutaneously. The effects of Seprehvir plus anti-PD-1 blockade on
antitumor efficacy were evaluated by measuring tumor volumes over
time. Survival data were evaluated for statistical significance
with Log-rank (Mantel-Cox) test. Upper left chart: PBS/Ctrl Ab
(n=10) and PBS/Anti-PD-1 antibody (n=11); Upper middle chart:
PBS/Ctrl Ab (n=10) and Seprehvir/Ctrl (n=12); Upper right chart:
PBS/Ctrl Ab (n=10) and Seprehvir/Anti-PD-1 antibody (n=11). Lower
chart: PBS/Ctrl Ab (n=10)--dotted line, left-hand side;
PBS/Anti-PD-1 antibody (n=11)--dashed and dotted line;
Seprehvir/Ctrl (n=12)--dashed line; Seprehvir/Anti-PD-1 antibody
(n=11)--solid line, right-hand side.
[0535] FIG. 30. Combination of Seprehvir with anti-PD-1 antibody
significantly prolongs survival in less immunogenic male to male
M3-9-M tumor model. Male C57BL/6 mice were injected with
5.times.10.sup.6 M3-9-M cells subcutaneously. The effects of
Seprehvir plus anti-PD-1 blockade on antitumor efficacy were
evaluated by measuring tumor volumes over time. Survival data were
evaluated for statistical significance with Log-rank (Mantel-Cox)
test. Upper left chart: PBS/Ctrl Ab (n=7) and PBS/Anti-PD-1
antibody (n=6); Upper middle chart: PBS/Ctrl Ab (n=7) and
Seprehvir/Ctrl (n=6); Upper right chart: PBS/Ctrl Ab (n=7) and
Seprehvir/Anti-PD-1 antibody (n=7). Lower chart: PBS/Ctrl Ab
(n=7)--dotted line, left-hand side; PBS/Anti-PD-1 antibody
(n=6)--dashed and dotted line; Seprehvir/Ctrl (n=6)--dashed line;
Seprehvir/Anti-PD-1 antibody (n=7)--solid line, right-hand
side.
[0536] FIG. 31. Checkpoint inhibition does not significantly alter
intra-tumoral viral kinetics. Female M3-9-M tumor-bearing mice were
treated with three doses of 10.sup.8 pfu of Seprehvir
intra-tumorally (i.tu.) followed by intra-peritoneal (i.p.)
injection of anti-PD-1 or control antibody. Tumors were harvested
3, 24, 72 and 168 hours after intra-peritoneal antibody injection
for plaque assay. Data are expressed as total plaque-forming units
(pfu) per tumor.
[0537] FIGS. 32A and 32B. Combination therapy induces more CD4+ and
CD8+ T cells (A) but the increase in CD4+ T cells in both female
(left) and male (right) is not due to higher numbers of CD25+CD4+
Treg cells (B). Female and male M3-9-M tumor-bearing mice received
three doses of intra-tumoral (i.tu.) Seprehvir injection followed
by intra-peritoneal (i.p.) injection of anti-PD-1 or control
antibody. Immune cell infiltrates in tumors were evaluated by flow
cytometry analyses 72 hours post intra-peritoneal antibody
injection. In each chart, columns from left to right are
PBS/Control antibody, PBS/Anti-PD-1 antibody, Seprehvir/Control
antibody and Seprehvir/Anti-PD-1 antibody.
[0538] FIG. 33. Combination therapy induces higher inflammatory
gene expression and lower immune suppressive gene expression.
Female M3-9-M tumor-bearing mice received three doses of
intra-tumoral (i.tu.) Seprehvir injection followed by
intra-peritoneal (i.p.) injection of anti-PD-1 or control antibody.
Tumors were harvested 72 hours post intra-peritoneal antibody
injection. T-bet (Th-1-related gene), Foxp3 (Treg-related gene),
IFN.gamma., IL-10, iNOS (M1 macrophage-related gene) and MRC-1 (M2
macrophage-related gene) were quantified by real-time. Date are
represented as relative RNA expression to gapdh. In each chart,
columns from left to right are PBS/Control antibody, PBS/Anti-PD-1
antibody, Seprehvir/Control antibody and Seprehvir/Anti-PD-1
antibody.
[0539] FIG. 34. Tumour microenvironment is remodeled to Th1 away
from Th2 by Seprehvir and combination with anti-PD-1. IFN.gamma.
and iNOS are used as Th1 markers and their proportion, relative to
the Th2 markers IL-10 and MRC-1 are significantly increased by
Seprehvir+anti-PD-1.
[0540] FIG. 35. Charts showing results of re-challenge experiment
in which mice cured of xenograft tumor are vaccinated against tumor
re-challenge via development of memory anti-tumor immunity. Cured
mice from FIG. 30 (n=1 for anti-PD-1 alone, n=2 for Seprehvir alone
and n=3 for the combination) were rechallenged by subcutaneous
implantation of M3-9-M cells. Tumors failed to form in any animal
but developed in 5/5 age-matched naive mice.
[0541] FIG. 36. Seprehvir directly interacts and activates human
PBMCs. Charts showing phenotypes of NK and CD4+ cells after
treatment with reovirus (reo), HSV1716, steroid or Braf inhibitor
or X-ray radiation treatment. PBMC were isolated from leukapheresis
cones, seeded at 2.times.10.sup.6 cells/ml and treated.+-.reovirus
or HSV1716 (MOI 1); dexamethasone (0.2 mM); PLX4720 (2 .mu.M); 2 Gy
XRT; 8 Gy XRT. After 24 or 48 h culture the cells were harvested,
stained for the markers indicated and analyzed by FACS. For each
entry 24 h data left bar shows 24 h data and right bar shows 48 h
data.
[0542] FIG. 37. Seprehvir directly interacts and activates human
PBMCs. Charts showing phenotypes of CD8+ cells after treatment with
reovirus (reo), HSV1716, steroid or Braf inhibitor or X-ray
radiation treatment. PBMC were isolated from leukapheresis cones,
seeded at 2.times.10.sup.6 cells/ml and treated.+-.reovirus or
HSV1716 (MOI 1); dexamethasone (0.2 mM); PLX4720 (2 .mu.M); 2 Gy
XRT; 8 Gy XRT. After 24 or 48 h culture the cells were harvested,
stained for the markers indicated and analyzed by FACS. For each
entry 24 h data left bar shows 24 h data and right bar shows 48 h
data.
[0543] FIG. 38. Seprehvir directly interacts and activates human
PBMCs. Charts showing phenotypes of CD14+ cells after treatment
with reovirus (reo), HSV1716, steroid or Braf inhibitor or X-ray
radiation treatment. PBMC were isolated from leukapheresis cones,
seeded at 2.times.10.sup.6 cells/ml and treated.+-.reovirus or
HSV1716 (MOI 1); dexamethasone (0.2 mM); PLX4720 (2 .mu.M); 2 Gy
XRT; 8 Gy XRT. After 24 or 48 h culture the cells were harvested,
stained for the markers indicated and analyzed by FACS. For each
entry 24 h data left bar shows 24 h data and right bar shows 48 h
data.
[0544] FIGS. 39A and 39B. Charts showing expression of cytokines in
PBMC after treatment with reovirus (reo), HSV1716, steroid or Braf
inhibitor or X-ray radiation treatment. PBMC were isolated from
leukapheresis cones, seeded at 2.times.10.sup.6 cells/ml and
treated.+-.reovirus or HSV1716 (MOI 1); dexamethasone (0.2 mM);
PLX4720 (2 .mu.M); 2 Gy XRT; 8 Gy XRT. After 24 or 48 h culture the
cells were harvested, stained for the markers indicated and
analyzed by FACS. (A) IL-6, IL-10, IFN.alpha., IFN.gamma.; (B)
TNF.alpha..
[0545] FIG. 40. Chart showing FACs analysis and table showing GFP
expression in tumor cell lines One58 (human mesothelioma), Ovcar3
(human ovarian cancer), T98 (human glioblastoma multiforme), Ln229
(human glioma). Human cancer cell lines were infected with
HSV1716gfp (HSV1716 modified to express GFP) at moi 0.5 and
cultured for 24 hours before addition of 10.sup.6 human PBMCs.
After 24 hours culture, PBMCs were decanted and cultured for 24-48
hours before analysed for expression of GFP. HSV1716 was found to
infect and transfer to a monocyte/macrophage rich subset of
PBMCs.
[0546] FIG. 41. HSV1716 infected human cancer cell lines stimulate
PBMC and Pleural Fluid Mononuclear Cell (PFMC) growth. Two separate
experiments are shown and PBMC/PFMC were decanted and cultured for
48 hours before MTS assay. Left chart, experiment 1 shows result of
MTS assay of PBMC/PFMC when added to human cancer cell lines
infected with HSV1716; in each entry from left to right bars
indicate T98 cells, One58 cells and Ovcar3 cells+/-virus infection.
Right chart shows experiment 2 result of MTS assay following
treatment of tumor cells lines Ln229, Ovcar3, T98 and One58 with
HSV1716 followed by the addition of PBMC/PFMC; in each entry, from
left to right bars indicate PBMC alone, PBMC+HSV1716, pleural fluid
mononuclear cells, pleural fluid mononuclear cells+HSV1716.
[0547] FIG. 42. Seprehvir preferentially infects monocytes in human
PBMCs. Charts results of FACs analysis for human monocytes and
lymphocytes infected with HSV1716gfp.
[0548] FIG. 43. Seprehvir infects and polarises human macrophages.
Charts show infection of macrophages with HSV1716gfp, macrophage
cell death, expression of viral genes in macrophages: ICP0, ICP8,
gB. All data were normalised to the house keeping gene GAPDH and 6
independent experiments were performed (n=6). X-axis 0=macrophages
(no virus).
[0549] FIG. 44. Charts showing expression of FasL, Bcl-2, LC3B and
Atg5 in macrophages 24 hours after infection with HSV1716 at moi of
5. All data were normalised to the house keeping gene GAPDH and 6
independent experiments were performed (n=6). X-axis 0=macrophages
(no virus).
[0550] FIG. 45. Charts showing mRNA expression of markers of
inflammation in human monocyte-derived macrophages 24 hours post
infection. All data were normalised to the house keeping gene GAPDH
and 6 independent experiments were performed (n=6). X-axis
0=macrophages (no virus).
[0551] FIG. 46. Charts showing mRNA expression of M1 macrophage
markers (NOS2, CXCL10) and M2 macrophage marker (MRC1) in human
monocyte-derived macrophages 24 hours post infection. All data were
normalised to the house keeping gene GAPDH and 6 independent
experiments were performed (n=6).
[0552] FIG. 47. Chart showing HSV1716 infection of 7 day human
monocyte derived macrophages significantly induces PCNA expression.
All data were normalised to the house keeping gene GAPDH and 4
independent experiments were performed (n=4).
[0553] FIG. 48. Chart showing oncolytic HSV therapy significantly
delays 975A2 mNB (murine neuroblastoma) tumor growth.
[0554] FIG. 49. Oncolytic HSV therapy recruits more T cells in
975A2 mNB tumor. Charts show cellular infiltrate into tumor of
different cell types. In all charts pairs of data points are shown
for treatment with PBS (left) and Seprehvir (right) for
administration of 1 (1.times.) or 3 (3.times.) doses.
[0555] FIG. 50. FACS analysis charts show expression of PD-L1 on
975A2 mNB cells.
[0556] FIG. 51. Oncolytic HSV therapy induces PD-L1 expression in
myeloid cells. Charts show PD-L1 expression in F4/80+ Macrophage
cells, myeloid derive suppressor cells (MDSCs) and neutrophils. For
bar charts, left bar=PBS, right bar=Seprehvir.
[0557] FIG. 52. Charts showing mean tumor volumes from data shown
for individual mice in FIG. 29 (left) demonstrates that the
combination of Seprehvir+anti-PD-1 is synergistic as the actual
combination effect on tumor growth is greater than the predicted
additive effect (right).
[0558] FIG. 53. Diagrammatic illustration of treatment cycles:
weekly, and twice weekly (one and two cycles).
[0559] FIG. 54. Charts showing immune cell proliferation and
IFN.gamma. levels in patient 09 (Example 1).
[0560] FIG. 55. Chart showing Seprehvir persistence in pleural
fluids (Example 1).
[0561] FIG. 56. Table showing summary of patient Th1, immune cell
and cytokine responses (Example 1).
[0562] FIG. 57. Table showing summary of patient anti-tumor IgG
responses.
[0563] FIG. 58. Table showing detection of HSV-1 DNA in patient
blood samples. "Pos"=positive for HSV-1 DNA, "Neg"=negative for
HSV-1 DNA.
[0564] FIG. 59. Chart showing titration of human macrophages at
various times after infection with 4 pfu/cell HSV1716 and culture
in normoxia or hypoxia. Approximately 300,000 primary human
macrophages were infected with 1,180,000 pfu HSV1716 with samples
collected at various times post infection and HSV1716 titrated on
Vero cells. Total titratable virus was graphed against time and the
dotted line represents the amount of input virus.
[0565] FIG. 60. Chart showing output (total pfu) from human
macrophages after 72 hrs of normoxia infection with HSV1716 at
various input moi. Approximately 300,000 primary human macrophages
were infected with HSV1716 at moi 40, 4, 0.4 and 0.04 with samples
collected at 72 hrs post infection only and HSV1716 titrated on
Vero cells.
[0566] FIG. 61. Table showing detection of HSV-1 DNA in patient
blood samples for 8 patients enrolled on NCT00931931.
IV=intravenous administration of Seprehvir, ITu=intratumoral
administration of Seprehvir, "Pos"=positive for HSV-1 DNA,
"Neg"=negative for HSV-1 DNA, nd=not done.
[0567] FIG. 62: Digital PET/CT images for patient HSV13 enrolled on
NCT00931931 at day 14 and day 28 post intravenous administration of
Seprehvir. Lesion is circled and SUV indicated.
[0568] FIG. 63: Digital PET/CT images for patient HSV13 enrolled on
NCT00931931 at day 14 and day 28 post intravenous administration of
Seprehvir. Transverse image through body. Lesion is circled and SUV
indicated.
[0569] FIG. 64: Digital PET/CT image for patient HSV07 enrolled on
NCT00931931 showing regions of tumor.
[0570] FIG. 65: Digital PET/CT images for patient HSV07 enrolled on
NCT00931931 prior to intratumoral administration of Seprehvir (top
left), at day 14 post intratumoral administration of Seprehvir (top
middle) two days prior to second intratumoral injection of virus on
6.27.14 (top right) and post second intratumoral injection (bottom
row).
[0571] FIG. 66. Table showing viability of SupT1, Toledo (ToIB) or
THP-1 cells 5 days after infection with either HSV1716 or HSV-1
17+.
[0572] FIG. 67. Charts showing FACS analysis of fresh human PBMC
fractions (monocytes or lymphocytes) mock infected (control) or
infected with HSV1716gfp (GFP) at moi 1.
[0573] FIG. 68. Chart showing stability of HSV1716 in PBS, whole
blood, plasma or cell fraction. PBS=top, approximately horizontal
line, whole blood=left most line at 2 minutes.
[0574] FIG. 69. Chart showing HSV1716 released for infection from
spiked whole blood, cell fraction or plasma during 72 hrs of
incubation with Vero cells. The dotted line represents the yield
from Vero cells infected with 10 pfu HSV1716. At each time point:
whole blood=left bar, cell fraction=middle bar, plasma=right
bar.
[0575] The details of one or more embodiments of the invention are
set forth in the accompanying description below including specific
details of the best mode contemplated by the inventors for carrying
out the invention, by way of example. It will be apparent to one
skilled in the art that the present invention may be practiced
without limitation to these specific details.
EXAMPLES
[0576] Malignant pleural mesothelioma (MPM) remains a major
challenge, with limited therapeutic options. Multifocal
intrapleural disease can cause disabling symptoms of pain and
breathlessness, in the absence of distant metastases, so an
intrapleural treatment approach is attractive.
[0577] SEPREHVIR.RTM. (HSV1716) is a mutant oncolytic herpes
simplex virus type 1 deleted in the RL1 gene which encodes the
protein ICP34.5, a specific determinant of virulence. Mutants
lacking the RL1 gene are capable of specific replication in cancer
cells and inducing anti-tumor immune responses. Clinical studies
with SEPREHVIR have been completed in adult glioma, melanoma,
squamous cell head and neck cancer, and studies are ongoing in
non-CNS solid tumors and MPM. In total, 98 patients have received
SEPREHVIR and the virus is well-tolerated with no spread to
surrounding normal tissue or no shedding in patients. SEPREHVIR
selectivity for replication only in tumor cells and intimations of
efficacy and immuno-stimulatory potential have been
demonstrated.
[0578] Cytokines are secreted intercellular signalling molecules
that regulate many different processes including inflammation, host
defence and cell differentiation. Cytokine profiles may help
understand changes in the pleural fluid samples in patients
following SEPREHVIR.RTM. administration.
[0579] Upon activation, naive CD4+ helper T cells differentiate
into distinct subsets. The development of the subsets is driven in
part by the cytokine milieu. Type 1 (Th1) cells help drive cellular
immunity against intracellular pathogens. IL-12 and IFN.gamma.
induce Th1 cell development. Th1 cells produce IFN-.gamma. and
IL-2, which provided a positive feedback loop to enhance Th1 cell
differentiation and NK cell and CD8+ T cell cytolytic activity.
[0580] Th2 cells play a crucial role in the humoral response
against extracellular pathogens. IL-4 drives development of Th2
cells, which subsequently produce IL-4, IL-5 and IL-13. These
cytokines induce B cell proliferation, antibody production, IgE
class switching and activate eosinophils respectively.
[0581] Another distinct helper T cell lineage, Th17 is important
for mucosal immunity. De-regulation of Th17 may significantly
contribute to the development of autoimmunity. IL-17 produced by
Th17 cells induces secretion of proinflammatory cytokines IL-6,
IL-8, GM-CSF and TNF.alpha.. Many of these molecules link innate
and adaptive immunity through the recruitment and activation of
innate immune cells.
[0582] Effective immune responses require finely tuned coordination
between pro- and anti-inflammatory signals. Proinflammatory
molecules play important roles in activating key immune players to
fight infection. IL-8 induces granulocyte migration and activates
neutrophil phagocytic activity. GM-CSF mobilizes monocytes into
infected tissue and activates macrophage and neutrophils.
TNF.alpha. is a multifunctional proinflammatory cytokine involved
with a number of processes including cell proliferation,
differentiation and apoptosis.
[0583] Uncontrolled inflammation may damage surrounding host
tissue. IL-10 is a prototypical anti-inflammatory cytokine that
serves to terminate the acute inflammatory response by inhibiting
Th1 cells function and pro-inflammatory cytokine production.
Example 1 --Cytokine Responses Following Intrapleural
Administration of Oncolytic HSV SEPREHVIR.RTM. in Patients with
Malignant Pleural Mesothelioma
[0584] We are currently conducting a phase I/11a trial at Cancer
Clinical Trials Centre, Weston Park Hospital, Sheffield and Queen
Elizabeth University Hospital, Glasgow, United Kingdom to determine
the safety and potential for efficacy of SEPREHVIR.RTM. given
intrapleurally to patients with malignant pleural mesothelioma
(MPM). Patients receive 1.times.10.sup.7 iu SEPREHVIR.RTM. through
their pleural catheter on one, two or four occasions each dose
given one week apart, in three separate patient cohorts. To date 11
patients have been treated, 3 in each 1 and 2 dose cohorts and 5 in
the 4 dose cohort and SEPREHVIR.RTM. has been well-tolerated with
few adverse events in any patients. An exploratory objective, to
assess tumor responses by CT using modified RECIST criteria, has
demonstrated disease stabilisation in 6/10 evaluable patients.
[0585] Pleural fluid and plasma samples have been collected pre-
and post-treatment and analysed to assess patient responses to
SEPREHVIR.RTM. administration.
1.1 HSV DNA
[0586] HSV DNA was detected in the pleural fluids of most patients
and in some persisted for at least two or four weeks
post-administration (FIG. 55).
1.2 Cytokine Analysis
[0587] Pleural fluid samples were collected from patients following
intrapleural administration of SEPREHVIR.RTM. and were analysed for
changes in the levels of the following cytokines, or potential
biomarkers: IFN-.gamma. (Interferon-gamma), IFN-.alpha.
(Interferon-alpha), the following Interleukins (IL): IL-1.alpha.,
IL-2, IL-4, IL-6, IL-10, IL-12, IL-21, IP-10 (IFN-.gamma. inducible
protein 10), MIG (monokine induced by IFN-.gamma.), TNF-.alpha.
(Tumor necrosis factor alpha), and VEGF (Vascular Endothelial
Growth Factor).
[0588] Changes in cytokine and chemokine levels may be indicative
of a developing immune response in the pleural space and changes in
potential biomarker levels may be indicative of patient responses
to treatment.
1.2.1 Materials and Methods
[0589] Commercially available ELISA kits were used to measure the
concentrations of these cytokines and potential biomarkers in
biological fluids. ELISA kits for quantifying cytokines, chemokines
and potential biomarkers in biological fluids were used exactly as
specified in the manufacturer's instructions. For example,
Novex.RTM. (Thermo Fisher) ELISA kits allow specific, quantitative
measurements of cytokines, chemokines and disease-related proteins
in various biological fluids. ELISA kits were selected on the basis
that they are compatible with biological fluids such as serum or
plasma.
[0590] For detection of human interferon-.gamma. ELISA Kit Cat#
KHC4021, 4022, 4021C (Invitrogen, Camarillo, Calif., USA) was used.
For detection of human VEGF ELISA Kit Cat# KHG0112, 0111
(Invitrogen, Camarillo, Calif., USA) was used.
[0591] Pleural fluid samples from patients were delivered on dry
ice, thawed and processed for subsequent analysis. 5-10 ml of each
pleural fluid were stored at -70.degree. C. in 15 ml centrifuge
tubes for analysis of cytokines and potential biomarkers.
[0592] Prior to using an ELISA kit, its compatibility with pleural
fluids and useful dilution range was tested. Two pleural fluids are
used for this test, one sample prior and one post administration of
SEPREHVIR.RTM. were diluted 1:10, 1:100 and 1:1000 using the
dilution buffer provided with the kit. One strip of eight wells was
removed from the kit and the undiluted, 1:10, 1:100 and 1:1000
dilutions for each samples were added to individual wells. The
ELISA protocol was then followed exactly as specified by the
manufacturer and the resultant OD450 nm readings identify the most
appropriate sample dilutions for use in the ELISA. The most
appropriate dilutions should generate an OD450 nm of between
0.5-1.5 within 15-30 mins. Pleural fluid samples were then analysed
at this appropriate dilution.
1.2.2 Results Detection of changes in levels of cytokines and
biomarkers (see FIGS. 1 to 14).
Th1 Associated Cytokines
IL-2:
[0593] Patients receiving 4 doses of SEPREHVIR.RTM. showed an
increase in IL-2 production (FIG. 5).
IL-12:
[0594] Patients receiving 4 doses of SEPREHVIR.RTM. showed an
increase in IL-12 production (FIG. 9).
[0595] IL-12, produced by dendritic cells, macrophages and human
B-lymphoblastoid cells, is known as a T cell stimulating factor and
involved in the differentiation of naive T cells into Th1 cells.
IL-12 is important within the immune response with various
activities including mediating the enhancement of the cytotoxic
activity of NK cells and CD8+ cytotoxic lymphocytes, stimulating
production of IFN-.gamma., TNF-.alpha. from T-cells and reduces
IL-4 mediated suppression of IFN-.gamma..
[0596] IL-12 has been shown to have anti-angiogenic abilities by
increasing production of IFN-.gamma. which causes the increased
production of IP-10, which mediates an anti-angiogenic effect.
IFN-.gamma.:
[0597] IFN-.gamma. levels were notably increased from low initial
levels in patients receiving single and multiple doses of
SEPREHVIR.RTM. (FIG. 2).
[0598] IFN-.gamma. functions include enhancing the cytotoxic
activity, activation, growth and differentiation of T-cells,
macrophages and NK cells. As well as the activation of other cells
types such as B-cells and fibroblasts. IFN-.gamma. production is a
characteristic of Th1 differentiation and promotes a Th1 immune
phenotype by causing naive CD4+ cells (Th0) to differentiate into
Th1 cells while suppressing Th2 cell differentiation. IFN-.gamma.
further enhances the immune response by stimulating macrophages
which upregulates antigen processing and presentation pathways,
promoting CD4+T cell activation and cell-mediated immunity. Through
upregulation of various cells, IFN-.gamma. directs the flow of
specific immune cells to the site of inflammation or infection
(Boehm, U., Klamp, T., Groot, M., Howard, J. C. (1997) Cellular
responses to interferon-gamma. Annu. Rev. Immunol. 15,
749-795).
[0599] IFN-.gamma. produced by APC (antigen presenting cells) that
secrete IFN-.gamma. may stimulate the self-activation and
activation of nearby cells. The production of IFN-.gamma. is
controlled by various cytokines, importantly IL-12 and IL-18
(Frucht, D. M., Fukao, T., Bogdan, C., Schindler, H., O'shea, J.,
Koyasu, S. (2001) IFN-gamma production by antigen-presenting cells:
mechanisms emerge. Trends Immunol. 22, 556-560). These cytokines
serve roles within the innate immune response, IL-12 is secreted by
macrophages which then attract NK cells to the site, while IL-12
continues to promote IFN-.gamma. synthesis. IFN-.gamma. is
negatively regulated by IL-4 and IL-10.
IP-10:
[0600] Patients receiving single and multiple doses showed a strong
upregulation of IP-10 (FIG. 12).
[0601] Interferon gamma-induced protein 10 (IP-10) is a chemokine
secreted by various cell types including monocytes, endothelial
cells and fibroblasts in response to IFN-.gamma.. IP-10 has various
roles within the immune system, arguably the most important of role
is being a potent chemoattractant for monocytes/macrophages, T
cells, NK cells and dendritic cells, IP-10 promotes anti-tumor
activity and inhibition of angiogenesis (Dufour. J. H., Dziejman.
M., Liu. M. T., Leung. J. H., Lane. T. E., Luster. A. D. (2002)
IFN-.gamma.-Inducible protein 10(IP-10) deficient mice reveal a
role for IP-10 in effector T cell generation and trafficking. Jour
Immunology. 168. 7. 3195-3204). IP-10 and other members of the
chemokine family including MIG, CXCL9, CXCL11 and CXCL4 have been
proposed as a therapeutic agent in the fight against cancer as they
induce injury to established tumor associated vasculature and
promote tumor necrosis (Homey, B., A. Muller, and A. Zlotnik. 2002.
Chemokines: agents for the immunotherapy of cancer? Nat. Rev.
Immunol. 2:175-184).
MIG:
[0602] Analysis of pleural fluid cytokines by AbCam ELISA indicated
that baseline levels of MIG (before treatment with SEPREHVIR.RTM.)
were high. Samples were diluted 1:100 before assay. Patients
receiving single and multiple doses showed a strong upregulation of
MIG (FIG. 13).
[0603] Monokine induced by gamma interferon (MIG), closely related
to the chemokine CXCL10, is a T cell and NK cell bearing the
chemokine receptor CXCR3 chemoattractant (Walser. C. T., Xinrong.
M., Kundu. N., Dorsey. R., Goloubeva. W. O., Fulton. M. A. (2007)
Immune-mediated Modulation of Breast Cancer Growth and Metastasis
by the Chemokine Mig (CXCL9) in a Murine Model. J Immunother 2007;
30:490-498). CXCR3 can regulate leukocyte trafficking, attracts Th1
cells and promotes Th1 cell maturation. MIG has been shown to have
anti-tumor activity in a number of tumor models as well as
stimulating T cells to the site of injury and having anti
angiogenic properties (Saudemont A, Jouy N, Hotuin D, et al. NK
cells that are activated by CXCL10 can kill dormant tumor cells
that resist CTL-mediated lysis and can express B7-H1 that
stimulates T cells. Blood. 2005; 105:2428-2435). Furthermore, there
is evidence to suggest NK cells that have been stimulated by MIG
have the potential to kill dormant tumor cells that have previously
been resistant to cell death (Saudemont. A., Jouy. N., Hetuin. D.,
Quesnel. B. (2005) NK cells that are activated by CXCL10 can kill
dormant tumor cells that resist CTL-mediated lysis and can express
B7-H1 that stimulates T cells. Blood. Vol 15. 6. 2428-2435).
TNF-.alpha.:
[0604] Patients showed a small increase in TNF-.alpha. production
(FIG. 11).
[0605] Tumor necrosis factor alpha is a multifunctional
inflammatory cytokine produced by macrophages/monocytes during
inflammation and implicated in signalling events that lead to cell
necrosis and apoptosis (Idriss. H. T and Naismith. H. J. (2000)
TNF.alpha. and the TNF receptor subfamily: Structure-function
relationship(s). Microscopy research and technique. 50. 184-195).
Although the exact mechanism is unknown, TNF.alpha. is critical in
efficient T cell immune responses, affecting T cell priming,
proliferation, recruitment and function. The link between
anti-TNF.alpha. therapies and increased incidence of malignancies
in Rheumatoid Arthritis patients has suggested a link between
TNF.alpha. in the development, progression and immune surveillance
of tumors as well as potentially possessing anti-tumor properties
(Calzascia T, Pellegrini M, Hall H, et al. TNF-.alpha. is critical
for antitumor but not antiviral T cell immunity in mice. The
Journal of Clinical Investigation 2007; 117(12):3833-3845.
doi:10.1172/JC132567).
Proinflammatory Cytokines
IL-6:
[0606] Analysis of pleural fluid cytokines by ELISA indicated that
baseline levels of IL-6 (before treatment with SEPREHVIR.RTM.) were
high. Samples were diluted 1:1000 before assay. In most patients,
even at multiple doses IL-6 levels did not rise notably compared to
baseline levels (FIG. 7).
[0607] Detection of high levels of IL-6 is consistent with previous
reports of detection of IL-6 in patients having malignant pleural
mesothelioma (T Nakano et al., Interleukin 6 and its relationship
to clinical parameters in patients with malignant pleural
mesothelioma. British Journal of Cancer (1998) 77(6), 907-912; Siti
N. Abdul Rahim et al., The role of interleukin-6 in malignant
mesothelioma Transl Lung Cancer Res 2015; 4(1):55-66).
[0608] IL-6 is a pro and anti-inflammatory cytokine which is
produced by a variety of cells such as T cells, B cells monocytes,
fibroblasts and keratinocytes and macrophages. IL-6 stimulates a
broad range of cellular and physical responses in the event of
infection or trauma. Recent research suggests IL-6 along with
TNF.alpha. and IL-1, are major proinflammatory cytokines, IL-6 is
an important modulator of CD4 T cell effector functions therefore
impacting the immune response and contributing to inflammation
(Dienz. O., Rincon. M. (2009). The effect of IL-6 on CD4 T cell
responses. Clin Immunol. 130(1): 27-33). In response to PAMPS
(pathogen-associated molecular patterns), which are located on the
cell surface and intracellular compartments, IL-6 is produced by
macrophages, causing a signalling cascade that produces an
inflammatory cytokine production. IL-6 may protect CD4 T cells from
undergoing apoptosis and stimulates T cell activation as well as T
cell migration. A major function of IL-6 is antibody induction
(Akira. S., Hirano. T., Taga. T., Kishimoto. T. (1990) Biology of
multifunctional cytokines: IL6 and related moplecules (IL1 and
TNF). The FASEB Journal. 4. 11. 2860-2867).
IL-1.alpha.:
[0609] IL-1.alpha. levels were essentially unchanged in patients
receiving single or multiple doses of SEPREHVIR.RTM. compared to
baseline levels (FIG. 4).
[0610] IL-1.alpha. possesses a strong proinflammatory effect.
IL-1.alpha. is multifunctional and produced by tissue macrophages,
monocytes, fibroblasts and dendritic cells. IL-1.alpha. enables
transmigration of immunocompetent cells to sites of infection and
considered a central cytokine in the regulation of immune
responses. The release of IL-1.alpha. can induce activation of NFkB
which will promote cell survival, proliferation and angiogenesis
(Wolf. J. S., Chen. Z., Dong. G., Sunwoo. J. B., Bancroft. C. C.,
Capo. D. E., Yeh. N. T., Mukaida., Waes. C. V. (2001) IL
(Interleukin)-1a Promotes Nuclear Factor-kB and AP-1-induced IL-8
Expression, Cell Survival, and Proliferation in Head and Neck
Squamous Cell Carcinomas. Clin Cancer Res. 7. 1812-1820).
Th2 Associated Cytokines:
IL-4:
[0611] IL-4 levels were essentially unchanged in patients receiving
single of multiple doses of SEPREHVIR.RTM. compared to baseline
levels (FIG. 6).
[0612] IL-4 stimulates the differentiation of naive T cells (Th0
cells) to effector T cells (Th2 cells), subsequently Th2 cells
produce additional IL-4 and have a role in a class switch response
to IgG1 and IgE isotopes of B-cells (Kabsech. M., Schedel. M.,
Carr. D., Woitsch. B., Fritzsch. C., Weiland. S. K., Mutius. E.
(2006) IL-4/IL-13 pathway genetics strongly influence serum IgE
levels and childhood asthma. Journal of Allergy and Clinical
Immuno. Vol 117. 2. 269-274). One of the biological activities of
IL-4 is the stimulation of activated B-cell and T-cell
proliferation. IL-4 is considered a key regulator in humoral and
adaptive immunity. IL-4 is known to decrease the production of Th1
cells, IFN gamma, macrophages and dendritic cell IL-12.
IL-10:
[0613] Patients receiving 4 doses of SEPREHVIR.RTM. showed an
increase in IL-10 production (FIG. 8). Although IL-10 is associated
with Th2 cells it acts to regulate the Th1 response, preventing an
excessive Th1 response. Its upregulation in patients exhibiting a
more pronounced Th1 response is consistent with this regulatory
function and confirms the authenticity of the Th1 response.
[0614] IL-10 is an anti-inflammatory cytokine primarily produced by
monocytes and to a lesser extent by Th2 lymphocytes, mastocytes and
in certain activated T and B cells. IL-10 limits the production of
proinflammatory cytokines (including IL-12, IL-6, IL-1.alpha.,
TNF.alpha., IL-8 and IP-10), resulting in the indirect inhibition
of Th1 cells (Couper. K. N., Blount. D. G., Riley. E. M. (2008)
IL-10: The master regulator of immunity to infection. Jour Immunol.
180. 5771-5777). However IL-10 can directly act on CD4+T cells
causing an inhibition of proliferation and production of IL-2,
IFN-.gamma., IL-4, IL-5 and TNF a, allowing IL-10 to directly
regulate the innate and adaptive Th1 and Th2 responses by limiting
T cell activation while inhibiting pro inflammatory responses
(Moore, K. W., R. de Waal Malefyt, R. L. Coffman, A. O'Garra. 2001.
Interleukin-10 and the interleukin-10 receptor. Annu. Rev. Immunol.
19: 683-765).
[0615] During infection IL-10 both regulates and inhibits pro
inflammatory cytokines to help prevent tissue damage which would
result in the over production of pro inflammatory cytokines.
Other Cytokines and Biomarkers:
IFN-.alpha.:
[0616] IFN-.alpha. levels were essentially unchanged in patients
receiving single of multiple doses of SEPREHVIR.RTM. compared to
baseline levels (FIG. 3).
[0617] Classed within the type I IFN family of interferons,
IFN-.alpha. are produced by dendritic cells in response to viral
infection and have immunomodulatory functions which causes immune
cell differentiation, activation and survival (Padovan, E.,
Spagnoli. G., Ferrantini. M., Heberer. M. (2002) IFN-.alpha.2a
induces IP-10/CXCL10 and MIG/CXCL9 production in monocyte-derived
dendritic cells and enhances their capacity to attract and
stimulate CD8.sup.+effector T cells. Journal of Leukocyte
Biologyvol. 71 no. 4 669-676).
VEGF:
[0618] VEGF levels increased in some patients but it was notable
that baseline levels of VEGF in these patients (before treatment
with SEPREHVIR.RTM.) were high (FIG. 14).
[0619] Vascular endothelial growth factor is a signal protein that
stimulates angiogenesis and vasculogenesis, VEGF is considered to
be an important factor in tumor growth (Carmeliet. P. (2005) VEGF
as a key mediator of angiogenesis in caner. Oncology. 69. 3. 4-10).
VEGF production can be induced in cells that are lacking oxygen,
released VEGF triggers a tyrosine kinase pathway leading to
angiogenesis, leading VEGF to be a potential target in the
treatment of cancer (Ohm, J., Gabrilovich. D., Sempowski. G.,
Kisseleva. E., Parman. K., Nadaf. S., Carbone. D. (2003) VEGF
inhibits T-cell development and may contribute to tumor-induced
immune suppression. Blood. 101. 12). VEGF has been shown to promote
monocytes/macrophage migration and increase the production of B
cells, however VEGF has also been shown to inhibit T cell
production and over all reducing immune cell function (Ferrara. N.,
Gerber H., LeCouter. J. (2003) The biology of VEGF and its
receptors. Nature Medicine 9, 669-676).
IL-21:
[0620] Some patients exhibited a small increase in IL-21 levels
(FIG. 10).
Induction of a Th1 Response Varies Between Patients
[0621] In some patients Seprehvir replicated and persisted in the
pleural fluid but did not induce a Th1 response, e.g. patient 02
(FIGS. 21 and 27), patient 05 (FIGS. 26 and 27).
[0622] In other patients Seprehvir was undetectable in the pleural
fluid but induced a robust Th1 response, e.g. patient 03 (FIGS. 22
and 27). The absence of detectable HSV DNA in the pleural fluid is
not inconsistent with an HSV mediated treatment effect. The levels
of the virus in the pleural fluids could be below the detection
limit of 100 pfu equivalents/ml or it could be persisting and
spreading intracellularly. In other studies we have noted viral
adsorbtion by cells and tissue within 1 hour of administration to
patients such that virus was not detectable in blood for a period
of time after which HSV DNA was detectable. This is consistent with
adsorption of virus following administration and re-emergence later
in fluid samples after build up of productive infection within the
tumor tissue.
[0623] Proliferation of immune cells and IFN.gamma. production was
measured in pleural fluid samples from patient 09 on day 1
(pre-treatment) and at days 11, 17, 22 and 29 post-administration
of Seprehvir (patient 09 received four doses of Seprehvir on days
1, 11, 15 and 22). Cells were stimulated in the presence of
anti-CD3 antibody and the whole cell extract from an MPM cell line
(MSTO211H). Control cells were incubated with the anti-CD3
antibody+PBS. Proliferation was measured by MTS assay and
proliferation was calculated as MTS at 96 hrs/MTS at 0 Hrs.
IFN.gamma. was measured in culture supernatants by ELISA at 96 hrs.
Results are shown in FIG. 54.
Comments
[0624] Analysis of pleural fluid cytokines by ELISA indicated that
SEPREHVIR.RTM. administration was associated with Th1-type
responses with increased levels of IFN.gamma., IP-10 and TNF.alpha.
and accompanied by increased levels of IL-10 in most patients.
[0625] Analysis of pleural fluid cytokines by ELISA indicated that
they were generally rich in IL-6, MIG and VEGF. Pleural fluids had
high levels of IL-6 and IL-12 and, in most patients, there were
moderate increases of both post SEPREHVIR.RTM. administration.
Pleural fluids were also rich in VEGF and levels increased in 4/9
patients post SEPREHVIR.RTM. administration.
[0626] IL-1.alpha., IL-4 and IFN.alpha. were not detected
pre-treatment and showed no response to SEPREHVIR.RTM.
administration.
[0627] Post SEPREHVIR administration there were increased levels of
IFN-.gamma., IP-10, MIG, TNF.alpha. and IL-6 in most patients,
including patients receiving only one dose of SEPREHVIR.RTM.. IL-2,
and IL-12 increases were most notable in patients receiving 4 doses
of SEPREHVIR.RTM..
[0628] Overall, these responses are consistent with development of
a Th1 response.
[0629] IL-1.alpha., IL-4, IL-21, and IFN.alpha. were not detected
pre-treatment and showed little or no response to SEPREHVIR.RTM.
administration, consistent with lack of development of a Th2
response.
Example 2--Novel Anti-Tumor Serum IgG Response
[0630] Patient serum samples were used to probe cell extracts in
order to investigate the possibility of an anti-tumor antibody
response to treatment with SEPREHVIR.RTM.. Cell extracts were
prepared from cell lines: MSTO-211H (mesothelioma; ATCC CRL-2081),
SPC111 (mesothelioma), and HuH7 (hepatic carcinoma), and contacted
with patient sera. IgG:target complexes were detected using an
anti-human IgG antibody by standard Western Blotting procedures
(FIG. 15).
[0631] Analysis of plasma samples indicated a strong anti-HSV IgG
responses post SEPREHVIR.RTM. administration, particularly after 2
and 4 doses. Intrapleural administration of SEPREHVIR.RTM. was
found to induce a novel anti-tumor IgG response against an antigen
present on MSTO-211H cells but not on SPC111 or HuH7 cells (FIG.
16-18) more so in patients receiving 4 doses of Seprehvir.
[0632] Thus, SEPREHVIR.RTM. has immunotherapeutic potential capable
of inducing novel anti-tumor immune responses in patients. This
result is consistent with induction of IgG B cells directed to
tumour antigens released during Seprehvir oncolysis and stimulated
through a Th1 response.
Example 3--Checkpoint Blockade Enhances Oncolytic Herpes
Virotherapy in Immunosuppressive Sarcoma Models
[0633] Most solid tumors are characterized by an immunosuppressive
microenvironment, limiting the effectiveness of antitumor
immunotherapeutics. Programmed cell death protein (PD)-1-mediated T
cell suppression via engagement of its ligand, PD-L1, is of
particular interest due to recent successes in selected adult
cancers. The utility of PD1-directed therapy for pediatric cancers
is unknown, especially given the paucity of mutations and thus
infrequent neoantigens in many types of childhood tumors. Oncolytic
virotherapy induces tumor shrinkage via a multistep process
including direct tumor cell lysis, induction of cytotoxic or
apoptosis-sensitizing cytokines, and induction of antitumor T cell
responses. We have demonstrated that intratumoral injection of an
oncolytic herpes virus induced growth delays and in some cases
durable remissions in several mouse models of rhabdomyosarcoma. The
effects were T cell-mediated, as surviving mice were resistant to
tumor rechallenge and efficacy was lost in athymic nude hosts. We
found these tumor models express PD-L1, suggesting that T cell
checkpoints may in part limit virus-induced antitumor immunity.
Here we show the implantable C57BL/6 rhabdomyosarcoma model,
M3-9-M, showed a moderate response to single-agent Seprehvir
(HSV1716), a virus currently in pediatric clinical trials
(NCT00931931). Single-agent PD-1 blockade also showed moderate but
significant tumor growth delay with no complete responses.
Combining these two therapies together substantially prolonged
overall survival with several complete responses post 60 days
treatment. Interestingly, mice that received combination therapy
showed more CD4+/CD8+ T cell recruitment to the tumor and displayed
higher immune inflammatory responses and a less immunosuppressive
microenvironment, as measured by the decreased proportion of
CD4+/CD25+/Fox3P+ Tregs and suppressive cytokines. Overall, our
data suggest the combination of PD-1 and oncolytic herpes
virotherapy may be an effective treatment strategy for some
cancers. Results are shown in FIG. 28-33.
[0634] We observed that: combination of oHSV treatment with immune
checkpoint inhibitor anti-PD-1 significantly prolongs survival in
both male to male and male to female rhabdomyosarcoma models;
greater antitumor efficacy was observed in male to female murine
rhabdomyosarcoma, suggesting that combination therapy favors more
immunogenic microenvironments; combination therapy resulted in more
CD4+/CD8+ T cell recruitment but did not affect in vivo virus
activity; combination therapy induces more inflammatory responses
and, although CD4+ T cell numbers increased, CD25+/CD4+ Treg
numbers were unchanged thus lowering the regulatory/suppressive
tumor microenvironment.
Experimental Methods and Results
[0635] C57BL/6 mice were injected with 5.times.10.sup.6 M3-9-M
cells subcutaneously. Tumors were treated intra-tumorally (i.tu.)
with Seprehvir when sizes reached 200-400 mm.sup.3.
Intra-peritoneal (i.p.) injection of anti-PD-1 antibody [anti-PD1
antibody rat monoclonal RMP1-14 (AbCam plc)] were given twice a
week after last dose of virus treatment. Tumor growth was monitored
twice a week. Mice were sacrificed when tumors reached 2,500
mm.sup.3 in volume or grew over 2 cm in length.
[0636] Female C57BL/6 mice were injected with 5.times.10.sup.6
M3-9-M cells subcutaneously. The effects of Seprehvir plus
anti-PD-1 blockade on antitumor efficacy were evaluated by
measuring tumor volumes over time. Survival data were evaluated for
statistical significance with Log-rank (Mantel-Cox) test. FIG. 29
shows the combination of Seprehvir and anti-PD-1 antibody to
significantly prolong survival with several complete responses in
the male to female M3-9-M tumor model.
[0637] Male C57BL/6 mice were injected with 5.times.10.sup.6 M3-9-M
cells subcutaneously. The effects of Seprehvir plus anti-PD-1
blockade on antitumor efficacy were evaluated by measuring tumor
volumes over time. Survival data were evaluated for statistical
significance with Log-rank (Mantel-Cox) test. FIG. 30 shows the
combination of Seprehvir and anti-PD-1 antibody to significantly
prolong survival in the less immunogenic male to male M3-9-M tumor
model.
[0638] Female M3-9-M tumor-bearing mice were treated with three
doses of 10.sup.8 pfu of Seprehvir intra-tumorally (i.tu.) followed
by intra-peritoneal (i.p.) injection of anti-PD-1 or control
antibody. Tumors were harvested 3, 24, 72 and 168 hours after
intra-peritoneal antibody injection for plaque assay. Data are
expressed as total plaque-forming units (pfu) per tumor. FIG. 31
shows checkpoint inhibition does not significantly alter
intra-tumoral viral kinetics.
[0639] Female M3-9-M tumor-bearing mice received three doses of
intra-tumoral (i.tu.) Seprehvir injection followed by
intra-peritoneal (i.p.) injection of anti-PD-1 or control antibody.
Immune cell infiltrates in tumors were evaluated by flow cytometry
analyses 72 hours post intra-peritoneal antibody injection. FIGS.
32A and 32B shows combination therapy induces more CD25+CD8+ memory
T cells but less CD25+CD4+ Treg cells.
[0640] Female M3-9-M tumor-bearing mice received three doses of
intra-tumoral (i.tu.) Seprehvir injection followed by
intra-peritoneal (i.p.) injection of anti-PD-1 or control antibody.
Tumors were harvested 72 hours post intra-peritoneal antibody
injection. T-bet (Th-1-related gene), Foxp3 (Treg-related gene),
IFN.gamma., IL-10, iNOS (M1 macrophage-related gene) and MRC-1 (M2
macrophage-related gene) were quantified by real-time PCR. FIG. 33
shows combination therapy induces higher inflammatory gene
expression and lower immune suppressive gene expression. Data are
represented as relative RNA expression to gapdh.
[0641] Combination of oncolytic HSV treatment with immune
checkpoint inhibitor anti-PD-1 significantly prolonged survival in
both male to male and male to female rhabdomyosarcoma models.
[0642] Greater antitumor efficacy was observed in male to female
murine rhabdomyosarcoma, suggesting that combination therapy favors
more immunogenic microenvironments.
[0643] Combination therapy did not result in more T cell
recruitment or affect in vivo virus activity.
[0644] Combination therapy induces more inflammatory responses with
less immune regulatory/suppressive responses.
Example 4--Seprehvir Directly Polarises PBMCs Phenotype to Th1
[0645] When human PBMCs were exposed directly to Seprehvir, the
virus induced a marked Th1 phenotype with increased production of
IFN.alpha. and IFN.gamma. and TNF.alpha.. IL-6 and regulatory IL-10
production were also stimulated and HSV was more effective than
Reovirus, dexamethasone, PLX4720 and ionising radiation. Thus
Seprehvir could influence these cells directly following their
recruitment into the tumour microenvironment. Exposure of PBMCs to
Seprehvir upregulated the expression of immune checkpoints in many
different subsets including NK, CD4+, CD8+ and CD14+ (monocytes)
cells (FIGS. 36 to 39).
Example 5 Seprehvir Infects and Polarises Human Macrophages
Potentially Inducing a Th1 Response Directly in Human PBMCs
[0646] On Day 7 following infection with HSV1716 expressing gfp,
human monocyte-derived macrophages demonstrated a significant
increase in infection which correlated with an increase in cell
death.
[0647] Infection was demonstrated via investigation of the
expression of viral proteins immediate early (ICP0) and late (gB)
genes indicating significant gene expression in macrophages (FIG.
43).
Mechanism of Cell Death in Human Macrophages
[0648] HSV1716 kills macrophages via apoptosis and in a Fas
dependent manner with both FasL and Bcl-2 gene expression
up-regulated 24 hours after infection with HSV1716 at an MOI of
5.
[0649] Consistent with this observation, expression of genes
involved in autophagy (Atg5 and LC3B) were not significantly
altered (FIG. 44).
HSV1716 Infection Induces an Inflammatory Phenotype in
Macrophages
[0650] HSV1716 infection of day 7 monocyte-derived macrophages
significantly induces mRNA expression of typical markers of
inflammation 24 hours post infection with significantly increased
expression of IL-6, IL-8, TNFalpha. Expression of IL-10, TGFbeta
and NFkappaB were not significantly enhanced (FIG. 45).
HSV1716 Infection Induces an Inflammatory Phenotype in
Macrophages
[0651] HSV1716 infection of day 7 monocyte-derived macrophages
significantly induces mRNA expression of typical inflammatory M1
macrophage markers (NOS2, and CXCL10) and significantly down
regulated expression of the M2 marker MRC1 expressed by
tumour-derived macrophages (FIG. 46). There were no significant
changes in two other M2 markers, Arg1 and VEGF.
HSV1716 Infection Induces PCNA Expression in Macrophages
[0652] HSV1716 infection of day 7 monocyte-derived macrophages
significantly induces PCNA expression which therefore could be a
potential mechanism for inducing viral replication, macrophage cell
death and M2 to M1 switching in tumor-dwelling monocytes and other
myeloid derived suppressor cells. Further studies are currently
being pursued to investigate siRNA knockdown of PCNA prior to
HSV1716 infection (FIG. 47).
[0653] Taken together, examples 4 and 5 suggest that Seprehvir is
capable of inducing Th1 responses leading to anti-tumor immunity by
two separate but interdependent mechanisms. Oncolytic replication
of cancer cells within the tumor microenvironment induces an
inflammatory response which attracts circulating immune cells and
initiates a Th1 response. Both tumor-resident and newly recruited
immune cells, especially monocytes and other myeloid-derived cells,
such as M2 macrophages (TAMs, tumor associated macrophages) are
susceptible to Seprehvir infection in this localised inflammatory
milieu. Their subsequent infection by Seprehvir progeny released
from lytic replication of cancer cells amplifies this Th1 response
as for example, macrophages are polarised to the aggressive
anti-tumor M1 phenotype. Tumor antigens released by oncolysis and
picked up by activated antigen presenting cells leads to the
development of anti-tumor immunity.
Example 6--a Phase I Study Investigating the Safety, Tolerability
and Efficacy of Intravenous Injections of the Selectively
Replication-Competent Herpes Simplex Virus Seprehvir in Patients
with Relapsed or Refractory Solid Tumours
Summary of Clinical Experience
[0654] To date ninety eight patients have received Seprehvir, in
the context of locally advanced disease, via a variety routes,
mostly intra-tumoural (n=83) and the remainder via intrapleural
(n=11) or intravenous (n=4) infusions, in the absence of any
definitely attributable Seprehvir-related toxicity.
[0655] Forty seven patients with brain tumours have received a
range of Seprehvir doses (10.sup.3 to 2.times.10.sup.6)
intratumorally (n=35) or peri-tumorally post resection (n=12) in 4
clinical studies, three in primary or recurrent glioma and 1 in
recurrent glioblastoma multiforme (GBM). No induction of
encephalitis or any re-activation of latent wild type HSV was
observed and no adverse clinical symptoms attributable to Seprehvir
were identified.
[0656] Two further clinical studies of Seprehvir have been
completed. The first of these, a study in melanoma patients
involved five patients with metastatic melanoma and accessible soft
tissue tumor nodules. No local or systemic toxicity associated with
Seprehvir was observed.
[0657] The second of these studies involved 20 patients with
resectable squamous cell carcinoma of the head and neck in which
patients received a single preoperative intratumoral injection
(either 1, 3 or 14 days prior to surgery) with Seprehvir at a dose
of 10.sup.5 i.u. (5 patients) or 5.times.10.sup.5 i.u. (15
patients). No toxicity was experienced by any of the patients and
evidence of virus in tumor tissue was observed.
[0658] Two clinical studies are currently on-going.
[0659] A Phase I/11a study in Malignant Pleural Mesothelioma is
investigating the safety, tolerability and biological effect of
single and repeat intra-pleural administration of Seprehvir at a
dose of 1.times.10.sup.7 iu. To date, three patients have received
a single dose of Seprehvir through their IPC, three have received
two doses and five have received four doses with recruitment of an
additional one patient required at the four dose level to complete
the trial. Seprehvir is well-tolerated with a limited number of
transient possibly-related adverse events identified.
[0660] In the Phase I dose escalation study in non-CNS tumours,
three patients have received a single intratumoral administration
of 1.times.10.sup.5 i.u. Seprehvir, two patients have received a
single intratumoral administration of 2.times.10.sup.6 i.u.
Seprehvir, one patient has received a single intratumoral
administration of 2.times.10.sup.6 i.u. Seprehvir on two separate
occasions and three patients have received a single intratumoral
administration of 1.times.10.sup.7 i.u. Seprehvir to date. The
intratumoral arm of this study is now closed to recruitment.
Study Rationale
[0661] Seprehvir is an oncolytic virus that replicates in and lyses
the dividing cells of tumours but fails to replicate in normal
post-mitotic cells. Seprehvir also has anti-cancer vaccination
potential with induction of anti-tumour immune responses observed
in mesothelioma (MPM) patients
[0662] Based on this selective replication phenotype and the lack
of attributable toxicity noted in preclinical systemic dosing
models, coupled with the clinical safety profile demonstrated in 96
patients treated by localised Seprehvir delivery, a study in
patients with recurrent/metastatic advanced solid tumours is
proposed. The starting dose will be 1.times.10.sup.7 i.u. based on
the current loco-regionally administered dose used in our MPM study
and supported by a murine biodistribution studies and the maximum
dose FDA-approved doses to be used in the systemic arm of the study
in non-CNS solid tumours in children and adolescents.
[0663] Since it is considered highly relevant to analyse tumour
tissue for evidence of Seprehvir replication and cell lysis,
pre-tumour biopsies and post treatment biopsy or resection will be
conducted for all patients.
Objectives and Endpoints
[0664] Primary objective: To evaluate the safety, tolerability and
tumour localisation of repeat IV administration of Seprehvir in
patients with relapsed or refractory solid tumours
[0665] Secondary objective: To evaluate the patient's immunological
response post-Seprehvir administration
[0666] Primary endpoint: Safety and tolerability, in terms of the
emergence of DLTs, will be assessed by conducting the following
safety assessments at pre-defined time-points during the study:
[0667] Physical examination, including vital signs [0668] ECG
[0669] Analysis of laboratory parameters as follows. [0670]
Haematology: full blood count including differential white cell
count, haemoglobin, and haematocrit; coagulation parameters
including prothrombin time (PT) and activated partial
thromboplastin time (APPT) [0671] Biochemistry: urea, creatinine,
sodium, potassium, total protein, total bilirubin, alanine
aminotransferase (ALT), aspartate aminotransferase (AST),
.gamma.-glutamyltranspeptidase, lactate dehydrogenase (LDH),
alkaline phosphatase, albumin, calcium, phosphorus, glucose,
creatine kinase [0672] Viral shedding in urine and buccal swab
samples
[0673] Evidence of Seprehvir replication will be assessed using
plasma/serum samples and tumour tissue by: [0674] PCR for the
detection of Seprehvir genomes [0675] IHC of Seprehvir antigens in
biopsy/resected tissue
[0676] Adverse events will be recorded throughout the study
period.
[0677] Secondary endpoints: The immune response to Seprehvir
administration will be assessed by conducting the following at
pre-defined time-points during the study: [0678] Measurement of
circulating anti-HSV IgG and IgM in plasma samples [0679] Analysis
of circulating and tumour localised immune cells using immune cell
profiling and emergence of anti-tumour immune responses [0680]
Pharmacodynamic assessments in plasma/serum samples and tumour
tissue [0681] Tumour markers (CEA, Ca19-9, Ca15-3, Ca125, LDH, PSA
as appropriate) [0682] Biomarkers of Seprehvir activity to include
but not limited to IFNgamma and related Th1 cytokines and
chemokines, HMGB1, HSP70 and 90 [0683] Histology and
immunohistochemistry for necrosis, apoptosis, immune
infiltration
Study Design
[0684] This Phase I study will run at two sites in the UK.
[0685] This is a Phase I, open-label, dose-escalation study to
evaluate the safety, tolerability and tumour localisation of
Seprehvir, a selectively replication-competent herpes simplex
virus, administered IV in 36-40 patients with histologically
confirmed unresectable advanced or metastatic solid tumours that
are refractory to standard therapy.
[0686] The study will follow a 3+3 design to explore the safety and
tolerability and tumour localisation of up to 8.times.IV
administrations of Seprehvir, at 2 dose levels (1.times.10.sup.7 iu
and 1.times.10.sup.8 iu).
[0687] The starting dose will be 1.times.10.sup.7 iu, administered
IV on 4 weekly occasions on days 1, 8, 15 and 22. The dose will
then escalate to 1.times.10.sup.8 iu and Seprehvir administered IV
on 4 weekly occasions on days 1, 8, 15 and 22. Two other dosing
regimen will be tested at 1.times.10.sup.8 iu. Patients will
receive either a single cycle of 4.times.IV Seprehvir on days 1, 5,
8 and 13 or two cycles of this dosing scheme one week apart.
[0688] The DLT assessment period will comprise the first 12 days
after last IV dose. Recruitment into each cohort will be
sequential, whereby the first patient to be treated must have
successfully completed the DLT assessment period without
experiencing a DLT, prior to the next patient being treated at that
dose level. The twelve-day dosing interval will be observed for all
subsequent patient(s). Initially three patients will be treated in
a given cohort. If any of these 3 patients experience a DLT during
their DLT assessment period, an additional 3 patients (total of
six) will be treated at that dose level. Following completion of
the DLT assessment period by the final patient in each cohort, all
available adverse event and laboratory safety data will be collated
and reviewed by the Principal Investigator and sponsor, and a
decision made regarding progression to the next dose level.
[0689] Patients who do not complete the DLT assessment period for
reasons other than toxicity will be replaced for the purpose of
toxicity evaluation.
[0690] Dose Limiting Toxicities
The definition of Seprehvir DLT will be made according to the
National Cancer Institute (NCI) Common Terminology Criteria for
Adverse Events [NCI CTCAE Version 4].
Haematological DLT:
[0691] Neutropenia<0.5.times.10.sup.9/L for >5 days [0692]
Neutropenia<1.times.10.sup.9/L with fever [0693]
Thrombocytopenia<25.times.10.sup.9/L accompanied by bleeding or
thrombocytopenia<10.times.10.sup.9/L
Non-Haematological DLT:
[0693] [0694] Any Grade 3 or 4 toxicity that is not related to
tumour progression with the exception of [0695] Grade 3 `flu-like
symptoms (including fever, chills and malaise) in the absence of
appropriate prophylaxis [0696] Grade 3 nausea, vomiting and
abdominal pain unless persisting for >2 days despite appropriate
prophylaxis [0697] Isolated laboratory abnormalities .gtoreq.Grade
3 that resolve to .ltoreq.Grade 1 in .ltoreq.7 days without
clinical sequelae or need for therapeutic intervention will not be
considered a DLT
[0698] If a patient develops an absolute neutrophil count (ANC)
<500/.mu.L or a platelet count <25,000/.mu.L, blood samples
must be collected every 2 to 3 days and study treatment withheld
until counts resolve or until ANC returns to >1000/.mu.L and
platelet counts return to >75,000/.mu.L.
Study Population
[0699] Inclusion criteria: [0700] 1. Patients with histologically
confirmed solid tumour who have exhausted all standard lines of
therapy for advanced or metastatic disease and/or for whom no
standard therapy exists [0701] 2. Previous treatment with
anticancer agent(s), including chemotherapy, immunotherapy,
biological or hormonal therapy (other than LHRH agonists), must be
completed .gtoreq.4 weeks (6 weeks for nitrosoureas or mitomycin C)
prior to administration of Seprehvir, and all associated toxicity
must be resolved to .ltoreq.grade 1 prior to administration of
Seprehvir [0702] 3. Previous radiation therapy must be completed
.gtoreq.14 days prior to administration of Seprehvir, and all
associated toxicity must be resolved to .ltoreq.Grade 1 prior to
administration of Seprehvir [0703] 4. Prior major surgery must be
completed within 4 weeks prior to Seprehvir administration [0704]
5. Age .gtoreq.18 years (at screening) [0705] 6. ECOG performance
status 0 or 1 at screening [0706] 7. Life expectancy >12 weeks
(at screening) as determined by the Principal
Investigator/Sub-Investigator [0707] 8. Ability to give written
informed consent as evidenced by signature on the patient consent
form, to communicate well with the investigator and to comply with
the expectations of the study [0708] 9. Male and female patients of
child-bearing potential must use an approved method of
contraception during the study and for 3 months after the last dose
of Seprehvir
Exclusion Criteria:
[0709] A patient will be excluded from the study if any of the
following apply: [0710] 1. Evidence of severe or uncontrolled
systemic disease, congestive cardiac failure >New York Heart
Association (NYHA) Class 2, myocardial infarction within 6 months,
or any medical or surgical condition that is deemed significant by
the Principal Investigator [0711] 2. Known hypersensitivity to any
Seprehvir excipients [0712] 3. Brain metastases that are associated
with a changing neurological deficit that has been documented to be
stable for <3 months, or for which systemic corticosteroids are
required [0713] 4. Laboratory values: [0714] a) ANC
.ltoreq.1500/.mu.L [0715] b) Platelet count .ltoreq.75,000/4 [0716]
c) Haemoglobin<9 g/dL [0717] d) Serum bilirubin
.gtoreq.1.5.times.upper limit of normal (ULN) unless Gilbert's
Disease (.gtoreq.2.times.ULN) is known to be the only underlying
hepatic disorder [0718] e) Aspartate aminotransferase (AST) and
alanine aminotransferase (ALT) .ltoreq.2.5.times.ULN (AST and ALT
.gtoreq.5.times.ULN for subjects with liver metastasis) [0719] f)
Creatinine clearance within the local laboratory normal range
[0720] g) >1+ proteinuria on consecutive testing at least 24
hours apart [0721] 5. Prior investigational agents for malignant or
non-malignant disease within 4 weeks or 5 half-lives (whichever is
shorter) prior to Day 1 [0722] 6. Previous treatment with viral
therapy of any kind within 8 weeks of entry to the study [0723] 7.
Active systemic bacterial or clinically proven infection with
hepatitis B (HBV) or C(HCV) or evidence of Human Immunodeficiency
Virus (HIV) infection [0724] 8. Pregnancy or lactation [0725] 9.
History of a second malignancy except those treated with curative
intent >3 years previously in the absence of relapse and basal
cell skin cancer or cervical cancer in situ
Treatment and Interventions
[0726] Patients will attend clinic study visits at screening and on
Days 1, 8, 15 and 22 for IV Seprehvir administration at the first
dose level of 1.times.10.sup.7 iu (FIG. 43). The dose will then be
escalated to 1.times.10.sup.8 iu and patients will attend clinic
study visits at screening and on Days 1, 8, 15 and 22 for IV
Seprehvir administration at this dose (FIG. 43). Two other dosing
regimen will be tested at 1.times.10.sup.8 iu. Patients will
receive either a single cycle of 4.times.IV Seprehvir on days 1, 5,
8 and 13 or two cycles of this dosing scheme one week apart (FIG.
43). Patients will then undergo biopsy or resection, 4-7 days after
the final dose.
Screening Period: Tumour biopsy within 14 days before the first
dose of Seprehvir (Day 1).
Treatment Cycle: Days 1 to 22, 1 to 13 or 1 to 32 (FIG. 53).
Duration and Frequency
[0727] Seprehvir will be administered on Day 1 of each 4 to 8 dose
cycle until development of severe toxicity or withdrawal of
consent.
Evaluation
[0728] Physical examination, vital signs, ECG, routine blood panel
(haematology, clinical chemistry coagulation), HSV immune
response(IgG/IgM), uUrine sample and buccal swab for assessment of
viral shedding taken at initiation of baseline assessment, followed
by days +1, +8, +15, +22, +36 if weekly injections, days +1, +5,
+8, +13, +26 if 2 injections/week or days +1, +5, +8, +13, +21,
+25, +28, +32 and +46 if 2.times.2 injections per week.
[0729] HSV bloods (IgG/IgM) and immune cell profiling (FACS) taken
at pre-screen, at time of biopsy/surgery, and at end of study
visit.
[0730] Additional viremia assessment (HSV-1 PCR blood) conducted at
3, 6 and 24 hrs post administration.
[0731] Seprehvir replication and immune cell recruitment (HSV-1 PCR
and IHC)--tumour tissue samples taken in pre-treatment biopsy and
post treatment surgery or biopsy.
[0732] Follow up: The End of Study Visit is to occur 14 days after
the subject has discontinued study treatment. All Seprehvir-related
toxicities will be followed until the End of Study Visit or until
all treatment-related toxicities have resolved to .ltoreq.Grade 2,
stabilized, or returned to baseline.
Example 7--Phase I Trial of HSV1716 in Patients with Non-Central
Nervous System (Non-CNS) Solid Tumors
[0733] Clinical trial NCT00931931 is an investigation into the use
of HSV1716 in patients with non-central nervous system (non-CNS)
solid tumors (typically sarcomas and neuroblastoma) and has a two
part study design. Part 1 of the study specifies a single dose of
virus. Participants who experience at least stable disease or
relapse following a determination of stable disease, may qualify
for subsequent doses in Part 2. There are two treatment arms: an
intratumoral route in which participants with localised disease
receive HSV1716 as an intratumoral injection; and an intravenous
route in which participants with metastatic disease receive HSV1716
intravenously.
[0734] FDA approval for systemic administration of Seprehvir in
clinical trial was supported by FDA-approved in vivo toxicology
& biodistribution studies for IV Seprehvir and extensive
preclinical efficacy studies in murine xenograft models.
[0735] Participants enrolled in the trial have now started to
receive HSV1716. 9 patients are enrolled in the intravenous arm.
Intravenous infusion has started at a conservative level with a
single dose of 2.times.10.sup.6 i.u. HSV1716. Initial results from
the first 4 patients demonstrate evidence that HSV1716 is reaching
tumor and is replicating therein.
[0736] PCR analysis of blood samples obtained from the patients at
several time points was used to identify the presence ("Pos") or
absence ("Neg") of HSV-1 DNA. Results are shown in FIGS. 58 and 61
and indicate no evidence of HSV1716 in the circulation immediately
following intravenous infusion (day 0 post infusion).
[0737] Blood samples were subjected to shell vial culture. All
cultures for viable HSV1716 were also negative at all time
points
[0738] However, by Day 4, it is notable that in 2/4 patients a
signal reappears in blood samples collected and analysed for HSV
DNA by PCR. This signal is consistent with an initial burst of
HSV1716 replication in tumor post administration and shedding of
HSV1716 DNA back into the circulation. In 1 of 4 patients, the
signal persisted to Day 14. This is encouraging given the treatment
involved a single dose at low titre. In 1 patient, the signal did
not materialise until day 28.
[0739] This data shows that herpes simplex virus administered to
the blood is immediately absorbed such that intact viral particles
are not detectable in the blood. Viral DNA is also not detectable
immediately following administration but is detectable several days
after administration. This supports the theory that HSV1716 is
quickly absorbed by cells or is neutralised following intravenous
administration, but is able to reach tumor tissue where it may
infect, replicate and lyse cells, lysis of tumor cells releasing
viral DNA which is detectable in the blood. Similar PCR results
have been seen at Day 4 following image-guided intratumoral
administration of HSV1716 in some patients in this study and the
similarity in pattern between the PCR bloods by both intratumoral
injection and IV infusion is significant.
[0740] Pharmacokinetic data from the first two patients treated
with intravenous Seprehvir indicates an initial loss of the input
signal during the first 24 hrs post IV infusion with subsequent
re-emergence of signal at day 4. The fourth IV patient had a
positive signal on day 28 (FIG. 61). Supportive of intratumoural
replication (intratumoral patients have also shown this pattern of
HSV emergence in the circulation).
[0741] To date, no virus has been detected in the circulation of
third IV patient.
Case Study--Patient HSV13.
[0742] This is the fourth patient to receive Seprehvir by
intravenous administration. The patient is a 25 year old Caucasian
male diagnosed with Ewing's Sarcoma (primary lesion in tibia,
metastatic lesion in lung) and enrolled on the study on Apr. 21,
2016. HSV13 received a single dose of 2.times.10.sup.6 pfu
Seprehvir by intravenous infusion.
[0743] PET/CT screening (FIGS. 62 and 63) revealed a low level of
standardized uptake value (SUV) at prescreen (no digital PET
available), day 14 post Seprehivir administration a flare up
(pseudo-progression) was noted with an increased SUV. At day 28
post Seprehivir administration a return to low level SUV was noted.
The effect of "pseudo-progression" shortly after treatment is
acknowledged for biologic agents.
[0744] A similar pattern has been seen in patient HSV06 (receiving
intratumoral Seprehvir).
Case Study--Patent HSV07
[0745] This patient received Seprehvir by intratumoral
administration. Patient is an 8 year old male with a diagnosis of
recurrent rhabdomyosarcoma (13 cm Stage III eRMS) in a
retroperitoneal location. Prior treatment includes surgery,
radiation (41.4 Gy tumor bed /36 Gy LN), chemotherapy (VAC per
D9803--remission; Relapse: VI, Cyclo/Topo, IE per ARST0121; PD:
Vinorelbine, oral cytox, temsirolimus; PD: Vinorelbine, oral cytox,
Avastin). Complications include AKI from obstructive uropathy and
ureteral stents, nephrostomies.
[0746] HSV07 showed interesting PET/CT findings (FIGS. 64 and 65).
A pre-injection PET on 28 May 2014 identified a site in tumour mass
for injection. PET scan day 14 post 1.sup.st injection indicated
stable disease at injection site--2.sup.nd injection administered.
PET hot spot at site distant from 1.sup.st injection on day 28. Hot
spot chosen as site for 2.sup.nd injection on 27th June 2014. PET
hot spot gone on 24 Jul. 2014.
Discussion
[0747] HSV is being detected in patients receiving a single
intravenous low dose (2.times.10.sup.6 pfu) of Seprehvir. This is a
low dose, similar to doses normally used in experiments with mice.
Experiments in mice typically use a dose of 1.times.10.sup.6 or
greater, meaning that following scale up for human administration
(considering human mass and blood volume) the expected dose
required would be about 1.times.10.sup.9 pfu or higher. Such a dose
would provide new challenges to (i) prove the safety of such a high
dose and (ii) manufacture sufficient quantities of virus. The
finding that an effect is present at a dose as low as
1.times.10.sup.6 pfu means that intravenous administration of doses
in the range 1.times.10.sup.7 to 1.times.10.sup.8 represents a
viable approach to treatment of tumors in human patients. Results
from our study of patients having mesothelioma (Example 1) are also
consistent with multiple systemic doses of Seprehvir leading to a
sustained Th1 response.
[0748] No HSV is being detected immediately following
administration of virus but, surprisingly, an HSV signal is
re-emerging in 3 out of 4 patients after at least several days. The
re-emergence of signal is consistent with results seen in patients
receiving intra-tumoral administration of Seprehvir by image guided
technology (see compare IV and ITu arms in FIG. 61).
[0749] The levels of HSV DNA detected by quantitative PCR are
approximately equivalent to the administered dose, which is a clear
indicator that virus is replicating. Our experiments on the
stability of Seprehvir in human blood (Examples 11 to 13) show that
Seprehvir has a short half-life in human blood. Our observation is
therefore consistent with sufficient virus reaching the tumor and
replicating therein.
[0750] Our intravenous infusion protocol has been well tolerated,
with no adverse reactions so far.
[0751] These observations are significant. Contrary to the
established view (e.g. see Russell et al., (Oncolytic virotherapy.
Nature Biotechnology Vol. 30 No. 7 Jul. 2012) and Seymour and
Fisher (British Journal of Cancer (2016) 114, 357-361)), the data
emerging from this trial indicates that Seprehvir can successfully
circumvent the innate obstacles presented by human blood and the
human immune system, can replicate and expand the viral population
to therapeutically effective levels and reach tumor tissue. This
opens the door to an alternative treatment of tumors that are
difficult to access by intratumoral injection.
Description of NCT00931931
[0752] Official title: A Phase I Dose Escalation Study of
Intratumoral or Intravenous Herpes Simplex Virus-1 Mutant HSV1716
in Patients with Refractory Non-Central Nervous System (Non-CNS)
Solid Tumors.
Purpose
[0753] Patients with relapsed solid tumors such as sarcomas and
neuroblastoma have a poor survival, generally <20%. There is an
urgent need for new treatments that are safe and effective.
[0754] HSV1716, an oncolytic virus, is a mutant herpes simplex
virus (HSV) type I, deleted in the RL1 gene which encodes the
protein ICP34.5, a specific determinant of virulence. Mutants
lacking the RL1 gene are capable of replication in actively
dividing cells but not in terminally differentiated cells--a
phenotype exploited to selectively kill tumor cells. In previous
clinical studies, HSV1716 has been shown to be safe when injected
at doses up to 10.sup.5 plaque forming units (pfu) directly into
human high-grade glioma and into normal brain adjacent to tumour,
following excision of high-grade glioma. In an extension study,
HSV1716 has been shown to be safe when injected at a dose of up to
10.sup.6 pfu directly into brain tumours.
[0755] Replication of HSV1716 in human glioblastoma in situ has
been demonstrated. Following a single administration of HSV1716 by
direct injection into active recurrent tumor or brain adjacent to
tumor, some patients have lived longer than might have been
expected. In part, this study seeks to evaluate the safety of a
single injection of HSV1716 in the treatment of extracranial solid
tumors in adolescents and young adults.
[0756] HSV1716 has also proved safe when given by direct
intra-tumoural injection in patients with squamous carcinoma of the
head and neck, and in patients with malignant melanoma.
[0757] Replication of HSV mutants in human sarcomas and
neuroblastoma in cultured cells and human xenograft models has been
demonstrated. This study is designed in two parts. PART 1 of the
study specifies a single dose of virus. Participants who experience
at least stable disease or relapse following a determination of
stable disease, may qualify for subsequent doses in PART 2. PART 2
requires signing of a separate consent.
Primary Outcome Measures:
[0758] To determine whether intratumoral injection or intravenous
infusions of HSV1716 is safe in adolescents and young adults with
non-CNS solid tumors.
Secondary Outcome Measures:
[0759] To measure antiviral immune response in patients with
refractory cancer treated with HSV1716.
Treatment Arms
[0760] Intratumoral route--participants with localized disease
receive HSV1716 as an intratumoral injection.
[0761] Intravenous route--participants with metastatic disease
receive HSV1716 intravenously.
Condition
[0762] Participants may have one of the following conditions:
Rhabdomyosarcoma, Osteosarcoma, Ewing Sarcoma, Soft Tissue Sarcoma,
Neuroblastoma, Wilms Tumor, Malignant Peripheral Nerve Sheath
Tumor, Clival Chordoma, Non-CNS Solid Tumors.
Eligibility
[0763] Ages Eligible for Study: 7 Years to 30 Years [0764] Genders
Eligible for Study: Both [0765] Accepts Healthy Volunteers: No
Inclusion Criteria:
[0766] Inclusion of Women and Minorities: The study is open to all
participants regardless of gender or ethnicity.
[0767] Inclusion for intratumoral injection: Subject must have 1-3
lesions amenable to HSV1716 administration by needle if
superficial; by needle and/or catheter if deep or pulmonary, via
interventional radiology without undue risk. Lesion(s) must meet
specific size criteria.
[0768] Inclusion for intravenous administration: Subject must have
metastatic disease or a lesion not deemed suitable for direct
injection.
[0769] Age: Subjects must be greater than or equal to 7 years and
less than or equal to 30 years of age at the time of signing
consent (study entry).
[0770] Histologic Diagnosis: Subjects must have had histologic
verification of a non-CNS solid tumor at original diagnosis. The
tumor must be amenable to HSV1716 administration without undue
risk. Disease must be considered refractory to conventional therapy
or for which no conventional therapy exists.
[0771] Metastatic Disease: Subjects who have metastasis to the
brain are eligible for the intratumoral arm of this study; however,
no metastatic sites within the brain will be considered for
injection. Subjects who have metastasis to the brain are eligible
for the intravenous arm of this study only if those metastases have
been treated and are no longer active.
[0772] Performance Level: Karnofsky greater than or equal to 50.
Subjects who are unable to walk because of paralysis, but who are
up in a wheelchair will be considered ambulatory for the purpose of
assessing the performance score.
[0773] Subjects must have fully recovered from the acute toxic
effects of all prior chemotherapy, immunotherapy, or radiotherapy
prior to entering this study;
[0774] Myelosuppressive chemotherapy: Must not have received within
28 days of entry onto this study (42 days if prior nitrosourea)
accompanied by hematopoietic recovery, or 14 days of stopping
non-myelosuppressive therapy as long as hematopoietic requirements
are met;
[0775] Biologic (anti-neoplastic agent): Must not have received
within 7 days of entry onto this study (21 days if prior VEGF-Trap
and at least 3 half lives after last dose of a monoclonal
antibody). For biologic agents that have known adverse events
occurring beyond 7 days after administration, this period must be
extended beyond the time during which adverse events are known to
occur;
[0776] No Radiation Therapy greater than or equal to 14 days for
local palliative XRT (small port): greater than or equal to 6
months must have elapsed if prior craniospinal XRT or if greater
than or equal to 50% radiation of pelvis; greater than or equal to
42 days must have elapsed if other substantial bone marrow
radiation;
[0777] Immunoablative or myeloablative Stem Cell Transplant (SCT):
greater than or equal to 6 months must have elapsed from prior
autologous transplant. Subjects must not have graft versus host
disease post autologous transplant;
[0778] Investigational agent: greater than or equal to 28 days must
have elapsed from treatment with a different phase I agent;
[0779] Subjects with seizure disorder may be enrolled if on
anticonvulsants and well controlled. At the time of enrollment,
specified CNS conditions must be less than or equal to Grade II
toxicity per CTCAE 3.0 criteria;
[0780] All subjects must have adequate blood counts defined as:
peripheral absolute neutrophil count (ANC) greater than or equal to
750/uL, Platelet count greater than or equal to 100,000/uL (may be
a post transfusion value), Hemoglobin greater than or equal to 9.0
gm/dL (may be a post transfusion value)
[0781] Adequate renal function defined as: Serum creatinine less
than or equal to 1.5.times.upper limit of normal (ULN) for age or
creatinine clearance or radioisotope GFR greater than or equal to
70 ml/min/1.73 m2;
[0782] Adequate liver function defined as: Total bilirubin less
than or equal to 2.0.times.ULN for age, and SGPT (ALT) less than or
equal to 2.5.times.ULN for age and albumin greater than or equal to
2 g/dL, GGT<2.5.times.ULN
[0783] Adequate cardiac function as defined by: Shortening fraction
>25% by echocardiogram or ejection fraction above the
institutional lower limit of normal by MUGA, No focal wall motion
abnormalities as determined by either of the above studies, EKG
without evidence of ischemia or significant arrythmia
[0784] Adequate coagulation as defined by: PT/INR and
PTT<1.5.times.ULN for age;
[0785] Infectious Disease: Documented evidence of negative tests
for the presence of Hepatitis B surface antigen, Hepatitis C
antibody, HIV1 and HIV2 antibodies within the three months
preceding study entry. Subjects who do not have such evidence must
undergo appropriate testing prior to virus administration;
Exclusion Criteria:
[0786] Stem cell transplant: No subjects who have received an
allogeneic hematopoietic stem cell transplant are eligible;
[0787] Pregnancy or Breast-Feeding: There is no available
information regarding human fetal or teratogenic toxicities.
Pregnant women are excluded and pregnancy tests must be obtained in
girls who are post-menarchal. Males or females of reproductive
potential may not participate unless they have agreed to use an
effective contraceptive method from the time of study entry to a
period of no less than four months post the final HSV1716
injection. For the same period of time, women who participate in
this study must agree not to breast feed;
[0788] Consent: Unable or unwilling to give voluntary informed
consent/assent; Leukemia: Subjects with leukemia are not eligible
for study participation;
[0789] Infection or any other severe systemic disease or medical or
surgical condition deemed significant by the principal
investigator;
[0790] Administration of any unlicensed or investigational agent
within 4 weeks of entry to the study;
[0791] Growth factor(s): No PEG-GCSF within 14 days of virus
injection (day 0);
[0792] Anti-HSV antivirals: Subjects whose physicians determine
that anti-HSV antiviral therapy (such as acyclovir, ganciclovir,
foscarnet, etc.) cannot be safely discontinued from 2 days prior to
the injection to 28 days following the injection should not be in
the study.
[0793] Subjects who have other conditions which in the opinion of
the investigator contra-indicate the receipt of HSV1716 or indicate
subject's inability to follow protocol requirements.
Example 8--a Phase Ib/2 Open-Label Evaluation of the Safety and
Efficacy of Intravenous Administration of Oncolytic Herpes Simplex
Virus HSV1716 and Pembrolizumab Compared to Pembrolizumab Alone and
HSV1716 Alone in Subjects with Stage III or Stage IV Head and Neck
Cancer
[0794] In the Phase Ib part of this study, the objective of the
study is to demonstrate tumor targeting of HSV1716 when
administered by intravenous administration in patients with any
operable head and neck cancer who are indicated to receive a tumor
resection. Each patient will receive up to 4 doses of HSV1716 by
intravenous infusion at two dose levels (1.times.10.sup.7 i.u. and
1.times.10.sup.8 i.u.). Each dose will be administered within 1 to
7 days of the previous dose. The final dose will be administered
within 1 to 14 days of the tumor resection. Patient tumor material
will be collected during the procedure and will be stored for
analysis to confirm evidence of HSV1716 localisation to tumor and
anti-tumor immunological or biological effect. Analysis will
involve shell vial culture, immunohistochemical analysis of tumor
tissue, qPCR to detect HSV DNA, and detection of immunological
response, e.g. infiltrating immune cells, cytokine response.
[0795] In addition, the safety and tolerability of the two dose
levels will be carefully monitored and compared during the period
up to tumor resection to confirm the maximum tolerated dose (MTD)
for the Phase II part of the Study. 3 patients will be recruited to
each dose level but in the event of a single Dose Limiting Toxicity
at any dose level, the cohort will be expanded to 6 patients
according to the usual "3+3" dose-escalation design.
[0796] In the Phase 2 part of this study, the objectives of this
study are to evaluate the following measures in an open-label,
multi-center, controlled study. Approximately 180 patients are to
be recruited and randomized 1:1:1 across each of the 3 treatment
arms: [0797] Arm 1: pembrolizumab alone; [0798] Arm 2: HSV1716
alone; [0799] Arm 3: HSV1716 and pembrolizumab.
[0800] Pembrolizumab (also known as MK-3475; lambrolizumab,
Keytruda.TM.; Merck, USA) is a humanised antibody that binds
PD-1.
Primary Outcome Measures:
[0801] Progression-free Survival (PFS) per immune related response
criteria ("irRC") for All Participants [0802] Overall Survival (OS)
for All Participants
Secondary Outcome Measures:
[0802] [0803] PFS per irRC in Participants with PD-L1-Positive
Expression [0804] OS in Participants with PD-L1-Positive Expression
[0805] Objective Response Rate (ORR) per irRC in All Participants
[0806] ORR per irRC in Participants with PD-L1-Positive Expression
[0807] Time to Tumor Progression (TTP) per irRC in All Participants
[0808] TTP per irRC in Participants with PD-L1-Positive Expression
[0809] Percentage of Participants Experiencing Grade 3-5 AEs [0810]
Time to First Grade 3-5 Adverse Event (AE) [0811] Percentage of
Participants Experiencing Viral Shedding of HSV1716 [0812]
Percentage of Participants Experiencing an Anti-viral immune
response to HSV1716
[0813] In Arm 1, pembrolizumab is administered intravenously at a
dose of 200 mg on day 1 of each 3 week cycle.
[0814] In Arms 2 and 3, HSV1716 is administered by intravenous
infusion at a dose of up to 1.times.10.sup.8 i.u. on each occasion
or at the dose of HSV1716 established as the MTD in the Phase 1b
part of the study. For intravenous infusion of HSV1716, vials of
HSV1716 will be diluted into 250 mL lactated Ringer's solution and
administered over one hour. Virus will be infused via peripheral IV
access. Standard hospital contact and respiratory precautions will
be followed, per institutional standards of operations for this
type of product. The dosing schedule for HSV1716 commences on day 1
and continues every week thereafter until up to 8 doses have been
administered (i.e. Days 1, 8, 15, 22, 29, 36, 43 and 50).
[0815] In Arm 3, pembrolizumab at a dose of 200 mg and HSV1716 at
up to 1.times.10.sup.8 i.u. are administered according to the
following schedule. The treatment may occur on the same day. Where
a delay in commencement of pembrolizumab is clinically justified, 1
cycle of HSV1716 may be given prior to commencement of
pembrolizumab.
TABLE-US-00001 Day Agent 1 8 15 22 29 36 43 50 HSV1716 + + + + + +
+ + Pembrolizumab + + +
[0816] In Arms 1 and 3, subjects shall continue dosing with
pembrolizumab therapy until a predetermined number of doses is
reached, dose limiting toxicity is observed or disease progression
is observed.
[0817] Times specified above are all subject to a tolerance of +1-3
days.
[0818] Results may be stratified by stage of disease, PD-L-1 status
of tumor, treatment cycles and anti-viral immune response.
[0819] Subjects are treated in each arm of the study until the
first to occur of: complete response; disease progression as per
the irRC; or intolerance of study treatment. For intravenous
infusion of HSV1716, vials of HSV1716 will be diluted into 250 mL
lactated Ringer's and administered over one hour. Virus will be
infused via peripheral IV access. Standard hospital contact and
respiratory precautions will be followed, per institutional
standards of operations for this type of product.
Example 9--Method for Selecting Patients for Treatment with a
Combination of HSV1716 and Pembrolizumab
[0820] Patients with head and neck cancer who are indicated for
surgery may receive up to 4 doses of HSV1716 by intravenous
infusion prior to surgical resection. Tumor tissue from the
resection may be analysed for evidence of HSV1716 targeting the
tumor and for an immunological or biological activity in response
to oncolytic immunotherapy. Patients demonstrating such activity
may be selected for cycles of HSV1716 therapy following surgery
with the aim of targeting residual tumor cells at the site of
surgical resection and/or metastatic disease.
Example 10--Preparation of a Vial of HSV1716 for Intravenous
Infusion
[0821] The total virus dose for each patient will be diluted into
250 mL lactated Ringer's and administered over one hour according
to the following instructions. Virus will be infused via peripheral
IV access. Standard hospital contact and respiratory precautions
will be followed, per institutional standards of operations for
this type of product.
[0822] Frozen vials of HSV1716 will be dispensed from the Pharmacy.
Preparation for intravenous administration will be performed within
an appropriate `clean` room. If transport to a `clean` room is
required, vials will be placed into a secondary container, labeled
appropriately and transported on dry ice. The label will include
"Route of administration--intravenous".
[0823] A 250 mL bag of lactated Ringer's solution for intravenous
infusion will also be dispensed from the Pharmacy and transported
as needed to the `clean` room in preparation for intravenous
administration. The lactated Ringer's solution to be maintained at
room temperature.
[0824] Defrost the vials of HSV1716 according to the manufacturer's
instructions. Once the vials are defrosted, they must be used
immediately.
[0825] Place the re-suspended vials and the bag containing 250 mL
of lactated Ringer's solution (IV bag) in a biosafety cabinet to
prepare the HSV1716 final drug product for intravenous
administration,
[0826] Aspirate 1 mL of the virus suspension from each vial into a
syringe ready for injection into the 250 mL bag of lactated
Ringer's via the inlet port. Gently mix the contents of the IV bag
using a backwards and forwards rocking motion.
[0827] Immediately following dilution of the investigational
product in the 250 ml of lactated Ringer's solution, label the IV
bag containing the HSV1716 final investigational drug product for
intravenous administration according to institutional policies and
applicable state and federal regulations. Immediately transfer to
the Principal Investigator or other staff as appropriate for
use.
[0828] Intravenous administration must be completed within a three
hour time period following preparation of the HSV1716 final drug
product.
[0829] Following the preparation of HSV1716 for intravenous use,
immediately place used vials on ice and return to the study
biosafety team for appropriate research purposes or
deactivation.
Example 11--HSV1716 and Human Primary Macrophages
[0830] 1) In an initial study human macrophages were infected with
HSV1716 at approximately 4 pfu/cell and the cells were then
incubated under normal and hypoxic conditions. Samples were removed
at various time points after infection (+1.5 hr, +24 hrs, +48 hrs
and +72 hrs) and titrated (FIG. 59).
[0831] Within 1 hour 90% of the virus had been adsorbed by the
macrophages and then no virus was detectable at 24 or 48 hrs in
either normoxia or hypoxia (detection limit of titration is 100
pfu/ml).
[0832] Significantly, virus was detectable at 72 hrs but the
amounts at this time were similar in the normoxic vs hypoxic
macrophages. This emergent virus is of significant interest as it
could either be the original input which had entered some transient
latent state or represent the first wave of replication in the
macrophages.
[0833] 2) Macrophages were infected with decreasing HSV1716 moi
(40, 4, 0.4 and 0.04) and samples were titrated after 72 hrs only.
Virus was detected from the macrophages infected at moi 40, 4 and
0.4 but not from those infected at 0.04 moi (FIG. 59).
Interestingly, the ratio of virus detected after 72 hrs relative to
the input pfu was approximately the same and similar to those from
the two other 72 hr normoxia/hypoxia time points shown in FIG.
60.
[0834] In summary, human primary macrophages were found to have a
high capacity to adsorb HSV1716, and active virus can be recovered
from the macrophages after 48 hrs in culture.
[0835] This data evidences that white blood cell components,
specifically monocyte derived cells, can rapidly absorb HSV1716 and
furthermore that HSV1716 is able to kill monocyte derived cells
after 96 hours from infection.
[0836] Accordingly, the inventors' have observed that following
infection of monocyte derived cells virus is not detected in the
infected cells but presence of virus is re-established upon
prolonged culture (FIG. 59). This is consistent with productive
infection of the cells, i.e. involving replication and cell lysis
by viral progeny, although the invention is not bound by such
theory. The finding that infection with oncolytic Herpes Simplex
Virus leads to cell death of monocyte derived cells means that
infected monocytes or monocyte derived cell may be used to deliver
the oncolytic Herpes Simplex Virus to the diseased tissue,
including to hypoxic areas of a tumor, subsequently allowing the
release of virus directly to the diseased tissue as the cell
dies.
[0837] The inventors also found that oncolytic Herpes Simplex Virus
replication in, and subsequent cell death (e.g. lysis) of, monocyte
derived cells is actually greater in hypoxic conditions. This
indicates that death of the monocyte derived cells occurs
(apparently preferentially) in hypoxic tumor environments and will
directly release the oncolytic Herpes Simplex Virus to the hypoxic
parts of a tumor that are otherwise difficult to access.
[0838] The data in FIGS. 58 and 59 support a hypothesis in which
intravenous administration of HSV1716 exposes the virus to a
population of macrophages or other monocyte derived cells that
transport the virus to hypoxic parts of the tumor.
Example 12--HSV1716 Infects but does not Replicate in Human
PBMC
[0839] Three human white blood cell lines were tested for HSV1716
infection and replication using HSV1716gfp, an HSV1716 variant
expressing gfp. SupT1 (ATCC, CRL-1942) is a T-cell lymphoma cell
line derived from the malignant cells collected from the malignant
pleural effusion of an 8 year old child with T-Cell Lymphoblastic
Lymphoma. THP-1 (ATCC, TIB-202) cells are monocytic cells derived
from a patient with acute monocytic leukaemia. Toledo (CRL-2631) is
a B-cell lymphoma cell line established from peripheral blood
leukocytes of a patient that originally had a diffuse large cell
lymphoma (DLCL). HSV1716gfp at moi 1 readily infected each of these
cell lines but failed to affect survival of the cells when assessed
by Trypan blue exclusion 5 days after virus infection (FIG. 66).
Wild-type HSV-1 17+ had some toxicity in SupT1 and THP-1 cells
compared to HSV1716.
[0840] Fresh human PBMC and monocytes were infected at moi 1 with
HSV1716gfp and green fluorescence was observed after 60 hrs of
infection using fluorescence microscopy. Light microscopy indicated
that only a subset of the PBMC were infected and there was no green
fluorescence in uninfected cells. Similar observations with
monocytes indicated that all of the monocytes were infected with
HSV1716gfp. Although the gfp fluorescence was much weaker in these
primary cells, all of the monocyte aggregates were positive for
green fluorescent and FACS analysis confirmed HSV1716 principally
infected the monocyte fraction of PBMC (FIG. 67).
[0841] HSV1716 is able to infect but not replicate in human
leukaemic cell lines. In the PBMC fraction of human blood, HSV1716
associates principally with monocytes. Human primary macrophages
have a high capacity to adsorb HSV1716 with no obvious
morphological signs of virus cytopathic effects at high moi. Live
virus was recovered from the macrophages after 48 hrs in
culture.
Example 13--HSV1716 Stability in Human Blood
[0842] Human blood is perceived as a very hostile environment to
oncolytic viruses and therefore there are apparent, major
limitations to intravenous delivery of oncolytic viruses. Hostile
factors to oncolytic virus persistence in blood include immune
attacks against the viruses mediated via complement, cytokines or
probably most critically, by neutralising antibodies in
seropositive patients. Therefore, most oncolytic virus studies to
date favour intratumoural or loco-regional delivery.
[0843] The stability of HSV1716 was assessed in human blood by
spiking samples of whole blood, plasma and cell fraction with a
known amount of HSV1716 and removing aliquots for titration at
various times after virus was added. An additional aliquot was
removed at each time point and added to Vero cells plated out in 60
mm dishes. The Vero cells were then incubated for 72 hrs and the
cells and medium were harvested, subjected to one freeze/thaw cycle
(-70.degree. C.) and titrated. This additional Vero cell culture
step was used to identify HSV1716 which was not available
immediately for infection, as in a titration assay, but is released
and becomes available to infect the Vero cells during the 72 hrs of
incubation. For example, HSV1716 may bind to or be taken up by
cells in the blood and released at later times or could be
transiently associated with plasma proteins that block
infection.
[0844] 5 ml each of whole blood, plasma and cell fraction were
obtained from a volunteer and were spiked with 100 ul clinical
grade HSV1716 (November fill, vial no 96, CRU-01A@ 2.times.10.sup.6
pfu/ml) which is equivalent to 200,000 pfu HSV1716 in each sample.
The expected titre in each of these samples is 4.times.10.sup.4
pfu/ml and, assuming that the average human blood volume is 5,000
ml, this equates to a systemic dose of 2.times.10.sup.8 pfu
HSV1716.
[0845] Additionally, 5 ml of PBS were spiked also with 200,000 pfu
HSV1716 and samples were titrated in parallel. The titre of this
spiked 5 ml of PBS spiked at the start of the experiment was
4.8.times.10.sup.4 pfu/ml which is within the acceptable range
(+/-0.5 log.sub.10) for the expected titre of 4.times.10.sup.4
pfu/ml.
[0846] The anti-HSV-1 IgG levels in the volunteer's plasma were
tested using a commercially available anti-HSV-1 IgG ELISA kit
(EUROIMMUN, D-23560 Lubeck, Seekamp 31). The kit was used exactly
as directed by the manufacturer and the anti-HSV-1 IgG levels are
determined in relative units/ml (RU/ml) with values derived from a
standard curve using samples of known anti-HSV-1 IgG levels
provided with the ELISA. The volunteer was seropositve for HSV-1
IgG with a high plasma concentration of anti-HSV-1 IgG of 600
RU/ml.
[0847] Two aliquots of 100 ul were removed from the spiked whole
blood, plasma, cell fraction and PBS at 2 mins, 5 mins, 10 mins, 20
mins, 60 mins, 120 mins, 180 mins and 240 mins after spiking with
HSV1716. The first 100 ul aliquot was titrated immediately for
half-life determination and the percentage virus remaining at each
sampling point was calculated using the titre of the PBS at the
start of the experiment (4.8.times.10.sup.4 pfu/ml) as 100%.
Results are presented graphically in FIG. 68 for samples collected
up to 10 mins after spiking. Titres of HSV1716 remain stable in PBS
during this time but decrease rapidly in whole blood, plasma and
the cell fraction during the first 5 minutes of incubation with
virus undetectable by titration within 10 mins. The estimated
half-lives for HSV1716 in whole blood, plasma or cell fractions are
approximately 2 mins 40 secs, 3 mins 10 secs or 3 mins 20 secs for
whole blood, plasma or cell fraction respectively. As it takes
approximately one minute for blood to complete a circuit in the
human circulation systems then at least two complete circuits will
be completed within the half-life of HSV1716. The other 100 ul
aliquot was added to Vero cells in a 60 mm plate and the plate
returned to the incubator and left for 72 hrs. HSV1716 always
became available to infect Vero cells from the 100 ul aliquot of
whole blood, plasma or cell fraction during the 72 hour incubation
period as indicated by titration of the harvested Vero cell extract
(FIG. 69). Titres of between 200,000 to 400,000 pfu/ml were
detected in the Vero cell extracts after the Vero cells had been
incubated with 100 ul spiked whole blood, plasma or cell fraction
indicating the availability of HSV1716 within these fractions to
infect cells. Titres obtained from Vero cells which received whole
blood or cell fraction tended to be higher than those from plasma
suggesting that more virus was available for release in these
fractions.
[0848] In parallel, Vero cells were infected with 10 or 100 pfu
HSV1716 diluted in PBS and these yielded 33,000 pfu (dotted line in
FIG. 69) or 420,000 pfu respectively indicating that between 10-100
pfu are released for infection of Vero cells from whole blood, cell
fraction or plasma during the 72 hrs incubation. It should be noted
that this probably represents an underestimation of the amount of
virus released from the 100 ul whole blood, cell fraction or plasma
added to the Vero cells as these may exert some neutralisation
effects on progeny virus propagated from the initial Vero cell
infections. The titre of the spiked whole blood, cell fraction and
plasma was approximately 40,000 pfu/ml HSV 1716 and, as the 100 ul
added to the Vero cells will contain 4000 pfu, then the 10-100 pfu
that infects the Vero cells during the 72 hrs incubation suggests
that between 0.25-2.5% of the input virus is available for
infection. This equates to 2.5.times.10.sup.5-2.5.times.10.sup.6
pfu from a 1.times.10.sup.8 pfu dose.
[0849] Results indicate that although HSV1716 has a short half life
in human blood, a small but significant proportion of the input
dose is not irreversibly neutralised and is continuously available
for infection. This proportion of the input HSV1716 may be bound to
or be taken up by cells in the blood and released at later times or
could be transiently associated with plasma proteins that block
infection.
* * * * *