U.S. patent application number 16/346310 was filed with the patent office on 2019-09-12 for combination therapy for treatment of brain cancers.
The applicant listed for this patent is DNATRIX, INC.. Invention is credited to Joanna J. Peterkin, Frank Tufaro.
Application Number | 20190275092 16/346310 |
Document ID | / |
Family ID | 62076437 |
Filed Date | 2019-09-12 |
![](/patent/app/20190275092/US20190275092A1-20190912-D00000.png)
![](/patent/app/20190275092/US20190275092A1-20190912-D00001.png)
![](/patent/app/20190275092/US20190275092A1-20190912-D00002.png)
![](/patent/app/20190275092/US20190275092A1-20190912-D00003.png)
![](/patent/app/20190275092/US20190275092A1-20190912-D00004.png)
![](/patent/app/20190275092/US20190275092A1-20190912-D00005.png)
![](/patent/app/20190275092/US20190275092A1-20190912-D00006.png)
![](/patent/app/20190275092/US20190275092A1-20190912-D00007.png)
United States Patent
Application |
20190275092 |
Kind Code |
A1 |
Tufaro; Frank ; et
al. |
September 12, 2019 |
COMBINATION THERAPY FOR TREATMENT OF BRAIN CANCERS
Abstract
Methods of treating a subject having a brain tumor comprising
(a) administering an oncolytic virus and (b) administering a
therapeutic antibody to said subject.
Inventors: |
Tufaro; Frank; (Rancho Santa
Fe, CA) ; Peterkin; Joanna J.; (Rancho Santa Fe,
CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
DNATRIX, INC. |
Houston |
TX |
US |
|
|
Family ID: |
62076437 |
Appl. No.: |
16/346310 |
Filed: |
November 1, 2017 |
PCT Filed: |
November 1, 2017 |
PCT NO: |
PCT/US2017/059611 |
371 Date: |
April 30, 2019 |
Related U.S. Patent Documents
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
|
|
62416075 |
Nov 1, 2016 |
|
|
|
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 9/0085 20130101;
A61K 9/0019 20130101; C12N 2710/10321 20130101; A61K 39/3955
20130101; A61K 2039/545 20130101; A61K 2039/505 20130101; A61P
35/00 20180101; A61K 45/06 20130101; C07K 16/2818 20130101; C12N
2710/10332 20130101; C07K 2317/24 20130101; A61K 35/761 20130101;
A61K 2039/54 20130101; A61K 35/761 20130101; A61K 2300/00 20130101;
A61K 39/3955 20130101; A61K 2300/00 20130101 |
International
Class: |
A61K 35/761 20060101
A61K035/761; C07K 16/28 20060101 C07K016/28; A61K 39/395 20060101
A61K039/395; A61K 9/00 20060101 A61K009/00; A61K 45/06 20060101
A61K045/06; A61P 35/00 20060101 A61P035/00 |
Claims
1. A method of treating a subject having brain tumor comprising:
(a) administering an oncolytic adenovirus to said subject; and (b)
administering an anti-PD-1 antibody to said subject.
2. The method of claim 1, wherein the oncolytic adenovirus is an
adenovirus serotype 5 strain, is selectively replication competent
in cells defective in the Rb/p16 tumor suppressor pathway, contains
a deletion of the 24 nucleotides encoding amino acids 122 to 129 of
the adenoviral E1A protein, and/or contains an integrin binding
RGD-4C motif.
3. The method of claims 1-2, wherein the oncolytic adenovirus is
DNX-2401.
4. The method of claims 1-3, wherein the anti-PD1 antibody is a
humanized antibody.
5. The method of claims 1-4, wherein the anti-PD1 antibody is
pembrolizumab.
6. The method of claims 1-5, wherein the oncolytic adenovirus is
delivered intratumorally.
7. The method of claim 6, wherein the oncolytic adenovirus is
delivered via cannula or needle.
8. The method of claims 1-7, wherein the oncolytic adenovirus is
delivered at 5.times.10.sup.8 viral particles per dose,
5.times.10.sup.9 viral particles per dose, or 5.times.10.sup.10
viral particles per dose.
9. The method of claims 1-8, wherein the anti-PD1 antibody is
delivered by intravenous infusion.
10. The method of claims 1-9, wherein the anti-PD1 antibody is
delivered at 200 mg per dose.
11. The method of claim 10, wherein the dose is administered over
30 minutes.
12. The method of claims 1-11, wherein a single dose of the
oncolytic adenovirus is provided prior to three consecutive doses
of the anti-PD1 antibody.
13. The method of claim 12, wherein the time between oncolytic
adenovirus administration and the first administration of the
anti-PD1 antibody is about 7-9 days.
14. The method of claims 12-13, wherein the time between sequential
administrations of the anti-PD1 antibody is about three weeks.
15. The method of claims 1-14, further comprising administering to
the subject one or more of a steroid, an anticonvulsant or an
antibody that inhibits vascular endothelial growth factor A.
16. The method of claim 12, wherein treating further comprises
additional administrations of the anti-PD1 antibody for up to a 105
weeks or 24 months from the date of oncolytic adenovirus
administration.
17. The method of claims 1-16, wherein said subject is evaluated
for one of more of overall survival, tumor response, clinical
benefit rate, Karnofsky performance status, neurologic status,
cytokine levels, lymphocyte levels, or a biomarker.
18. The method of claim 17, wherein said biomarker is PD-1 level or
PDL-1 level.
19. The method of claim 18, wherein tumor response is measured by
MRI.
20. The method of claims 1-19, further comprising
stereotactically-guided biopsy of the brain tumor.
21. The method of claims 1-20, wherein the brain tumor is
glioblastoma multiforme.
22. The method of claims 1-20, wherein the brain tumor is
gliosarcoma.
23. The method of claims 1-22, wherein the subject exhibits an
overall survival of at least 9 months, 12 months, 15 months, 18
months, 24 months, 36 months or 48 months, any interval
therebetween.
24. The method of claims 1-22, wherein the subject exhibits an
increase in overall survival, as compared to an untreated control
subject, of at least 6 months, 9 months, 12 months, 15 months, 18
months, 24 months, 36 months or 48 months, any interval
therebetween.
25. The method of claims 1-22, wherein the tumor exhibits reduced
growth, no growth, a 10% reduction in tumor mass, a 20% reduction
in tumor mass, a 30% reduction in tumor mass, a 40% reduction in
tumor mass, a 50% reduction in tumor mass, a 60% reduction in tumor
mass, a 70% reduction in tumor mass, an 80% reduction in tumor
mass, a 90% reduction in tumor mass, or a 100% reduction in tumor
mass following initiation of treatment.
26. The method of claims 1-22, wherein the Karnofsky performance
status improves or remains unchanged following step (b).
27. The method of claims 1-22, wherein there is a statistically
measurable clinical benefit.
28. The method of claims 1-27, wherein the subject exhibits
recurrent or progressive brain tumor following previous
treatment.
29. The method of claims 1-28, wherein the previous treatment was
chemotherapy, radiotherapy, or antibody therapy.
30. The method of claims 1-29, wherein the subject is a human.
31. A composition comprising: (a) an oncolytic adenovirus; and (b)
an anti-PD-1 antibody.
32. The composition of claim 31, wherein the oncolytic adenovirus
is an adenovirus serotype 5 strain, and/or wherein the oncolytic
adenovirus is selectively replication competent in cells defective
in the Rb/p16 tumor suppressor pathway, and/or wherein the
oncolytic adenovirus comprises a deletion of the 24 nucleotides
encoding amino acids 122 to 129 of the adenoviral E1A protein,
and/or wherein the oncolytic adenovirus comprises an integrin
binding RGD-4C motif.
33. The composition of claims 31-32, wherein the oncolytic
adenovirus is DNX-2401.
34. The composition of claims 31-33, wherein the anti-PD1 antibody
is a humanized antibody.
35. The composition of claims 31-34, wherein the anti-PD1 antibody
is pembrolizumab.
36. The composition of claims 31-35, wherein the composition is
formulated for intratumoral delivery.
37. The composition of claims 31-35, wherein the composition is
formulated for intravenous infusion.
38. The composition of claims 31-37, wherein the oncolytic
adenovirus is comprised in a unit dose of 5.times.10.sup.8 viral
particles per dose, 5.times.10.sup.9 viral particles per dose, or
5.times.10.sup.10 viral particles per dose.
39. The composition of claims 31-38, wherein the anti-PD1 antibody
is comprised in a unit dose of 200 mg.
40. The composition of claims 31-39, further comprising one or more
of a steroid, an anticonvulsant or an antibody that inhibits
vascular endothelial growth factor A.
41. A composition comprising an oncolytic adenovirus of adenovirus
serotype 5 strain, wherein the oncolytic adenovirus is selectively
replication competent in cells defective in the Rb/p16 tumor
suppressor pathway, wherein the oncolytic adenovirus comprises a
deletion of the 24 nucleotides encoding amino acids 122 to 129 of
the adenoviral E1A protein, and wherein the oncolytic adenovirus
comprises an integrin binding RGD-4C motif.
42. The composition of claim 41, wherein the oncolytic adenovirus
is DNX-2401.
43. The composition of claims 41-42, wherein the oncolytic
adenovirus is comprised in a unit dose of 5.times.10.sup.8 viral
particles per dose.
44. The composition of claims 41-42, wherein the oncolytic
adenovirus is comprised in a unit dose of 5.times.10.sup.9 viral
particles per dose.
45. The composition of claims 41-42, wherein the oncolytic
adenovirus is comprised in a unit dose of 5.times.10.sup.10 viral
particles per dose.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application claims the benefit under 35 U.S.C. .sctn.
119(e) of U.S. Provisional Patent Application No. 62/416,075 filed
Nov. 1, 2016, which application is incorporated herein by reference
in its entirety.
FIELD OF THE INVENTION
[0002] The present application is directed to the fields of
medicine, oncology, viral therapy and immune therapy. In
particular, it is directed to the use, in combination, of an
oncolytic adenovirus and an antibody directed to PD-1, in the
treatment of brain cancers, such as glioblastomas and
gliosarcomas.
BACKGROUND
Description of the Related Art
[0003] Cancers are a large family of diseases characterized by the
uncontrolled growth of cells in a body. It is one of the leading
causes of death, with an estimated 14.1 million new cases worldwide
per year (in 2012), and an estimated 1.6 million new cases (in
2016) in the US alone.
[0004] Numerous therapies have been developed to treat cancer,
including for example, surgical removal of the cancer, use of
chemotherapeutic drugs (i.e., use of drugs which kill cancer
cells), and use of radiation therapy. Nevertheless, it is estimated
that in 2016 cancer will kill an estimated 600,000 people in the US
alone.
[0005] One particularly difficult form of cancer to treat,
recurrent glioblasoma (GBM), only has a median survival time of 3
to 7 months. Although surgery for recurrent disease is performed in
some patients, it has yet to be established whether surgery alone
prolongs survival and/or improves the quality of life for patients
with recurrent disease. Furthermore, only two drugs, bevacizumab
(Avastin.RTM.) and carmustine (Gliadel.RTM.) have been approved for
recurrent glioblastoma, the latter as an adjunct to surgery. To
date, bevacizumab has not been shown to improve survival.
[0006] DNX-2401 is currently being investigated for patients with
recurrent GBM in several Phase I clinical studies as monotherapy or
with other agents. To date, DNX-2401 has been well tolerated with
no unexpected clinically significant related toxicity reported
across all studies. In the first-in-human Phase I dose-escalating
study, no dose-limiting toxicity (DLT) or clinically significant
adverse events (AEs) directly related to virus were observed at any
dose level. Overall, AEs were generally mild to moderate in
severity and considered unrelated to DNX-2401.
[0007] The first-in-human Phase I study evaluated intratumoral
administration alone (group A) and intratumoral administration
followed 14 days later by tumor resection and subsequent intramural
delivery (group B) of DNX-2401 throughout the resected tumor
cavity. When administered as a single intratumoral injection, 52%
of subjects (13 of 25) achieved clinical benefit [complete response
(CR)+partial response (PR)+stable disease (SD)] per Macdonald
criteria, including 3 durable CRs. The 1-year survival rate for
this group was 32% (n=8), regardless of DNX-2401 dose
concentration. Four of these 8 subjects (20% overall) survived at
least 3 years, including 4 subjects who all achieved a PR (n=1) or
CR (n=3). As of February 2016, 3 subjects remain alive 3.8, 4.1,
and 4.3 years post-treatment.
[0008] Early phase studies with DNX-2401 have demonstrated that the
drug has a favorable safety profile and strong tumor-killing
potential in patients with recurrent malignant glioma.
Additionally, it has been observed that DNX-2401: 1) replicates in
human tumors for a period of weeks to months, 2) elicits tumor
necrosis within weeks of injection, 3) leads to long-term tumor
destruction detectable by Magnetic Resonance Imaging (MRI) and 4)
triggers intratumoral immune cell infiltration. Accumulating
evidence shows a correlation between tumor-infiltrating lymphocytes
(TILs) in cancer tissue and prognosis in various
malignancies..sup.i,ii,iii,iv,v,vi,vii,viii,ix,x,xi,xiii,xiv
[0009] Pembrolizumab)(KEYTRUDA.RTM., a humanized monoclonal
antibody against the programmed death receptor-1 (PD-1) protein,
has been developed by Merck & Co. for the treatment of cancer.
Pembrolizumab is approved for treatment of melanoma in several
countries; in the United States (US) it is indicated for the
treatment of advanced, unresectable or metastatic malignant
melanoma in patients with disease progression after prior treatment
with ipilimumab and, for BRAF V600 mutation-positive patients, a
BRAF inhibitor, while in the European Union (EU) it is approved for
the treatment of advanced (unresectable or metastatic) melanoma in
adults. Pembrolizumab has also been granted approval in the US for
the treatment of patients with metastatic non-small cell lung
cancer (NSCLC) whose tumors express PD-L1 as determined by an
FDA-approved test and who have disease progression on or after
platinum-containing chemotherapy. Pembrolizumab has demonstrated
initial clinical efficacy in single arm studies as monotherapy for
patients with multiple tumor types as determined by response rate.
Ongoing clinical trials are being conducted in a number of other
advanced solid tumor indications including glioblastoma and
hematologic malignancies.
[0010] All of the subject matter discussed in the Background is not
necessarily prior art and should not be assumed to be prior art
merely as a result of its discussion in the Background section.
Along these lines, any recognition of problems in the prior art
discussed in the Background or associated with such subject matter
should not be treated as prior art unless expressly stated to be
prior art. Instead, the discussion of any subject matter in the
Background should be treated as part of the inventor's approach to
the particular problem, which in and of itself, may also be
inventive.
SUMMARY
[0011] In accordance with the present disclosure, there is provided
a method of treating a subject having brain tumor comprising (a)
administering an oncolytic adenovirus to said subject; and (b)
administering an anti-PD-1 antibody to said subject. The oncolytic
adenovirus may be an adenovirus serotype 5 strain, may be
selectively replication competent in cells defective in the Rb/p16
tumor suppressor pathway, may contain a deletion of the 24
nucleotides encoding amino acids 122 to 129 of the adenoviral ElA
protein, and/or may contain an integrin binding RGD-4C motif. The
oncolytic adenovirus may be DNX-2401. The anti-PD1 antibody may be
a humanized antibody, such as pembrolizumab. The brain tumor may be
glioblastoma multiforme or gliosarcoma. The subject may be a
human.
[0012] The oncolytic adenovirus is delivered intratumorally, such
as via cannula or needle. The oncolytic adenovirus may be delivered
at 5.times.10.sup.8 viral particles per dose, 5.times.10.sup.9
viral particles per dose, or 5.times.10.sup.10 viral particles per
dose. The anti-PD1 antibody may be delivered by intravenous
infusion. The anti-PD1 antibody may be delivered at 200 mg per
dose. The dose may administered over 30 minutes. A single dose of
the oncolytic adenovirus may be is provided prior to three
consecutive doses of the anti-PD1 antibody. The time between
oncolytic adenovirus administration and the first administration of
the anti-PD1 antibody may be about 7-9 days. The time between
sequential administrations of the anti-PD1 antibody may be about
three weeks.
[0013] The method may further comprise administering to the subject
one or more of a steroid, an anticonvulsant or an antibody that
inhibits vascular endothelial growth factor A. Treating may further
comprise additional administrations of the anti-PD1 antibody for up
to a 105 weeks or 24 months from the date of oncolytic adenovirus
administration. The subject may be evaluated for one of more of
overall survival, tumor response, clinical benefit rate, Karnofsky
performance status, neurologic status, cytokine levels, lymphocyte
levels, or a biomarker, such as PD-1 level or PDL-1 level. Tumor
response may be measured by MRI. The method may further comprise
stereotactically-guided biopsy of the brain tumor.
[0014] The subject may exhibit an overall survival of at least 9
months, 12 months, 15 months, 18 months, 24 months, 36 months or 48
months, any interval therebetween. The subject may exhibit an
increase in overall survival, as compared to an untreated control
subject, of at least 6 months, 9 months, 12 months, 15 months, 18
months, 24 months, 36 months or 48 months, any interval
therebetween. The tumor may exhibit reduced growth, no growth, a
10% reduction in tumor mass, a 20% reduction in tumor mass, a 30%
reduction in tumor mass, a 40% reduction in tumor mass, a 50%
reduction in tumor mass, a 60% reduction in tumor mass, a 70%
reduction in tumor mass, an 80% reduction in tumor mass, a 90%
reduction in tumor mass, or a 100% reduction in tumor mass
following initiation of treatment. The Karnofsky performance status
may measurably and/or statistically improve or remain unchanged
following step (b). The subject may exhibit recurrent or
progressive brain tumor following previous treatment, such as
chemotherapy, radiotherapy, or antibody therapy.
[0015] In another embodiment, there is provided a composition
comprising (a) an oncolytic adenovirus; and (b) an anti-PD-1
antibody. The oncolytic adenovirus may be an adenovirus serotype 5
strain, and/or may be selectively replication competent in cells
defective in the Rb/p16 tumor suppressor pathway, and/or may
comprise a deletion of the 24 nucleotides encoding amino acids 122
to 129 of the adenoviral E1A protein, and/or may comprise an
integrin binding RGD-4C motif. The oncolytic adenovirus may be
DNX-2401. The anti-PD1 antibody may be a humanized antibody, such
as pembrolizumab.
[0016] The composition may be formulated for intratumoral delivery,
or formulated for intravenous infusion. The composition may
comprise oncolytic adenovirus in a unit dose of 5.times.10.sup.8
viral particles per dose, 5.times.10.sup.9 viral particles per
dose, or 5.times.10.sup.10 viral particles per dose. The anti-PD1
antibody may be comprised in a unit dose of 200 mg. The composition
may further comprise one or more of a steroid, an anticonvulsant or
an antibody that inhibits vascular endothelial growth factor A.
[0017] In yet another embodiment, there is provided a composition
comprising an oncolytic adenovirus of adenovirus serotype 5 strain,
wherein the oncolytic adenovirus is selectively replication
competent in cells defective in the Rb/p16 tumor suppressor
pathway, wherein the oncolytic adenovirus comprises a deletion of
the 24 nucleotides encoding amino acids 122 to 129 of the
adenoviral E1A protein, and wherein the oncolytic adenovirus
comprises an integrin binding RGD-4C motif. The oncolytic
adenovirus may be DNX-2401. The oncolytic adenovirus may be
comprised in a unit dose of 5.times.10.sup.8 viral particles per
dose, a unit dose of 5.times.10.sup.9 viral particles per dose, or
a unit dose of 5.times.10.sup.10 viral particles per dose.
[0018] Embodiments discussed in the context of methods and/or
compositions of the disclosure may be employed with respect to any
other method or composition described herein. Thus, an embodiment
pertaining to one method or composition may be applied to other
methods and compositions of the disclosure as well.
[0019] As used herein the specification, "a" or "an" may mean one
or more. As used herein in the claim(s), when used in conjunction
with the word "comprising", the words "a" or "an" may mean one or
more than one.
[0020] The use of the term "or" in the claims is used to mean
"and/or" unless explicitly indicated to refer to alternatives only
or the alternatives are mutually exclusive, although the disclosure
supports a definition that refers to only alternatives and
"and/or." As used herein "another" may mean at least a second or
more.
[0021] Throughout this application, the term "about" is used to
indicate that a value includes the inherent variation of error for
the device, the method being employed to determine the value, or
the variation that exists among the study subjects.
[0022] Other objects, features and advantages of the present
disclosure will become apparent from the following detailed
description. It should be understood, however, that the detailed
description and the specific examples, while indicating preferred
embodiments of the disclosure, are given by way of illustration
only, since various changes and modifications within the spirit and
scope of the disclosure will become apparent to those skilled in
the art from this detailed description.
BRIEF DESCRIPTION OF THE DRAWINGS
[0023] The following drawings form part of the present
specification and are included to further demonstrate certain
aspects of the present disclosure. The disclosure may be better
understood by reference to one or more of these drawings in
combination with the detailed description of specific embodiments
presented herein.
[0024] FIG. 1. Overall Study Design.
[0025] FIG. 2. Study Drug Administration
[0026] FIG. 3. Dose Limiting Toxicity Study Design.
[0027] FIG. 4. Imaging and Treatment after First Radiologic
Evidence of Progressive Disease.
[0028] FIG. 5. Specific Laboratory Evaluations.
[0029] FIGS. 6A and 6B. Pembrolizumab Dose Modification Guidelines
for Drug-Related Adverse Events.
DETAILED DESCRIPTION OF THE INVENTION
[0030] As discussed above, the present invention provides
compositions and methods for treating a cancer (e.g., a brain
cancer such as glioblastoma), comprising the step of administering
to a subject an oncolytic adenovirus and an anti-PI1 antibody. As
is discussed in more detail above, although representative
embodiments of an oncolytic adenovirus (e.g., DNX-2401) and an
anti-PD1 antibody (e.g., pembrolizumab) have both shown efficacy in
distinct types of cancers, they have not been utilized in a
clinical setting in concert for a single type of cancer, e.g.,
brain cancer.
I. CANCER
[0031] "Cancer" as utilized herein refers to a large family of
diseases characterized by the uncontrolled growth of cells in a
body. Representative forms of cancer include carcinomas, sarcomas,
myelomas, leukemia's, lymphomas, and mixed types of the above.
[0032] Representative forms of cancer include carcinomas, sarcomas,
myelomas, leukemia's, lymphomas, and mixed types of the above.
Further examples include, but are not limited to bile duct cancer,
brain cancers such as glioblastomas, breast cancer, cervical
cancer, CNS tumors (such as a glioblastoma, astrocytoma,
medulloblastoma, craniopharyogioma, ependymoma, pinealoma,
hemangioblastoma, acoustic neuroma, oligodendroglioma, menangioma,
neuroblastoma and retinoblastomas), colorectal cancer, endometrial
cancer, hematopoietic cell cancers including leukemia's and
lymphomas, hepatocellular cancer, kidney cancer, laryngeal cancer,
lung cancer, melanoma, oral cancer, ovarian cancer, pancreatic
cancer, prostate cancer, squamous cell carcinoma, and thyroid
cancer. Cancers may be diffuse (e.g., leukemia's), comprise solid
tumors (e.g., sarcomas such as fibrosarcoma, myxosarcoma,
liposarcoma, chondrosarcoma and osteogenic sarcoma), or some
combination of these (e.g., a metastatic cancer having both solid
tumors and disseminated or diffuse cancer cells).
[0033] Within particularly preferred embodiments of the invention
the cancer is a brain tumor or intracranial neoplasm. All types of
brain tumors may produce symptoms that vary depending on the part
of the brain involved. These may include headaches, seizures,
problem with vision, vomiting, and mental changes. The headache is
classically worse in the morning and goes away with vomiting. More
specific problems may include difficulty in walking, speaking and
with sensation. As the disease progresses unconsciousness may
occur.
[0034] The cause of most brain tumors is unknown. Risk factors that
may occasionally be involved include a number of inherited
conditions known as neurofibromatosis as well as exposure to the
industrial chemical vinyl chloride, the Epstein-Barr virus, and
ionizing radiation. The most common types of primary tumors in
adults are: meningiomas (usually benign), and astrocytomas such as
glioblastomas. In children, the most common type is a malignant
medulloblastoma. Diagnosis is usually by medical examination along
with computed tomography or magnetic resonance imaging. This is
then often confirmed by a biopsy. Based on the findings, the tumors
are divided into different grades of severity.
[0035] Treatment may include some combination of surgery, radiation
therapy and chemotherapy. Anticonvulsant medication may be needed
if seizures occur. Dexamethasone and furosemide may be used to
decrease swelling around the tumor. Some tumors grow gradually,
requiring only monitoring and possibly needing no further
intervention. Treatments that use a person's immune system are
being studied. Outcome varies considerably depending on the type of
tumor and how far it has spread at diagnosis. Glioblastomas usually
have poor outcomes while meningiomas usually have good outcomes.
The average five-year survival rate for brain cancer in the United
States is 33%.
[0036] A. Glioblastoma
[0037] Glioblastoma multiforme (GBM), also known as glioblastoma
and grade IV astrocytoma, is the most common and most aggressive
cancer that begins within the brain. Signs and symptoms are
initially non-specific. They may include headaches, personality
changes, nausea, and symptoms similar to that of a stroke.
Worsening of symptoms is often rapid. This can progress to
unconsciousness.
[0038] The cause of most cases is unclear. Uncommon risk factors
include genetic disorders such as neurofibromatosis and Li Fraumeni
syndrome and previous radiation therapy. Glioblastomas represent
15% of brain tumors. They can either start from normal brain cells
or develop from an already existing low-grade astrocytoma. The
diagnosis is typically made by a combination of CT scan, MRI scan,
and tissue biopsy.
[0039] There is no clear way to prevent the disease. Typically
treatment involves surgery after which chemotherapy and radiation
therapy is used. The medication temozolomide is frequently used as
part of chemotherapy. High dose steroids may be used to help reduce
swelling and decrease symptoms. It is unclear if trying to remove
all or simply most of the cancer is better.
[0040] Despite maximum treatment, the cancer usually recurs. The
most common length of survival following diagnosis is 12 to 15
months with less than 3 to 5% of people surviving greater than five
years. Without treatment survival is typically 3 months. About 3
per 100,000 people develop the disease a year. It most often begins
around 64 years of age and occurs more commonly in males than
females. Immunotherapy is being studied in glioblastoma with
promising results.
[0041] Although common symptoms of the disease include seizure,
nausea and vomiting, headache, memory loss, and hemiparesis, the
single most prevalent symptom is a progressive memory, personality,
or neurological deficit due to temporal and frontal lobe
involvement. The kind of symptoms produced depends highly on the
location of the tumor, more so than on its pathological properties.
The tumor can start producing symptoms quickly, but occasionally is
an asymptomatic condition until it reaches an enormous size.
[0042] For unknown reasons, GBM occurs more commonly in males. Most
glioblastoma tumors appear to be sporadic, without any genetic
predisposition. No links have been found between glioblastoma and
smoking, consumption of cured meat, or electromagnetic fields.
Alcohol consumption may be a possible risk factor. Glioblastoma has
been associated with the viruses SV40, HHV-6, and cytomegalovirus.
There also appears to be a small link between ionizing radiation
and glioblastoma. Some also believe that there may be a link
between polyvinyl chloride (which is commonly used in construction)
and glioblastoma. A 2006 analysis links brain cancer to lead
exposure in the work-place. There is an association of brain tumor
incidence and malaria, suggesting that the anopheles mosquito, the
carrier of malaria, might transmit a virus or other agent that
could cause glioblastoma or that the immunosuppression associated
with malaria could enhance viral replication. Also HHV-6
reactivates in response to hypersensitivity reactions from drugs
and environmental chemicals. Other risk factors include being male
(slightly more common in men than women), being over 50 years old,
being Caucasian, Hispanic, or Asian, having a low-grade astrocytoma
(brain tumor), which often, given enough time, develops into a
higher-grade tumor, or having one of the following genetic
disorders is associated with an increased incidence of gliomas:
Neurofibromatosis, Tuberous sclerosis, Von Hippel-Lindau disease,
Li-Fraumeni syndrome, or Turcot syndrome.
[0043] Glioblastoma multiforme tumors are characterized by the
presence of small areas of necrotizing tissue that are surrounded
by anaplastic cells. This characteristic, as well as the presence
of hyperplastic blood vessels, differentiates the tumor from Grade
3 astrocytomas, which do not have these features.
[0044] GBMs usually form in the cerebral white matter, grow
quickly, and can become very large before producing symptoms. Less
than 10% form more slowly following degeneration of low-grade
astrocytoma or anaplastic astrocytoma. These are called secondary
GBMs and are more common in younger patients (mean age 45 versus 62
years). The tumor may extend into the meninges or ventricular wall,
leading to high protein content in the cerebrospinal fluid (CSF)
(>100 mg/dL), as well as an occasional pleocytosis of 10 to 100
cells, mostly lymphocytes. Malignant cells carried in the CSF may
spread (rarely) to the spinal cord or cause meningeal gliomatosis.
However, metastasis of GBM beyond the central nervous system is
extremely unusual. About 50% of GBMs occupy more than one lobe of a
hemisphere or are bilateral. Tumors of this type usually arise from
the cerebrum and may rarely exhibit the classic infiltration across
the corpus callosum, producing a butterfly (bilateral) glioma.
[0045] The tumor may take on a variety of appearances, depending on
the amount of hemorrhage, necrosis, or its age. A CT scan will
usually show an inhomogeneous mass with a hypodense center and a
variable ring of enhancement surrounded by edema. Mass effect from
the tumor and edema may compress the ventricles and cause
hydrocephalus.
[0046] Four subtypes of glioblastoma have been identified: (i)
classical, which includes ninety-seven percent of tumors in the
`classical` subtype carry extra copies of the epidermal growth
factor receptor (EGFR) gene, and most have higher than normal
expression of epidermal growth factor receptor (EGFR), whereas the
gene TP53, which is often mutated in glioblastoma, is rarely
mutated in this subtype; (ii) proneural, which often has high rates
of alterations in TP53, and in PDGFRA, the gene encoding a-type
platelet-derived growth factor receptor, and in IDH1, the gene
encoding isocitrate dehydrogenase-1; (iii) mesenchymal, which is
characterized by high rates of mutations or other alterations in
NF1, the gene encoding Neurofibromin 1 and fewer alterations in the
EGFR gene and less expression of EGFR than other types; and (iv)
neural, which was typified by the expression of neuron markers such
as NEFL, GABRA1, SYT1 and SLC12A5. Many other genetic alterations
have been described in glioblastoma, and the majority of them are
clustered in three pathways, the P53, RB, and the PI3K/AKT.
Glioblastomas have alterations in 64-87%, 68-78% and 88% of these
pathways, respectively.
[0047] Another important alteration is methylation of MGMT, a
"suicide" DNA repair enzyme. Methylation is described to impair DNA
transcription and therefore, expression of the MGMT enzyme. Since
an MGMT enzyme can only repair one DNA alkylation due to its
suicide repair mechanism, reverse capacity is low and methylation
of the MGMT gene promoter greatly affects DNA-repair capacity.
Indeed, MGMT methylation is associated with an improved response to
treatment with DNA-damaging chemotherapeutics, such as
temozolomide.
[0048] Cancer cells with stem cell-like properties have been found
in glioblastomas (this may be a cause of their resistance to
conventional treatments, and high recurrence rate). These so-called
glioblastoma stem-like cells reside in a niche around arterioles,
which protects these cells against therapy by maintaining a
relatively hypoxic environment. A biomarker for cells in
glioblastomas that exhibit cancer stem cell properties, the
transcription factor Hes3, has been shown to regulate their number
when placed in culture.
[0049] The IDH1 gene encodes for the enzyme isocitrate
dehydrogenase 1 and is frequently mutated in glioblastoma (primary
GBM: 5%, secondary GBM >80%). By producing very high
concentrations of the "oncometabolite" D-2-hydroxyglutarate and
dysregulating the function of the wild-type IDH1-enzyme it induces
profound changes to the metabolism of IDH1-mutated glioblastoma,
compared with IDH1 wild-type glioblastoma or healthy astrocytes.
Among others, it increases the glioblastoma cells' dependence on
glutamine or glutamate as an energy source. It has been
hypothesized that IDH1-mutated glioblastoma are in a very high
demand for glutamate and use this amino acid and neurotransmitter
as a chemotactic signal. Since healthy astrocytes excrete
glutamate, IDH1-mutated glioblastoma cells do not favor dense tumor
structures but instead migrate, invade and disperse into healthy
parts of the brain where glutamate concentrations are higher. This
may explain the invasive behaviour of these IDH1-mutated
glioblastoma.
[0050] Furthermore, glioblastoma multiforme exhibits numerous
alterations in genes that encode for ion channels, including
upregulation of gBK potassium channels and ClC-3 chloride channels.
It has been hypothesized that by upregulating these ion channels,
glioblastoma tumor cells can facilitate increased ion movement over
the cell membrane, thereby increasing H.sub.2O movement through
osmosis, which aids glioblastoma cells in changing cellular volume
very rapidly. This is helpful in their extremely aggressive
invasive behavior, because quick adaptations in cellular volume can
facilitate movement through the sinuous extracellular matrix of the
brain.
[0051] When viewed with MRI, glioblastomas often appear as
ring-enhancing lesions. The appearance is not specific, however, as
other lesions such as abscess, metastasis, tumefactive multiple
sclerosis, and other entities may have a similar appearance.
Definitive diagnosis of a suspected GBM on CT or MRI requires a
stereotactic biopsy or a craniotomy with tumor resection and
pathologic confirmation. Because the tumor grade is based upon the
most malignant portion of the tumor, biopsy or subtotal tumor
resection can result in undergrading of the lesion. Imaging of
tumor blood flow using perfusion MRI and measuring tumor metabolite
concentration with MR spectroscopy may add value to standard MRI in
select cases by showing increased relative cerebral blood volume
and increased choline peak respectively, but pathology remains the
gold standard for diagnosis and molecular characterization.
[0052] It is important to distinguish primary glioblastoma from
secondary glioblastoma. These tumors occur spontaneously (de novo)
or have progressed from a lower-grade glioma, respectively. Primary
glioblastomas have a worse prognosis, different tumor biology and
may have a different response to therapy, which makes this a
critical evaluation to determine patient prognosis and therapy.
Over 80% of secondary glioblastoma carries a mutation in IDH1,
whereas this mutation is rare in primary glioblastoma (5-10%).
Thus, IDH1 mutations are a useful tool to distinguish primary and
secondary glioblastomas since histopathologically they are very
similar and the distinction without molecular biomarkers is
unreliable.
[0053] It is very difficult to treat glioblastoma due to several
complicating factors. For example, .sup.the tumor cells are very
resistant to conventional therapies, the brain is susceptible to
damage due to conventional therapy, the brain has a very limited
capacity to repair itself, many drugs cannot cross the blood-brain
barrier to act on the tumor, and treatment of primary brain tumors
and brain metastases consists of both symptomatic and palliative
therapies.
[0054] Supportive treatment focuses on relieving symptoms and
improving the patient's neurologic function. The primary supportive
agents are anticonvulsants and corticosteroids.
[0055] Historically, around 90% of patients with glioblastoma
underwent anticonvulsant treatment, although it has been estimated
that only approximately 40% of patients required this treatment.
Recently, it has been recommended that neurosurgeons not administer
anticonvulsants prophylactically, and should wait until a seizure
occurs before prescribing this medication. Those receiving
phenytoin concurrent with radiation may have serious skin reactions
such as erythema multiforme and Stevens-Johnson syndrome.
[0056] Corticosteroids, usually dexamethasone given 4 to 8 mg every
4 to 6 h, can reduce peritumoral edema (through rearrangement of
the blood-brain barrier), diminishing mass effect and lowering
intracranial pressure, with a decrease in headache or
drowsiness.
[0057] Palliative treatment usually is conducted to improve quality
of life and to achieve a longer survival time. It includes surgery,
radiation therapy, and chemotherapy. A maximally feasible resection
with maximal tumor-free margins is usually performed along with
external beam radiation and chemotherapy. Gross total resection of
tumor is associated with a better prognosis.
[0058] Surgery is the first stage of treatment of glioblastoma. An
average GBM tumor contains 10.sup.11 cells, which is on average
reduced to 10.sup.9 cells after surgery (a reduction of 99%).
Benefits of surgery include resection for a pathological diagnosis,
alleviation of symptoms related to mass effect, and potentially
removing disease before secondary resistance to radiotherapy and
chemotherapy occurs.
[0059] The greater the extent of tumor removal, the better. Removal
of 98% or more of the tumor has been associated with a
significantly longer healthier time than if less than 98% of the
tumor is removed in retrospective analyses. The chances of
near-complete initial removal of the tumor may be increased if the
surgery is guided by a fluorescent dye known as 5-aminolevulinic
acid. GBM cells are widely infiltrative through the brain at
diagnosis, and so despite a "total resection" of all obvious tumor,
most people with GBM later develop recurrent tumors either near the
original site or at more distant locations within the brain. Other
modalities, typically radiation and chemotherapy, are used after
surgery in an effort to suppress and slow recurrent disease.
[0060] Subsequent to surgery, radiotherapy becomes the mainstay of
treatment for people with glioblastoma. It is typically performed
along with giving temozolomide (TMZ). A pivotal clinical trial
carried out in the early 1970s showed that among 303 GBM patients
randomized to radiation or nonradiation therapy, those who received
radiation had a median survival more than double those who did not.
Subsequent clinical research has attempted to build on the backbone
of surgery followed by radiation. On average, radiotherapy after
surgery can reduce the tumor size to 10.sup.7 cells. Whole-brain
radiotherapy does not improve when compared to the more precise and
targeted three-dimensional conformal radiotherapy. A total
radiation dose of 60-65 Gy has been found to be optimal for
treatment.
[0061] GBM tumors are well known to contain zones of tissue
exhibiting hypoxia which are highly resistant to radiotherapy.
Various approaches to chemotherapy radiosensitizers have been
pursued with limited success. As of 2010, newer research-approaches
included preclinical and clinical investigations into the use of an
oxygen diffusion-enhancing compound such as trans-sodium
crocetinate (TSC) as radiosensitizers, and as of 2015 a clinical
trial was underway.
[0062] Boron neutron capture therapy has been tested as an
alternative treatment for glioblastoma multiforme but is not in
common use.
[0063] Most studies show no benefit from the addition of
chemotherapy. However, a large clinical trial of 575 participants
randomized to standard radiation versus radiation plus temozolomide
chemotherapy showed that the group receiving temozolomide survived
a median of 14.6 months as opposed to 12.1 months for the group
receiving radiation alone. This treatment regime is now standard
for most cases of glioblastoma where the person is not enrolled in
a clinical trial. Temozolomide seems to work by sensitizing the
tumor cells to radiation.
[0064] High doses of temozolomide in high-grade gliomas yield low
toxicity, but the results are comparable to the standard doses.
Antiangiogenic therapy with medications such as bevacizumab control
symptoms but do not affect overall survival.
[0065] Alternating electric field therapy is an FDA-approved
therapy for newly diagnosed and recurrent glioblastoma. In 2015,
initial results from a phase-three randomized clinical trial of
alternating electric field therapy plus temozolomide in newly
diagnosed glioblastoma reported a three-month improvement in
progression-free survival, and a five-month improvement in overall
survival compared to temozolomide therapy alone, representing the
first large trial in a decade to show a survival improvement in
this setting. Despite these results, the efficacy of this approach
remains controversial among medical experts.
[0066] The median survival time from the time of diagnosis without
any treatment is 3 months, but with treatment survival of 1-2 years
is common. Increasing age (>60 years of age) carries a worse
prognostic risk. Death is usually due to widespread tumor
infiltration with cerebral edema and increased intracranial
pressure.
[0067] A good initial Karnofsky Performance Score (KPS) and MGMT
methylation are associated with longer survival. A DNA test can be
conducted on glioblastomas to determine whether or not the promoter
of the MGMT gene is methylated. Patients with a methylated MGMT
promoter have longer survival than those with an unmethylated MGMT
promoter, due in part to increased sensitivity to temozolomide.
This DNA characteristic is intrinsic to the patient and currently
cannot be altered externally. Another positive prognostic marker
for glioblastoma patients is mutation of the IDH1 gene, which can
be tested by DNA-based methods or by immunohistochemistry using an
antibody against the most common mutation, namely IDH1-R132H.
[0068] More prognostic power can be obtained by combining the
mutational status of IDH1 and the methylation status of MGMT into a
two-gene predictor. Patients with both IDH1 mutations and MGMT
methylation have the longest survival, patients with an IDH1
mutation or MGMT methylation an intermediate survival and patients
without either genetic event have the shortest survival.
[0069] Long-term benefits have also been associated with those
patients who receive surgery, radiotherapy, and temozolomide
chemotherapy. However, much remains unknown about why some patients
survive longer with glioblastoma. Age of under 50 is linked to
longer survival in glioblastoma multiforme, as is 98%+resection and
use of temozolomide chemotherapy and better Karnofsky performance
scores. A recent study confirms that younger age is associated with
a much better prognosis, with a small fraction of patients under 40
years of age achieving a population-based cure. The
population-based cure is thought to occur when a population's risk
of death returns to that of the normal population, and in GBM, this
is thought to occur after 10 years.
[0070] B. Gliosarcoma
[0071] Gliosarcoma is a rare type of glioma, a cancer of the brain
that comes from glial, or supportive, brain cells, as opposed to
the neural brain cells. Gliosarcoma is a malignant cancer, and is
defined as a glioblastoma consisting of gliomatous and sarcomatous
components.
[0072] It is estimated that approximately 2.1% of all glioblastomas
are gliosarcomas. Although most gliomas rarely show metastases
outside the cerebrum, gliosarcomas have a propensity to do so, most
commonly spreading through the blood to the lungs, and also liver
and lymph nodes.
[0073] Knowledge about this entity is limited to small
retrospective case series and case reports. In general, the
epidemiology and natural history of GSM appears similar to
glioblastoma (GBM). No patient or treatment factors have been
unequivocally identified that distinguish outcomes of GSM from GBM.
Gliosarcomas have an epidemiology similar to that of glioblastomas,
with the average age of onset being 54 years, and males being
affected twice as often as females. They are most commonly present
in the temporal lobe.
[0074] Due to small patient numbers, available case series are not
sufficiently powered to precisely characterize GSM. Modest, yet
clinically meaningful, differences between GSM and GBM may surface
with examination of a larger series. To refine our understanding of
GSM, we used the Surveillance, Epidemiology, and End Results (SEER)
database to identify and analyze more than 300 adult GSM patients
and compare them to adult GBM patients.
II. ACTIVE AGENTS
[0075] A. Oncolytic Adenoviruses
[0076] "Adenovirus" (Ad) refers to a large (approximately 36 kb)
DNA virus that infects humans, but which also display a broad host
range. Physically, adenovirus is an icosahedral virus containing a
double-stranded, linear DNA genome. There are approximately 50
serotypes of human adenoviruses, which are divided into six
families based on molecular, immunological, and functional
criteria. By adulthood, virtually every human has been infected
with the more common adenovirus serotypes, the major effect being
cold-like symptoms.
[0077] Adenoviral infection of host cells results in adenoviral DNA
being maintained episomally, which reduces the potential
genotoxicity associated with integrating vectors. In addition,
adenoviruses are structurally stable, and no genome rearrangement
has been detected after extensive amplification. Adenovirus can
infect most epithelial cells regardless of their cell cycle stage.
So far, adenoviral infection appears to be linked only to mild
disease such as acute respiratory disease in humans.
[0078] Members of any of the 57 human adenovirus serotypes (HAdV-1
to 57) may incorporate heterologous nucleic acid encoding an immune
cell stimulatory receptor agonist according to the invention. Human
Ad5 is well characterized genetically and biochemically (GenBank
M73260; AC 000008). Thus, in a preferred embodiment, the oncolytic
adenovirus is a replication competent Ad5 serotype or a hybrid
serotype comprising an Ad5 component. The adenovirus may be a wild
type strain but is preferably genetically modified to enhance tumor
selectivity, for example by attenuating the ability of the virus to
replicate within normal quiescent cells without affecting the
ability of the virus to replicate in tumor cells. Non-limiting
examples of replication competent oncolytic adenoviruses
encompassed by the present invention include Delta-24,
Delta-24-RGD, ICOVIR-5, ICOVIR-7, ONYX-015, ColoAd1, H101 and
AD5/3-D24-GMCSF. Onyx-015 is a hybrid of virus serotype Ad2 and Ad5
with deletions in the E1B-55K and E3B regions to enhance cancer
selectivity. H101 is a modified version of Onyx-015. ICOVIR-5 and
ICOVIR-7 comprise an Rb-binding site deletion of E1A and a
replacement of the E1A promoter by an E2F promoter. ColoAd1 is a
chimeric Add11p/Ad3 serotype. AD5/3-D24-GMCSF (CGTG-102) is a
serotype 5/3 capsid-modified adenovirus encoding GM-CSF (the Ad5
capsid protein knob is replaced with a knob domain from serotype
3).
[0079] Oncolytic adenoviruses injected into a tumor induce cell
death and release of new adenovirus progeny that, by infecting the
neighbor cells, generates a treatment wave that, if not halted, may
lead to the total destruction of the tumor. Significant antitumor
effects of Delta-24 have been shown in cell culture systems and in
malignant glioma xenograft models. Delta-24-RGD has shown
surprising anti-tumor effects in a Phase I clinical trial and is
currently the subject of additional clinical trials. Although lysis
of tumor cells is the main anti-cancer mechanism proposed for
Delta-24-RGD oncolytic adenovirus, data from the Phase I clinical
trial in patients with recurrent glioma and other observations
indicate that the direct oncolytic effect may be enhanced by the
adenovirus-mediated trigger of anti-tumor immune response.
[0080] The infectious cycle of the adenovirus takes place in 2
steps: the early phase which precedes initiation of the replication
of the adenoviral genome, and which permits production of the
regulatory proteins and proteins involved in the replication and
transcription of the viral DNA, and the late phase which leads to
the synthesis of the structural proteins. The early genes are
distributed in 4 regions that are dispersed in the adenoviral
genome, designated E1 to E4 ("E" denotes "early"). The early
regions comprise at least-six transcription units, each of which
possesses its own promoter. The expression of the early genes is
itself regulated, some genes being expressed before others. Three
regions, E1, E2, and E4 are essential to replication of the virus.
Thus, if an adenovirus is defective for one of these functions this
protein will have to be supplied in trans, or the virus cannot
replicate.
[0081] The E1 early region is located at the 5' end of the
adenoviral genome, and contains 2 viral transcription units, E1A
and E1B. This region encodes proteins that participate very early
in the viral cycle and are essential to the expression of almost
all the other genes of the adenovirus. In particular, the E1A
transcription unit codes for a protein that transactivates the
transcription of the other viral genes, inducing transcription from
the promoters of the E1B, E2A, E2B, E3, and E4 regions and the late
genes.
[0082] Within one embodiment of the invention, one or more
heterologous sequences can be incorporated into a nonessential
region of the adenovirus. Within a particularly preferred
embodiment of the invention one or more heterologous sequences can
be integrated in place of all or part of the E3 region.
Representative examples include cytokines, chemokines and
checkpoint inhibitors. Within one embodiment the heterologous
sequence code for an OX40 agonist (e.g., OX40L), GITR, anti-PD-1,
and/or anti-CTLA-3. Within another embodiment the heterologous
nucleic acid sequence encodes an inhibitor of an immune checkpoint
protein selected from the group consisting of CTLA4, PID-I, PD-LI,
PD-L2, B7-H3, B7-H4, TIM3, GALS, LAG3, VISTA, KIR, and/or BTLA.
Within yet another embodiment the heterologous nucleic acid
sequence encodes an agonist of an immune co-stimulatory receptor
selected from the group consisting of CD28, OX40 (CD 134),
glucocorticoid-induced TNF-receptor (GITR), CD137 (4-1BB), herpes
virus entry mediator A (HVEM), inducible T-cell costimulator (Icos
or cD278), cD27, Cl)40, and/or CD226. Representative examples are
disclosed in more detail in PCT/US2014/066920 and U.S. Provisional
No. 62/342,482, filed May 27, 2016, both of which are incorporated
by reference in their entirety.
[0083] The adenovirus enters the permissive host cell via a cell
surface receptor, and it is then internalized. The viral DNA
associated with certain viral proteins needed for the first steps
of the replication cycle enters the nucleus of the infected cells,
where transcription is initiated. Replication of the adenoviral DNA
takes place in the nucleus of the infected cells and does not
require cell replication. New viral particles or virions are
assembled after which they are released from the infected cells,
and can infect other permissive cells.
[0084] The adenovirus is an attractive delivery system. Embodiments
of the invention can utilize manufacturing process with maximum
yields of 1.times.10.sup.5 viral particles per cell. The process
can be free of or essentially free of protein, serum, and animal
derived components making it suitable for a broad range of both
prophylactic and therapeutic vaccine products.
[0085] Several factors favor the use of oncolytic adenoviruses for
the treatment of brain tumors. First, gliomas are typically
localized, and therefore an efficient local approach should be
sufficient to cure the disease without any detrimental effects on
other parts of the body. Second, gliomas harbor several populations
of cells expressing different genetic abnormalities. Thus, the
spectrum of tumors sensitive to the transfer of a single
therapeutic gene to cancer cells may be limited. Third, replication
competent adenoviruses can infect and destroy cancer cells that are
arrested in G.sub.0. Since gliomas invariably include non-cycling
cells, this property is important. Finally, the p16-Rb pathway is
abnormal in the majority of gliomas, thus making Delta-24
adenovirus particularly effective for treating these tumors,
although the loss of the retinoblastoma tumor suppressor gene
function has been associated with the causes of various types of
tumors and is not limited to treatment of gliomas. The oncolytic
adenoviruses of the instant invention are replication-competent in
cells with a mutant Rb pathways.
[0086] If an adenovirus has been mutated so that it is
conditionally replicative (replication-competent under certain
conditions), a helper cell may be required for viral replication.
When required, helper cell lines may be derived from human cells
such as human embryonic kidney cells, muscle cells, hematopoietic
cells or other human embryonic mesenchymal or epithelial cells.
Alternatively, the helper cells may be derived from the cells of
other mammalian species that are permissive for human adenovirus.
Such cells include, for example Vero cells or other monkey
embryonic mesenchymal or epithelial cells. In certain aspects a
helper cell line is 293. Various methods of culturing host and
helper cells may be found in the art, for example (Racher, A. J.,
Fooks, A. R. & Griffiths, J. B. Biotechnol Tech (1995) 9:
169.)
[0087] Adenoviruses can be isolated using different methodologies.
Most often, after transfection of the Ad genome, adenoviral plaques
are isolated from the agarose overlaid cells and the viral
particles are expanded for analysis. For detailed protocols the
skilled artisan is referred to (Graham, F. L., and Prevec, L.
(1991). Manipulation of adenovirus vectors. Methods Mol Biol 7,
109-128.).
[0088] Alternative technologies for the generation of adenovirus
vectors include utilization of the bacterial artificial chromosome
(BAC) system, in vivo bacterial recombination in a recA+bacterial
strain utilizing two plasmids containing complementary adenoviral
sequences, and the yeast artificial chromosome (YAC) system (PCT
publications 95/27071 and 96/33280, which are incorporated herein
by reference).
[0089] Representative examples of adenovirus vectors suitable for
use within the present invention include those described in US
Publication Nos. 2009/0175830, 2014/0377221, 2014/0377294,
2015/0306160, 2016/0289645, and 2016/0143967, and U.S. Pat. Nos.
6,210,946, 6,284,742, 6,312,699, 6,555,368, 6,649,396, 6,815,200,
6,824,771, 6,841,540, 6,955,808, 7,045,348, 7,297,542, 8,168,168
and 9,061,055, all of which are incorporated by reference in their
entirety.
III. DNX-2401
[0090] In one particularly preferred embodiment, the replication
competent oncolytic adenovirus is Delta-24 or Delta-24-RGD.
Delta-24 is described in U.S. Patent Application Publication Nos.
20030138405, and 20060147420, each of which are incorporated herein
by reference (see also "'"). The Delta-24 adenovirus is derived
from adenovirus type 5 (Ad-5) and contains a 24-base-pair deletion
within the CR2 portion of the E1A gene that encompasses the area
responsible for binding Rb protein (nucleotides 923-946)
corresponding to amino acids 122 to 129 in the encoded E1A protein
(Fueyo J et al., Oncogene, 19:2-12 (2000)). Delta-24-RGD further
comprises an insertion of the RGD-4C sequence (which binds strongly
to av133 and av135 integrins) into the HI loop of the fiber knob
protein (Pasqualini R. et al., Nat Biotechnol, 15:542-546 (1997)).
The E1A deletion increases the selectivity of the virus for cancer
cells; the RGD-4C sequence increases the infectivity of the virus
for gliomas and for several other tumors which express low level of
adenovirus receptors.
[0091] A. Therapeutic Uses
[0092] A stereotactic or image-guided injection device guides the
delivery of DNX-2401 into the tumor through a small hole in the
skull. The virus establishes an active infection by replicating in
and killing tumor cells with high potency and specificity. Results
from Phase I clinical studies indicate that DNX-2401: (i) can
replicate in human tumors for a period of weeks to months, (ii) can
elicit tumor necrosis within weeks of injection, (iii) can trigger
intratumoral immune cell infiltration, and (iv) can lead to long
term tumor destruction detectable by MRI.
[0093] DNX-2401 has been well tolerated. To date, it is associated
with limited toxicity as noted during several completed or ongoing
Phase I clinical studies investigating DNX-2401 as a single agent
or together with other agents for recurrent disease. The maximum
dose evaluated in a first-in-human Phase I trial for subjects with
recurrent glioma was 3e10 vp without any dose-limiting toxicity
reported (Study ID01-310). Clinical benefit (CR+PR+SD) was observed
in 52% (n=13) of the subjects enrolled (n=25) in the intratumoral
injection group (group A) of the study, based on
Investigator-assessed MRI. Overall, 28% of subjects achieved a CR
(n=3) or PR (n=4). The subject that achieved a complete response at
the lowest dose level for any responding subject (1e8 vp) remains
alive 4.2 years following DNX-2401 treatment. In a subset of
subjects, pseudo-progression noted on MRI was observed prior to
tumor regression, similar to objective response kinetics observed
with several other immunotherapies. The 1-year survival rate for
this group was 32% (n=8), irrespective of DNX-2401 dose.
Encouragingly, four of these 8 subjects (20% overall) survived at
least 3 years, including 4 subjects who achieved a PR (n=1) or CR
(n=3). As of February 2016, 3 subjects remain alive 3.8, 4.1, and
4.3 years post-treatment. (Refer to the current Investigator's
Brochure for further updates)
[0094] DNX-2401 contains two stable genetic changes in the
adenovirus dsDNA genome that cause it to (1) replicate selectively
in Rb-pathway deficient cells and (2) infect cells that express
RGD-binding integrins more efficiently. A deletion of 24 bases in
the E1A gene and insertion of an integrin-binding motif (RGD) in
the H1 loop of the fiber accomplished this. Because virtually all
tumor cells, including GBM, harbor defects in the Rb pathway or are
already in the cell cycle, DNX-2401 replicates in and kills these
tumor cells selectively and efficiently. DNX-2401 is superior to
wild type adenovirus with respect to replication in rapidly growing
tumor cells, an unprecedented property for an oncolytic
virus..sup.xvii,sviii,xxiv
[0095] B. Clinical Experience in Glioma
[0096] Study ID01-310. DNX-2401 was evaluated in a first in human,
two-group, dose-escalating Phase I study (Study ID01-310) in 37
subjects with recurrent high-grade malignant glioma at a single
center; The University of Texas MD Anderson Cancer Center. Group A
investigated a single direct intratumoral injection of DNX-2401
into recurrent tumor without further resection (n=25). Group B
investigated a biological endpoint that was to demonstrate
intratumoral virus replication and included only subjects with
resectable tumors (n=12). These subjects received two injections;
an initial intratumoral injection into biopsy-confirmed glioma on
Day 0, followed 14 days later by tumor resection and subsequent
injection of a divided dose of DNX-2401 into the post-resection
tumor bed.
[0097] Safety.
[0098] No dose-limiting toxicity (DLT) was reported during clinical
study ID01-310 at any dose level, including the highest dose
evaluated (3e10 vp), and there have been no clinically significant
unexpected DNX-2401-related safety concerns observed to date in
ongoing Phase I studies with 3e10 vp. Therefore, a maximum
tolerated dose (MTD) was not reached prior to study completion.
Adverse events were generally mild to moderate in severity, and
unrelated to virus following both types of administration (i.e.,
intratumoral and intramural). Analysis of subject sera, saliva,
nasopharyngeal secretions and urine did not demonstrate any
significant virus shedding. Only one serious adverse event report
of Grade 1 headache/head pain, Grade 1 confusion and Grade 2 fever
was considered possibly related to virus for one subject. All
deaths were considered unrelated to DNX-2401.
[0099] Efficacy.
[0100] Although study ID01-310 was a dose-escalation study designed
to evaluate a dose concentration of DNX-2401 that spanned four
orders of magnitude, all subjects were included in the secondary
efficacy analysis.
[0101] In group A (n=25, intratumoral injection), subjects were
enrolled across 8 dose cohorts (1e7 to 3e10 vp). Clinical benefit
(as measured by CR+PR+SD) was seen in 52% of subjects (n=13),
including 28% who achieved a best response of PR (n=4) or CR (n=3).
The subject that achieved a complete response at the lowest dose
level for any responding subject (1e8 vp) remains alive 4.2 years
following DNX-2401 treatment. In a subset of subjects,
pseudo-progression noted on MRI was observed prior to tumor
regression, similar to objective response kinetics observed with
several other immunotherapies..sup.xxi The 1-year survival rate for
this group was 32% (n=8), irrespective of DNX-2401 dose. Four of
these 8 subjects (20% overall) survived at least 3 years, including
4 subjects that all achieved a PR (n=1) or CR (n=3). As of February
2016, 3 subjects remain alive 3.8, 4.1, and 4.3 years
post-treatment.
[0102] In group B (n=12, intratumoral (within the tumor) and
intramural (tumor bed) injection; biological endpoint arm),
subjects were enrolled across 4 dose cohorts (1e7 to 3e8 vp; total
exposure after two injection: 2e7 to 6e8 vp). Subjects (n=12)
completed treatment successfully. Because tumor resection was part
of the treatment plan for group B, only a limited number of
subjects (25%, 3 of 12) had measurable disease 14 days following
intratumoral injection, which would have been essential to evaluate
changes in tumor burden and response per investigator assessment.
Of these 3 subjects, a best response of PR was achieved for 1
subject (33%) and SD for 2 subjects (67%). Nine subjects (75%) had
no measurable disease as a result of surgery and were not
considered evaluable for clinical response.
[0103] Immune Response to DNX-2401.
[0104] An immunological evaluation of resected tumors after
DNX-2401 treatment was conducted on a subset of subjects, based on
tumor availability. Two tumors from subjects enrolled in group A
(intratumoral injection) were resected several months after
DNX-2401 administration in response to what appeared to be tumor
progression or pseudo-progression. In both instances, pathologists
reported that the tumors were 80% and 90% necrotic with the
remaining tumor infiltrated by immune cells, including CD4 and CD8
T-cells. Analysis of resected tumors from subjects enrolled in
group B (intratumoral and intramural injection) provided further
evidence of tumor infiltration by macrophages at two weeks,
followed by the influx of T cells after several months. Overall,
this dynamic immune response may account for the persistence of
anti-glioma effects as well as the delayed treatment effect
observed during this study.
[0105] Study D24GBM.
[0106] Study D24GBM is an ongoing single center, Phase I,
Investigator-sponsored study being conducted at the University
Clinic of Navarra (Spain). The study is investigating a single dose
of 3e10 vp of DNX-2401 followed two weeks later by dose-dense
temozolomide administered for 8 weeks in subjects with glioblastoma
at first recurrence. Subjects receive either 1) a single
intratumoral injection during biopsy without further resection, 2)
an intramural injection and injection into the resection cavity
after tumor resection, or 3) an intratumoral injection of partially
resected tumor with injection into the resected cavity. As of
November 2015, all 31 planned subjects have received DNX-2401.
Based on preliminary data, no significant safety findings related
to DNX-2401 have been observed.
[0107] Study 2401BT-IFN-001.
[0108] Study 2401BT-IFN-001 (TARGET I) is an ongoing, randomized,
multi-center Phase 1b trial to investigate the safety of
administering 3e10 vp DNX-2401 into recurrent tumor followed 2
weeks later by treatment with gamma interferon in subjects with
recurrent glioblastoma or gliosarcoma. Enrolled subjects are
randomized to either DNX-2401 with interferon-gamma or to DNX-2401
alone in a 2:1 manner. As of March 2016, 26 of 24-36 evaluable
subjects have been enrolled. The protocol was amended to include
delivery of DNX-2401 by a cannula at a dose of 5e10 vp to a subset
of subjects. Preliminary data is not yet available.
[0109] C. Clinical Experience in Other Indications
[0110] Study 0643.
[0111] DNX-2401 has also been evaluated systemically in a completed
single-center, Phase I study in subjects (n=21) with recurrent
malignant gynecologic disease conducted as an
Investigator-sponsored study at the University of Alabama at
Birmingham. The study evaluated the feasibility and utility of
intraperitoneally delivered DNX-2401 for subjects with recurrent
epithelial ovarian cancer who had persistent or recurrent disease
following debulking and paclitaxel/platinum-based chemotherapy.
Cohorts of eligible subjects were treated with virus for three days
that was administered via an intraperitoneal catheter, to determine
the MTD. No DLTs were observed and an MTD was not reached with the
highest DNX-2401 dose tested: 1e12 vp/day for 3 days. No
vector-related Grade 3 or 4 toxicities were reported, and no
clinically significant laboratory abnormalities were noted. Adverse
events were limited to Grade 1/2 fever, fatigue, or abdominal pain.
Overall, AEs were considered unlikely or unrelated to virus and
deemed to be associated with underlying disease or the route of
delivery. Viral shedding studies showed insignificant shedding in
the serum, saliva, and urine. The maximum tolerated dose was not
reached. Over a one-month follow-up, 15 (71%) patients had stable
disease and six (29%) had progressive disease. Seven patients had a
decrease in CA-125; four had a drop >20%.
[0112] D. DNX-2401 Dosing
[0113] An MTD was not reached in Phase I studies. The potential
additive effects of intratumoral DNX-2401, followed by intravenous
pembrolizumab treatment are not known. Therefore, an initial phase
of dose escalation consisting of 3-cohorts (5e8 vp, 5e9 vp, and
5e10 vp; up to 12 subjects, total) to determine a safe intratumoral
dose of DNX-2401 in association with pembrolizumab is justified.
Single and repeat administration of DNX-2401 has been well
tolerated in multiple animal toxicity studies (e.g., MTD of ICB
7.5e10 vp in cotton rats) and early phase human studies (ID01-310,
D24GBM; preliminary 2401BT-IFN-001). In addition, intraperitoneal
DNX-2401 administered at a dose of up 1e12 vp/day for 3 consecutive
days has been shown to be well tolerated in humans. Modifications
to declared dose, volume, timing, may be necessary in certain
circumstances; however, changes will require approval from a
Medical Monitor or designee prior to implementation.
[0114] E. Method of Delivery
[0115] DNX-2401 is a conditionally replicative virus capable of
replication in tumor cells; however, the vector does not replicate
in surrounding normal brain tissue. Experience has shown that
direct intratumoral injection of DNX-2401 is safe and effective for
delivering DNX-2401 into the target tumor and increases the
potential of tumor cell exposure to the virus. Based on this
evidence, direct intratumoral administration of DNX-2401 via the
Alcyone MEMS Cannula (AMC.TM.) was selected for this Phase II study
to optimize virus delivery and provide a standardized
technique.
[0116] In a particular aspect, replication competent adenoviruses
are delivered via intratumoral injection into the brain. Direct
injection into the brain may be accomplished by a fine catheter or
cannula. With certain embodiments, the replication competent
oncolytic adenoviruses can be delivered by a microelectromechanical
(MEMS) system under MR intra-procedural guidance. In particular,
intratumoral injection into the brain is accomplished without
significant reflux or backflow by using cannula such as Alcyone
Lifesciences' Alcyone MEMS Cannula (AMC). Representative examples
of devices are described in U.S. Pat. No. 8,992,458 and U.S. Patent
Publications 2013/0035660, 2013/0035574 and 2013/0035560, each of
which are hereby incorporated by reference in their entirety.
[0117] F. Anti-PD-1 Therapeutics
[0118] Binding of PD-L1 on tumor cells to PD-1 on activated
effector T cells results in activation of PI3 kinase-signaling
cascade which in turn blocks the production of cytotoxic mediators
required for killing tumor cells. As used herein, a PD-L1 or PD-1
antagonist is a molecule that disrupts the interaction between
PD-L1 and PD-1. In one aspect, the anti-PD-1 therapeutic is an
antibody. Representative examples include the anti-PD-L1 antibody
MPDL3280A, or an anti-PD-1 antibody such as nivolumab
(Opdivo--Bristol Myers Squibb), or, lambrolizumab. Within another
embodiment the anti-PD-1 therapeutic is Pembrolizumab (as is
discussed in more detail below. Other drugs in early stage
development targeting PD-1 receptors (checkpoint inhibitors) are
Pidilizumab (CT-011, Cure Tech) and BMS-936559 (Bristol Myers
Squibb). Both Atezolizumab (MPDL3280A, Roche) and Avelumab (Merck
KGaA, Darmstadt, Germany & Pfizer) target the similar PD-L1
receptor.
[0119] Other anti-PD-1 therapeutics that can be delivered include
those described in US Patent Application Publication Nos.
2009/0217401, 20110195068 and 20120251537 and U.S. Pat. No.
8,217,149, the contents of each which are incorporated herein by
reference.
IV. PEMBROLIZUMAB
[0120] Pembrolizumab)(KEYTRUDA.RTM., a humanized monoclonal
antibody against the programmed death receptor-1 (PD-1) protein,
has been developed by Merck & Co. for the treatment of cancer.
Pembrolizumab is approved for treatment of melanoma in several
countries; in the US it is indicated for the treatment of advanced,
unresectable or metastatic malignant melanoma in patients with
disease progression after prior treatment with ipilimumab and, for
BRAF V600 mutation-positive patients, a BRAF inhibitor, while in
the EU it is approved for the treatment of advanced (unresectable
or metastatic) melanoma in adults. Pembrolizumab has also been
granted approval in the US for the treatment of patients with
metastatic NSCLC whose tumors express PD-L1 as determined by an
FDA-approved test and who have disease progression on or after
platinum-containing chemotherapy.
[0121] Pembrolizumab has demonstrated initial clinical efficacy in
single arm monotherapy trials in subjects with multiple tumor types
as determined by response rate. Ongoing clinical trials are being
conducted in a number of other advanced solid tumor indications and
hematologic malignancies.
[0122] A. Pharmaceutical and Therapeutic Background
[0123] The importance of intact immune surveillance in controlling
outgrowth of neoplastic transformation has been known for decades.
Accumulating evidence shows a correlation between
tumor-infiltrating lymphocytes (TILs) in cancer tissue and
prognosis in various malignancies..sup.Error! Bookmark not
defined.,Error! Bookmark not defined.,Error! Bookmark not
defined.,Error! Bookmark not defined.,Error! Bookmark not
defined.,Error! Bookmark not defined.,Error! Bookmark not
defined.,Error! Bookmark not defined.,Error! Bookmark not
defined.,Error! Bookmark not defined.,Error! Bookmark not
defined.,Error! Bookmark not defined.,Error! Bookmark not defined.
In particular, the presence of CD8+ T cells and the increased ratio
of CD8+ effector T cells/FoxP3+ regulatory T cells seem to
correlate with improved prognosis and long-term survival in many
solid tumors..sup.xxvii,xxvii,xxix,xxx,xxxi,xxxii
[0124] The PD-1 receptor-ligand interaction is a major pathway
hijacked by tumors to suppress immune control. The normal function
of PD-1, expressed on the cell surface of activated T cells under
healthy conditions, is to down-modulate unwanted or excessive
immune responses, including autoimmune reactions. PD-1 (encoded by
the gene Pdcd1) is an Ig superfamily member related to CD28 and
CTLA-4, which has been shown to negatively regulate antigen
receptor signaling upon engagement of its ligands (PD-L1 and/or
PD-L2). The structures of murine PD-1 alone and in complex with its
ligands were first resolved and more recently the NMR-based
structure of the human PD-1 extracellular region and analyses of
its interactions with its ligands were also reported. PD-1 and
family members are type I transmembrane glycoproteins containing an
Ig Variable-type (V-type) domain responsible for ligand binding and
a cytoplasmic tail which is responsible for the binding of
signaling molecules. The cytoplasmic tail of PD-1 contains 2
tyrosine-based signaling motifs, an immunoreceptor tyrosine-based
inhibition motif (ITIM) and an immunoreceptor tyrosine-based switch
motif (ITSM). Following T cell stimulation, PD-1 recruits the
tyrosine phosphatases SHP-1 and SHP-2 to the ITSM motif within its
cytoplasmic tail, leading to the dephosphorylation of effector
molecules, such as CD3.zeta., PKC.theta. and ZAP70, which are
involved in the CD3 T cell signaling cascade. The mechanism by
which PD-1 down-modulates T cell responses is similar to, but
distinct from that of CTLA-4. PD-1 was shown to be expressed on
activated lymphocytes, including peripheral CD4+ and CD8+ T cells,
B cells, T regs and Natural Killer cells. Expression has also been
shown during thymic development on CD4-CD8-(double-negative) T
cells, as well as subsets of macrophages and dendritic cells. The
ligands for PD-1 (PD-L1 and PD-L2) are constitutively expressed or
can be induced in a variety of cell types. PD-L1 is expressed at
low levels on various non-hematopoietic tissues, most notably on
vascular endothelium, whereas PD-L2 protein is only detectably
expressed on antigen-presenting cells found in lymphoid tissue or
chronic inflammatory environments..sup.Error! Bookmark not defined.
Both ligands are type I transmembrane receptors containing both
IgV- and IgC-like domains in the extracellular region and short
cytoplasmic regions with no known signaling motifs. Binding of
either PD-1 ligand to PD-1 inhibits T cell activation triggered
through the T cell receptor. PD-L2 is thought to control immune T
cell activation in lymphoid organs, whereas PD-L1 serves to dampen
unwarranted T cell function in peripheral tissues. Although healthy
organs express little, if any, PD-L1, a variety of cancers were
demonstrated to express abundant levels of this T cell inhibitor,
which, via its interaction with the PD-1 receptor on tumor-specific
T cells, plays a critical role in immune evasion by tumors. As a
consequence, the PD-1/PD-L1 pathway is an attractive target for
therapeutic intervention in cancer.
[0125] B. Non-Clinical Studies with Pembrolizumab
[0126] Therapeutic studies in mouse models have shown that
administration of antibodies blocking PD-1/PD-L1 interaction
enhances infiltration of tumor-specific CD8+ T cells and ultimately
leads to tumor rejection, either as a monotherapy or in combination
with other treatment modalities..sup.xlix,l,li,lii,liii,liv,lv
Anti-mouse PD-1 or anti-mouse PD-L1 antibodies have demonstrated
antitumor responses in models of squamous cell carcinoma,
pancreatic carcinoma, melanoma, acute myeloid leukemia and
colorectal carcinoma..sup.Error! Bookmark not defined.,Error!
Bookmark not defined.,Error! Bookmark not defined., In such
studies, tumor infiltration by CD8+ T cells and increased
IFN-.gamma., granzyme B and perforin expression were observed,
indicating that the mechanism underlying the antitumor activity of
PD-1 checkpoint inhibition involved local infiltration and
activation of effector T cell function in vivo..sup.Error! Bookmark
not defined. Experiments have confirmed the in vivo efficacy of
anti-mouse PD-1 antibody as a monotherapy, as well as in
combination with chemotherapy, in syngeneic mouse tumor models.
[0127] C. Pembrolizumab Dose Selection
[0128] The dose of pembrolizumab to be studied in this trial is 200
mg given intravenously every 3 weeks (Q3W). The dose recently
approved in the United States and several other countries for
treatment of melanoma subjects is 2 mg/kg Q3W. Information on the
rationale for selecting 200 mg Q3W is summarized below.
[0129] In KEYNOTE-001, an open-label Phase I study conducted to
evaluate the safety, tolerability, pharmacokinetics (PK),
pharmacodynamics (PD), and anti-tumor activity of pembrolizumab
when administered as monotherapy. The dose escalation portion of
this trial evaluated three dose levels, 1 mg/kg, 3 mg/kg and 10
mg/kg, administered every 2 weeks (Q2W) and dose expansion cohorts
evaluated 2 mg/kg Q3W and 10 mg/kg Q3W in subjects with advanced
solid tumors. All dose levels were well tolerated and no
dose-limiting toxicities were observed. This first-in-human study
of pembrolizumab showed evidence of target engagement and objective
evidence of tumor size reduction at all dose levels. No maximum
tolerated dose (MTD) has been identified. In addition, two
randomized cohort evaluations of melanoma subjects receiving
pembrolizumab at a dose of 2 mg/kg versus 10 mg/kg Q3W have been
completed, and one randomized cohort evaluating 10 mg/kg Q3W versus
10 mg/kg Q2W has also been completed. The clinical efficacy and
safety data demonstrate a lack of important differences in efficacy
or safety profile across doses.
[0130] An integrated body of evidence suggests that 200 mg every 3
weeks (Q3W) is expected to provide similar response to 2 mg/kg Q3W,
10 mg/kg Q3W and 10 mg/kg Q2W. Previously, a flat pembrolizumab
exposure-response relationship for efficacy and safety has been
found in subjects with melanoma in the range of doses between 2
mg/kg and 10 mg/kg. Exposures for 200 mg Q3W are expected to lie
within this range and will be close to those obtained with 2 mg/kg
Q3W dose.
[0131] A population pharmacokinetic (PK) model, which characterized
the influence of body weight and other patient covariates on
exposure, has been developed. The PK profile of pembrolizumab is
consistent with that of other humanized monoclonal antibodies,
which typically have a low clearance and a limited volume of
distribution. The distribution of exposures from the 200 mg fixed
dose are predicted to considerably overlap those obtained with the
2 mg/kg dose and importantly will maintain individual patient
exposures within the exposure range established in melanoma as
associated with maximal clinical response. Pharmacokinetic
properties of pembrolizumab, and specifically the weight-dependency
in clearance and volume of distribution are consistent with no
meaningful advantage to weight-based dosing relative to fixed
dosing.
[0132] In translating to other tumor indications, similarly flat
exposure-response relationships for efficacy and safety as observed
in subjects with melanoma can be expected, as the anti-tumor effect
of pembrolizumab is driven through immune system activation rather
than through a direct interaction with tumor cells, rendering it
independent of the specific tumor type. In addition, available PK
results in subjects with melanoma, NSCLC, and other tumor types
support a lack of meaningful difference in pharmacokinetic
exposures obtained at tested doses among tumor types. Thus the 200
mg Q3W fixed-dose regimen is considered an appropriate fixed dose
for other tumor indications as well.
[0133] A fixed dose regimen will simplify the dosing regimen to be
more convenient for physicians and to reduce potential for dosing
errors. A fixed dosing scheme will also reduce complexity in the
logistical chain at treatment facilities and reduce wastage. The
existing data suggest 200 mg Q3W as the appropriate dose for
pembrolizumab.
[0134] D. Commercial Dosage Forms
[0135] There are currently two formulations of KEYTRUDA.RTM..
KEYTRUDA.RTM. for injection is a sterile, preservative-free, and
white to off-white lyophilized powder in single-use vials. Each
vial is reconstituted and diluted for intravenous infusion. Each 2
mL of reconstituted solution contains 50 mg of pembrolizumab
formulated in L-histidine (3.1 mg), polysorbate 80 (0.4 mg), and
sucrose (140 mg) and may contain hydrochloric acid/sodium hydroxide
to adjust pH to 5.5. KEYTRUDA.RTM. solution for injection is a
sterile, preservative-free, clear to slightly opalescent, colorless
to slightly yellow solution that requires dilution for intravenous
infusion. Each vial contains 100 mg of pembrolizumab in 4 mL of
solution. Each 1 mL of solution contains 25 mg of pembrolizumab
formulated in: L-histidine (1.55 mg), polysorbate 80 (0.2 mg),
sucrose (70 mg), and water for injection, USP.
V. SUBJECTS
[0136] An evaluable subject is defined as an eligible subject who
receives at least one dose, or part of one dose, of study drug
(DNX-2401 and pembrolizumab) and one who completes the Day 28 study
visit.
[0137] Subjects who discontinue study participation for any reason,
other than progressive disease or study treatment-related toxicity,
prior to completing the Day 28 study visit, will not be evaluable
and will be replaced but will continue to be monitored for safety
and survival.
[0138] Subjects in general should meet the following inclusion
criteria for treatment. [0139] .gtoreq.18 years of age on the day
of informed consent [0140] A single glioblastoma or gliosarcoma
tumor with histopathological confirmation (if a prior diagnosis
exists such as anaplastic astrocytoma, or other tumor types that
have progressed to GBM, inclusion may be allowed following
discussion with a Medical Monitor or designee) [0141] Subjects with
contiguous tumors may be eligible with approval [0142] First or
presenting second recurrence of glioblastoma or gliosarcoma at the
time of consent [0143] Gross total or partial tumor resection,
including tumor debulking, is not possible or planned [0144] A
single measurable tumor that is at least 10.0 mm longest diameter
(LDi).times.10.0 mm shortest diameter (SDi) and this tumor does not
exceed 40.0 mm in LDi or SDi [0145] The measurable area of the
tumor is solid/nodular and is not cystic (most tumors will have
cystic components and subjects who have a tumor that is partially
cystic may be eligible for the study with approval or if the
surgeon can perform an injection within the measurable solid parts
of the tumor) [0146] Willing to provide a tumor tissue sample for
biomarker analysis and mutational status from an archived slide or
newly obtained stereotactic core biopsy [0147] Tumors must be
accessible for stereotactic injection [0148] Confirmatory evidence
of tumor recurrence (e.g., progression after last treatment) on the
screening MRI 15 days to minus 72 hours, inclusive, prior to
planned start time [0149] Tumor wherein the location will not risk
delivery of virus into the ventricular system [0150] Tumor
recurrence or progression documented after previously failing
surgical resection, chemotherapy and/or radiation [0151] Resolution
of toxic effect(s) of the most recent prior chemotherapy to Grade 1
or less (except neuropathy and alopecia) [0152] Note: Subjects with
.ltoreq.Grade 2 neuropathy and/or alopecia are an exception to this
criterion and they will not be excluded for these reasons alone.
[0153] Demonstrate adequate organ function as defined below: [0154]
Hematological [0155] Absolute neutrophil count
(ANC).gtoreq.1,500/mcL [0156]
WBC.gtoreq.2.5.times.10.sup.3/mm.sup.3 [0157]
Platelets.gtoreq.100,000/mcL [0158] Hemoglobin.gtoreq.10 g/dL or
.gtoreq.5.6 mmol/L [0159] Renal [0160]
Creatinine.ltoreq.1.5.times.ULN [0161] BUN.ltoreq.1.5.times.ULN
[0162] Hepatic [0163] Total bilirubin.ltoreq.1.5.times.ULN or
Direct bilirubin.ltoreq.ULN for subjects with total bilirubin
levels >1.5.times.ULN [0164] AST (SGOT) and ALT
(SGPT).ltoreq.2.5.times.ULN [0165] Coagulation [0166] International
Normalized Ratio (INR).ltoreq.1.5.times.ULN [0167] Prothrombin Time
(PT).ltoreq.1.5.times.ULN [0168] Activated Partial Thromboplastin
Time (aPTT).ltoreq.1.5.times.ULN [0169] Adequate venous access
[0170] Karnofsky performance status.gtoreq.70% [0171] Afebrile at
baseline/Day 0 prior to DNX-2401 administration (i.e.,
<38.0.degree. C.)
[0172] Prior anti-tumor therapies must have been completed within
the following time periods prior to DNX-2401 injection: [0173] 4
weeks after nitrosoureas [0174] 2 weeks after vincristine [0175] 3
weeks after procarbazine or temozolomide [0176] 4 weeks after
bevacizumab, other antibody therapy or other anti-angiogenic
therapy to treat glioblastoma [0177] 5 half-lives for other
anti-cancer agents including investigational agents (or 2 weeks
after the last dose when the half-life is unknown). A discussion of
these agents will take place with a Medical Monitor or designee
prior to establishing eligibility. [0178] For applicable screening
candidates, external beam radiotherapy (>5000 cGy) must have
been completed at least 12 weeks prior to DNX-2401
administration
[0179] Females should not be pregnant and should take steps to
remain non-pregnant, such as by using twice the normal protection
of birth control (i.e., double-barrier) by using a condom and
spermicidal jelly or foam, or a diaphragm and spermicidal jelly or
foam. A spermicidal jelly or foam should be used in addition to a
barrier method (e.g., condom or diaphragm). Male subjects should
use an acceptable method of contraception throughout the study
starting with DNX-2401 administration through 180 days after the
single dose of DNX-2401 and 120 days after the last dose of
pembrolizumab.
[0180] Patients must be willing and able to provide informed
consent, undergo and comply with all study assessments and adhere
to the protocol schedule, and agree not to donate blood or gametes
following virus administration.
VI. EXAMPLES
[0181] The following examples are included to demonstrate preferred
embodiments of the disclosure. It should be appreciated by those of
skill in the art that the techniques disclosed in the examples
which follow represent techniques discovered by the inventor to
function well in the practice of the disclosure, and thus can be
considered to constitute preferred modes for its practice. However,
those of skill in the art should, in light of the present
disclosure, appreciate that many changes can be made in the
specific embodiments which are disclosed and still obtain a like or
similar result without departing from the spirit and scope of the
disclosure.
EXAMPLES
Example 1
Study Parameters
[0182] Objectives.
[0183] Primary objectives are to evaluate the safety of escalating
doses of DNX-2401 and the overall safety of the declared dose of
intratumoral DNX-2401 when followed by sequential intravenous
administration of pembrolizumab, and to determine the objective
response rate (ORR). Secondary objectives are to determine overall
survival (OS-12) and to determine the clinical benefit rate (CBR;
defined as CR+PR+SD) per RANO criteria and RANO criteria modified
to account for pseudo-progression (e.g., iRANO). Exploratory
objectives are to evaluate overall survival (OS), OS-8, progression
free survival (PFS), progression free survival at six months
(PFS-6), time to tumor response, duration of response, per RANO
criteria and RANO criteria modified to account for
pseudo-progression (e.g., iRANO), to evaluate changes in Karnofsky
performance status (KPS) and neurologic status, and to evaluate
cytokines, lymphocyte sub-types and other potential biomarkers
(e.g., PD-1, PD-L1 expression, genetic mutation).
[0184] Subjects. Inclusion criteria are set forth above. Exclusion
criteria are as follows: [0185] Recurrent GBM with multiple
(.gtoreq.2) separate enhancing tumors (measurable or
non-measurable) [0186] Tumor shape that is bi-lobular or multifocal
[0187] Tumor involvement that would require ventricular, brainstem
or posterior fossa injection or access through a ventricle or risk
of ventricular penetration in order to deliver DNX-2401 [0188]
Tumor involves both hemispheres or involves the subependyma or
suspected cerebrospinal fluid (CSF) dissemination (extension of the
tumor across the midline of the corpus callosum is consistent with
contralateral hemisphere (bi-hemispheric) involvement which is
prohibited) [0189] Documented extracranial metastases [0190]
Requires treatment with high-dose systemic corticosteroids defined
as dexamethasone >4 mg/day or bioequivalent for at least 3
consecutive days within 2 weeks of start of pembrolizumab (an
alternative management strategy may be considered following prior
consideration) [0191] Uncontrolled blood-sugar levels defined as
HbA1c >7% on 2 separate measurements [0192] Active autoimmune
disease that requires, or has required, systemic treatment in the
past 2 years (i.e., with use of disease-modifying agents,
corticosteroids or immunosuppressive drugs) or replacement therapy
(e.g., thyroxine, insulin, or physiologic corticosteroid
replacement therapy for adrenal or pituitary insufficiency, etc.)
is not considered a form of systemic treatment [0193] Previous
treatment with anti-PD1 or PD-L1 agents including pembrolizumab
[0194] Evidence of active, non-infectious pneumonitis [0195] A
history of interstitial lung disease [0196] Transfusions or
medications (e.g., G-CSF) to treat pancytopenia or other
hematological conditions within 4 weeks prior to DNX-2401
administration [0197] Prior gene transfer therapy or prior therapy
with a cytolytic virus of any type [0198] Live vaccines of any kind
within 45 days prior to DNX-2401 administration and while
participating in the study other than seasonal influenza killed
virus vaccines for injection [0199] Major surgery within 4 weeks
and minor surgery within 2 weeks of DNX-2401 administration (if
subject received major surgery, they must have fully recovered from
the toxicity and/or complications from the intervention prior to
receiving DNX-2401) [0200] Currently participating and receiving
investigational agent(s) or has participated in a study of an
investigational agent(s) and received investigational agent(s) or
used an investigational device within 30 days prior to DNX-2401
administration [0201] Any contraindication for undergoing MRI
[0202] Is pregnant or breastfeeding, or expecting to conceive or
father children during the study, starting with the screening visit
through 180 days after the single dose of DNX-2401 and 120 days
after the last dose of pembrolizumab [0203] Evidence of active
uncontrolled infection or an unstable or severe intercurrent
medical condition that requires treatment and/or precludes surgery
[0204] History of prior malignancy except for curatively treated
basal or squamous cell carcinoma of the skin (non-melanoma skin
cancer), cervical or vaginal intra-epithelial neoplasia,
non-invasive breast cancer in situ or localized prostate cancer
with a current prostate-specific antigen (PSA) of <4.0 ng/mL
(mcg/L) (subjects with other curatively treated malignancies who
have had no evidence of metastatic disease and a >2 year
disease-free interval may be enrolled after approval) [0205] Any
medical condition that precludes intratumoral injection into the
brain tumor [0206] Immunocompromised subjects or those with
autoimmune conditions, active hepatitis (HAV, HBV, HCV), known
history of active TB (Mycobacterium tuberculosis), or human
immunodeficiency virus (HIV) seropositivity (TB testing is required
for subjects recently exposed to persons with active tuberculosis
or who have traveled recently to areas where TB is endemic) [0207]
Active hepatitis B is defined as anti-HBc core antibody and HBsAg
surface antigen positivity [0208] Evidence of bleeding diathesis or
use of anticoagulant medication or any medication that may increase
the risk of bleeding that cannot be stopped more one week prior to
surgery (low weight heparin and drugs in the anticoagulant class
(e.g., Lovenox (enoxaparin) administered on a temporary, limited
basis for post-procedure deep vein thrombosis (DVT) prophylaxis are
permitted) [0209] Encephalitis, multiple sclerosis or other central
nervous system (CNS) infection or primary CNS disease that would
interfere with subject evaluation [0210] Li-Fraumeni Syndrome or
with a known germ line defect in the retinoblastoma gene or its
related pathways [0211] Significant systemic or major illnesses
including but not limited to: congestive heart failure, ischemic
heart disease, kidney disease or renal failure, or organ
transplantation [0212] Alcohol or substance abuse or alcohol
dependency that is active within 12 months prior to screening that
has caused health consequences [0213] History or current diagnosis
of any medical or psychological condition that in the
Investigator's opinion, might interfere with the subject's ability
to participate in the study or the inability to obtain informed
consent because of cognitive, psychiatric or complicating medical
problems
[0214] Study Duration.
[0215] Up to 105 weeks of treatment (approximately 24 months) with
follow-up through 90 days following the final dose of
pembrolizumab. Thereafter, long-term survival follow-up will be
performed every 16 weeks for response, survival, study drug-related
adverse events and adverse events resulting in death.
[0216] Study Design.
[0217] The protocol involves a single intratumoral administration
of DNX-2401 with sequentially administered intravenous
pembrolizumab for subjects with first or second glioblastoma or
gliosarcoma recurrence for whom gross total or partial resection is
not possible or planned. In the initial phase of the study, up to a
total of 12 eligible subjects will participate in the
dose-escalation process and will be enrolled using a 3+3 study
design in dose-escalating cohorts as follows: [0218] Cohort 1:
Single dose DNX-2401 (5.times.10.sup.8 vp) delivered intratumorally
followed by sequential intravenous pembrolizumab every 3 weeks
(Q3W) [0219] Cohort 2: Single dose DNX-2401 (5.times.10.sup.9 vp)
delivered intratumorally followed by sequential intravenous
pembrolizumab every 3 weeks (Q3W) [0220] Cohort 3: Single dose
DNX-2401 (5.times.10.sup.10 vp) delivered intratumorally followed
by sequential intravenous pembrolizumab every 3 weeks (Q3W)
[0221] Three (3) subjects will be enrolled in Cohort 1 and will be
followed for 21 days after the initial dose of pembrolizumab for
evaluation of dose-limiting toxicity (DLT). If no DLTs are
observed, and following a review of clinical data (e.g., laboratory
results, adverse events), the next cohort will be enrolled. If, in
any cohort, 1 DLT is observed out of the initial 3 subjects, then 3
additional subjects will be added to the same cohort. The dose
escalation will continue until at least 2 subjects experience DLTs
or the highest dose cohort of 5.times.. 10.sup.10 vp is completed
and all subjects have been closely monitored for safety for 21 days
following administration of the first dose of pembrolizumab on Day
7. If 2 or more subjects have a DLT, then the previous dose will be
considered the declared Phase 2 dose. If, after safety review of
all dosed cohorts, there are no DLTs in the first 3 subjects or
<2 DLTs in the expanded cohort of 6 subjects, this cohort will
be deemed safe and tolerable.
[0222] Following the initial enrollment and dose-determination
phase, enrollment will continue at the declared dose for up to 36
additional eligible subjects. These subjects will receive a single
intratumoral DNX-2401 dose via cannula on Day 0, or via needle, and
managed per the institution's standard policy for post-operative
care. All subjects who enroll will also begin pembrolizumab
infusions 7-9 days following administration of DNX-2401.
[0223] Prior to virus delivery, a stereotactically-guided biopsy of
the tumor will be obtained and analyzed via frozen section to
confirm the presence of recurrent tumor at the planned intratumoral
injection target. In addition to histopathological analysis, the
tissue will be analyzed for mutational status, other potential
biomarkers, and archived.
[0224] Pembrolizumab will be infused intravenously at a dose of 200
mg over 30 minutes every 3 weeks beginning on Day 7 (+2-day window)
and continuing for up to 105 weeks or until progressive disease is
confirmed. On Day 7, if subjects are experiencing any AEs following
the DNX-2401 administration procedure on Day 0, the AEs must be
.ltoreq.Grade 1 or returned to baseline status prior to receiving
the first dose of intravenous pembrolizumab.
[0225] Subjects who complete the treatment phase and End of Study
visit will be subsequently monitored through Week 115 (i.e., 12
weeks or 90 days following final dose of pembrolizumab at Week
103). End of study, post-pembrolizumab interval safety status will
be reviewed at the first long-term survival follow-up visit (Week
119). Thereafter, subjects will be followed for response, survival,
study drug-related adverse events, and adverse events resulting in
death (regardless of relationship) every 16 weeks.
[0226] Subjects who withdraw or terminate from the study early will
have an assessment of adverse events and serious adverse events
(including Events of Clinical Interest (ECI)) for 90 days
post-study drug administration or 30 days post-study drug
administration if anti-cancer therapy is initiated. Thereafter,
subjects will be followed for response, survival, study
drug-related adverse events, and adverse events resulting in death
(regardless of relationship) every 8 weeks.
[0227] Safety monitoring will be performed throughout the study.
Monitoring will include adverse events and serious adverse events
(including Events of Clinical Interest (ECI)), laboratory
evaluations, physical and neurological examinations, vital sign
measurements, and MRI, including the post-DNX-2401 injection
procedure MRI (within 36 hours post DNX-2401 administration) to
assess for hematoma.
[0228] Additionally, other exploratory assessments may include
changes in cytokines, lymphocyte sub-types and other potential
biomarkers (e.g., PD-1 and PD-L1). Changes in KPS, and neurologic
status will also be evaluated.
Tumor response will be determined by MRI performed every 4 weeks up
to Week 28. The Week 28 visit corresponds to approximately 6 months
post-DNX-2401 administration and may take place no sooner than 182
days following Day 0. In support of this, the Week 28 (Cycle 10)
visit is conducted between 194 and 198 days post-DNX-2401
administration. After Week 28 (Month 6) tumor response will
continue to be followed with MRI every 8 weeks until Week 105
(approximately 24 months) post-DNX-2401 and every 16 weeks
thereafter (long-term survival follow-up). Because DNX-2401 appears
to work, at least in part, by eliciting an antitumor immune
response, objective response will be evaluated by RANO criteria and
RANO criteria modified to account for pseudo-progression (e.g.,
iRANO).
[0229] In addition to Investigator assessments, MR images will be
collected for independent central review. Partial and complete
responses will be confirmed at a minimum of 4 weeks later (28
days+3-day window) by repeat MRI. In the case of suspected
progressive disease by MRI, a confirmatory MRI will be performed at
a minimum of 4 weeks later (28 days +3-day window). The
neuroradiologist or other qualified PI-designee will make tumor
measurements and assessments required for clinical treatment
decisions. In the event of progressive disease on MRI in a subject
who is clinically stable, the investigator may exercise discretion
by keeping the subject on-study. Because tumor regression has been
demonstrated >5 months or later, following the intratumoral
delivery of DNX-2401, every effort should be made to manage
clinical signs and symptoms before initiating an alternative
therapy.
[0230] If necessary, each subject should receive the lowest
possible stable steroid dose for 7 days prior to DNX-2401
administration and for 3 days prior to any response/disease
assessment MRI. For suspected autoimmune events or conditions and
pseudoprogression and/or tumor swelling (e.g., appearance of
central nervous system (CNS) symptoms or worsening thereof),
pulse-dosing of steroids (e.g., 4 mg BID.times.3 days, then 4 mg
daily.times.2 days, then off) is recommended.
[0231] During the two weeks preceding the first dose of
pembrolizumab, steroids are to be restricted and may not exceed 4
mg dexamethasone per day for 3 consecutive days within 2 weeks
prior to the first dose of pembrolizumab on study Day 7 (+2-day
window) per protocol eligibility exclusion criterion. Following
completion of the Week 4 (Day 28) study visit, in the event that
pulse-dosing with steroids is not effective in the control of tumor
swelling/edema, low-dose (3-5 mg/kg every 2 weeks.times.3)
bevacizumab (Avastin.RTM.) may be used at the Principal
Investigator's discretion to address cerebral edema in lieu of
systemic steroid administration.
[0232] If bevacizumab (Avastin.RTM.) is used to address cerebral
edema, the lowest possible stable dose should be used for 3 days
prior to any response/disease assessment MRI. Avastin.RTM. should
not be initiated within 28 days before or after Day 0/surgery to
administer DNX-2401 or subsequent surgery for other reasons due to
known complications in surgery/wound healing and hemorrhage.
[0233] Anticonvulsants may also be administered to manage and
prevent seizures in subjects with a pre-existing medical history of
seizures, otherwise they should not be administered
prophylactically.
[0234] Safety Assessments.
[0235] Safety assessments will include: collection of adverse event
and serious adverse event data (including Events of Clinical
Interest (ECI)), physical and neurological examinations, laboratory
examinations, vital signs measurements, and MRI (including the MRI
obtained within 36 hours post DNX-2401 administration to assess for
possible post-procedure hematoma). The safety reporting period
begins at Informed Consent for SAEs and Day 0 for AEs and continues
through 12 weeks (90 days) after the last dose of study drug. Study
drug-related adverse events and adverse events that result in death
(regardless of relationship) will be reported from Day 0 and
through long-term survival follow-up every 16 weeks (every 8 weeks
for cases of early termination).
[0236] Statistical Methods.
[0237] Sample size: Up to a total of 48 evaluable subjects may be
enrolled. In the initial dose escalation phase, the safety and
incidence of dose limiting toxicities (DLTs) that occur within 21
days post the first infusion of pembrolizumab for each cohort will
be summarized.
[0238] The primary endpoint [objective response rate (ORR)] defined
as a complete response (CR) or a partial response (PR), will be
tested in a single arm (group) design. The sample size estimation
was based on a historical control response rate equal to
5%..sup.Error! Bookmark not defined.,Error! Bookmark not defined.
The null hypothesis to be tested is that the DNX-2401 treated
population has the same ORR as the historical control rate. A
one-sided test with an alpha=0.05 and N of 36 (declared Phase 2
dose) will yield 80% power for an alternative hypothesis of
ORR=18%.
[0239] Analysis Sets:
[0240] Safety Analysis Set (SAS): This set will include subjects
who receive at least one dose (or part of one dose) of study drug
(DNX-2401 and/or pembrolizumab). Typically the SAS would include
subjects who do not have measurable tumor at baseline; however, to
be eligible for the study, all subjects must have measurable
disease.
[0241] Evaluable Analysis Set (EAS):
[0242] Based on the intent-to-treat principle, this set will
include subjects who receive at least one dose (or part of one
dose) of DNX-2401 and pembrolizumab and who have measurable tumor
at baseline.
[0243] Safety Analyses:
[0244] All safety analyses will be performed on the Safety Analysis
Set. Adverse events (AEs) will be graded according to Version 4.03
(Jun. 14, 2010) of the National Cancer Institute-Common Terminology
Criteria for Adverse Events (NCI-CTCAE), or according to a
protocol-defined severity scale determined by the Investigator for
events not listed in the CTCAE. DNX-2401 treatment-emergent AEs are
those defined as having an onset after the start time of DNX-2401
administration and continue through week 105 (approximately 24
months) or Early Termination, whichever occurs first. Pembrolizumab
treatment-emergent AEs are those defined as having an onset after
the first dose of pembrolizumab up through 90 days after the last
dose of pembrolizumab or Early Termination (whichever comes first).
Treatment-emergent AEs for DNX-2401 and pembrolizumab will be
summarized separately using the most current version of the Medical
Dictionary for Regulatory Activities (MedDRA) by System Organ Class
(SOC) and Preferred Term (PT), classified from
Investigator-reported verbatim terms. The number and percentage of
subjects with at least one occurrence of a preferred term will be
summarized, according to the most severe grade using a 5-point
scale (Grade 1-5). The number of events per preferred term will
also be summarized. Maximum severity and causality (relationship to
study drug) will be summarized separately.
[0245] The number and percentage of subjects reporting AEs, SAEs,
related AEs, related SAEs, .gtoreq.Grade 3 AEs, related
.gtoreq.Grade 3 AEs, and AEs leading to withdrawal, or treatment
discontinuation will be summarized by dose according to SOC and
preferred term. AEs will also be presented in subject listings. The
duration of AEs will be determined and included in listings, along
with the action taken and outcome. Subject listings and narratives
will also be provided for SAEs, AEs resulting in discontinuation,
and death.
[0246] Medical history and concurrent illnesses will be summarized
and may be examined as potential confounders in the
treatment-response relationship. Prior and concomitant medications
and therapies will be coded using the latest version of the World
Health Organization Drug Dictionary (WHO-DD) and will be
summarized.
[0247] Laboratory results will be classified according to NCI-CTCAE
and summarized by parameter and by time point. Laboratory results
not corresponding to a coded term will not be graded. The incidence
of laboratory abnormalities will be summarized. The worst on-study
grade after DNX-2401 administration and after pembrolizumab will be
summarized. The incidence of .gtoreq.Grade 3 laboratory
abnormalities and shifts in toxicity grading from baseline to
highest grade post-baseline will also be displayed. Laboratory
values will be listed by subject and values outside of a normal
reference range will be flagged. Pregnancy test results will be
listed separately.
[0248] Vital sign measurements will be summarized by visit using
descriptive statistics. Physical examination findings will be
summarized by visit and will also be presented in subject
listings.
[0249] Efficacy Analyses:
[0250] All efficacy analyses will be performed on the Evaluable
Analysis Set (EAS). The primary efficacy endpoint will be the
objective response rate (ORR), defined as the sum (and percent) of
those subjects with a complete response (CR) or partial response
(PR) as the best overall response recorded from the start of
treatment until progressive disease or at the latest time point
obtained.
[0251] Objective response determined from MRI review will be based
upon the RANO criteria and RANO criteria modified to account for
pseudo-progression (e.g., iRANO) with the primary analysis based on
RANO criteria (as assessed by the central reader). ORR will be
summarized by the number and percentage of subjects. Corresponding
95% and 90% confidence intervals (CIs) will be based upon the
binomial exact method.
[0252] Complete, partial responses and suspected progression will
be confirmed. Any partial or complete response noted on MRI will be
confirmed by repeat MRI at a minimum of 4 weeks (28 days with
+3-day window) later. In the case of suspected tumor progression by
MRI, and with the absence of deteriorating clinical status as
determined by the Investigator, a repeat MRI will be performed, if
possible, at a minimum of 4 weeks (28 days with +3-day window)
later in an attempt to confirm progressive disease. In the case of
suspected progressive disease, confirmation by tumor biopsy is
strongly recommended. Additionally, if the tumor is resected prior
to initiating an alternative cancer therapy, tissue should also be
collected for histopathology, molecular assays (e.g., sequencing,
gene expression, immunohistochemistry) and archived for future
analysis related to this study.
[0253] Secondary efficacy endpoints will include OS-12 and CBR.
OS-12 will be summarized using Kaplan-Meier (KM) methods; all
estimates will be presented with 95% confidence intervals. CBR will
be summarized as the number and proportion of subjects with
clinical benefit, along with 95% CIs for the proportion of subjects
with clinical benefit based on the binomial exact method.
[0254] Additional efficacy endpoints will be summarized based on
the EAS. Other exploratory efficacy endpoints will include: OS,
OS-8, PFS, PFS-6, time to response, duration of response per RANO
criteria and RANO criteria modified to account for
pseudo-progression (e.g., iRANO). All such endpoints will be
summarized using Kaplan-Meier methods with estimates presented
along with 95% CIs.
[0255] OS-8 and OS-12 will be presented as 8-month and 12-month
survival estimates along with 95% CIs. The OS endpoint is defined
as the overall survival profile of all subjects while on study and
will be summarized with KM survival curves as well as quartile
estimates (25%, median, 75%) and associated 95% CIs.
[0256] Population subgroup analyses (e.g., cytokines, lymphocyte
sub-types, if performed) may be determined and subgroup analyses
performed. In addition, correlation analyses based on parameters
such as immunological effects may also be performed.
[0257] Changes from baseline in KPS and neurologic status will also
be summarized at each time point by subject.
[0258] All of the compositions and/or methods disclosed and claimed
herein can be made and executed without undue experimentation in
light of the present disclosure. While the compositions and methods
of this disclosure have been described in terms of preferred
embodiments, it will be apparent to those of skill in the art that
variations may be applied to the compositions and/or methods and in
the steps or in the sequence of steps of the method described
herein without departing from the concept, spirit and scope of the
disclosure. More specifically, it will be apparent that certain
agents which are both chemically and physiologically related may be
substituted for the agents described herein while the same or
similar results would be achieved. All such similar substitutes and
modifications apparent to those skilled in the art are deemed to be
within the spirit, scope and concept of the disclosure as defined
by the appended claims.
[0259] The following are some specific numbered embodiments of the
systems and processes disclosed herein. These embodiments are
exemplary only. It will be understood that the invention is not
limited to the embodiments set forth herein for illustration, but
embraces all such forms thereof as come within the scope of the
above disclosure. [0260] 1) A method of treating a subject having
brain tumor comprising: [0261] (a) administering an oncolytic
adenovirus to said subject; and [0262] (b) administering an
anti-PD-1 antibody to said subject. [0263] 2) The method of
embodiment 1, wherein the oncolytic adenovirus is an adenovirus
serotype 5 strain, is selectively replication competent in cells
defective in the Rb/p16 tumor suppressor pathway, contains a
deletion of the 24 nucleotides encoding amino acids 122 to 129 of
the adenoviral E1A protein, and/or contains an integrin binding
RGD-4C motif. [0264] 3) The method of embodiments 1-2, wherein the
oncolytic adenovirus is DNX-2401. [0265] 4) The method of
embodiments 1-3, wherein the anti-PD1 antibody is a humanized
antibody. [0266] 5) The method of embodiments 1-4, wherein the
anti-PD1 antibody is pembrolizumab. [0267] 6) The method of
embodiments 1-5, wherein the oncolytic adenovirus is delivered
intratumorally. [0268] 7) The method of embodiment 6, wherein the
oncolytic adenovirus is delivered via cannula or needle. [0269] 8)
The method of embodiments 1-7, wherein the oncolytic adenovirus is
delivered at 5.times.10.sup.8 viral particles per dose,
5.times.10.sup.9 viral particles per dose, or 5.times.10.sup.10
viral particles per dose. [0270] 9) The method of embodiments 1-8,
wherein the anti-PD1 antibody is delivered by intravenous infusion.
[0271] 10) The method of embodiments 1-9, wherein the anti-PD1
antibody is delivered at 200 mg per dose. [0272] 11) The method of
embodiment 10, wherein the dose is administered over 30 minutes.
[0273] 12) The method of embodiments 1-11, wherein a single dose of
the oncolytic adenovirus is provided prior to three consecutive
doses of the anti-PD1 antibody. [0274] 13) The method of embodiment
12, wherein the time between oncolytic adenovirus administration
and the first administration of the anti-PD1 antibody is about 7-9
days. [0275] 14) The method of embodiments 12-13, wherein the time
between sequential administrations of the anti-PD1 antibody is
about three weeks. [0276] 15) The method of embodiments 1-14,
further comprising administering to the subject one or more of a
steroid, an anticonvulsant or an antibody that inhibits vascular
endothelial growth factor A. [0277] 16) The method of embodiment
12, wherein treating further comprises additional administrations
of the anti-PD1 antibody for up to a 105 weeks or 24 months from
the date of oncolytic adenovirus administration. [0278] 17) The
method of embodiments 1-16, wherein said subject is evaluated for
one of more of overall survival, tumor response, clinical benefit
rate, Karnofsky performance status, neurologic status, cytokine
levels, lymphocyte levels, or a biomarker. [0279] 18) The method of
embodiment 17, wherein said biomarker is PD-1 level or PDL-1 level.
[0280] 19) The method of embodiment 18, wherein tumor response is
measured by MRI. [0281] 20) The method of embodiments 1-19, further
comprising stereotactically-guided biopsy of the brain tumor.
[0282] 21) The method of embodiments 1-20, wherein the brain tumor
is glioblastoma multiforme. [0283] 22) The method of embodiments
1-20, wherein the brain tumor is gliosarcoma. [0284] 23) The method
of embodiments 1-22, wherein the subject exhibits an overall
survival of at least 9 months, 12 months, 15 months, 18 months, 24
months, 36 months or 48 months, any interval therebetween. [0285]
24) The method of embodiments 1-22, wherein the subject exhibits an
increase in overall survival, as compared to an untreated control
subject, of at least 6 months, 9 months, 12 months, 15 months, 18
months, 24 months, 36 months or 48 months, any interval
therebetween. [0286] 25) The method of embodiments 1-22, wherein
the tumor exhibits reduced growth, no growth, a 10% reduction in
tumor mass, a 20% reduction in tumor mass, a 30% reduction in tumor
mass, a 40% reduction in tumor mass, a 50% reduction in tumor mass,
a 60% reduction in tumor mass, a 70% reduction in tumor mass, an
80% reduction in tumor mass, a 90% reduction in tumor mass, or a
100% reduction in tumor mass following initiation of treatment.
[0287] 26) The method of embodiments 1-22, wherein the Karnofsky
performance status improves or remains unchanged following step
(b). [0288] 27) The method of embodiments 1-22, wherein there is a
statistically measurable clinical benefit. [0289] 28) The method of
embodiments 1-27, wherein the subject exhibits recurrent or
progressive brain tumor following previous treatment. [0290] 29)
The method of embodiments 1-28, wherein the previous treatment was
chemotherapy, radiotherapy, or antibody therapy. [0291] 30) The
method of embodiments 1-29, wherein the subject is a human. 31) A
composition comprising: [0292] (a) an oncolytic adenovirus; and
[0293] (b) an anti-PD-1 antibody. [0294] 32) The composition of
embodiment 31, wherein the oncolytic adenovirus is an adenovirus
serotype 5 strain, and/or wherein the oncolytic adenovirus is
selectively replication competent in cells defective in the Rb/p16
tumor suppressor pathway, and/or wherein the oncolytic adenovirus
comprises a deletion of the 24 nucleotides encoding amino acids 122
to 129 of the adenoviral E1A protein, and/or wherein the oncolytic
adenovirus comprises an integrin binding RGD-4C motif. [0295] 33)
The composition of embodiments 31-32, wherein the oncolytic
adenovirus is DNX-2401. [0296] 34) The composition of embodiments
31-33, wherein the anti-PD1 antibody is a humanized antibody.
[0297] 35) The composition of embodiments 31-34, wherein the
anti-PD1 antibody is pembrolizumab. [0298] 36) The composition of
embodiments 31-35, wherein the composition is formulated for
intratumoral delivery. [0299] 37) The composition of embodiments
31-35, wherein the composition is formulated for intravenous
infusion. [0300] 38) The composition of embodiments 31-37, wherein
the oncolytic adenovirus is comprised in a unit dose of
5.times.10.sup.8 viral particles per dose, 5.times.10.sup.9 viral
particles per dose, or 5.times.10.sup.10 viral particles per dose.
[0301] 39) The composition of embodiments 31-38, wherein the
anti-PD1 antibody is comprised in a unit dose of 200 mg. [0302] 40)
The composition of embodiments 31-39, further comprising one or
more of a steroid, an anticonvulsant or an antibody that inhibits
vascular endothelial growth factor A. [0303] 41) A composition
comprising an oncolytic adenovirus of adenovirus serotype 5 strain,
wherein the oncolytic adenovirus is selectively replication
competent in cells defective in the Rb/p16 tumor suppressor
pathway, wherein the oncolytic adenovirus comprises a deletion of
the 24 nucleotides encoding amino acids 122 to 129 of the
adenoviral E1A protein, and wherein the oncolytic adenovirus
comprises an integrin binding RGD-4C motif. [0304] 42) The
composition of embodiment 41, wherein the oncolytic adenovirus is
DNX-2401. [0305] 43) The composition of embodiments 41-42, wherein
the oncolytic adenovirus is comprised in a unit dose of
5.times.10.sup.8 viral particles per dose. [0306] 44) The
composition of embodiments 41-42, wherein the oncolytic adenovirus
is comprised in a unit dose of 5.times.10.sup.9 viral particles per
dose. [0307] 45) The composition of embodiments 41-42, wherein the
oncolytic adenovirus is comprised in a unit dose of
5.times.10.sup.10 viral particles per dose.
[0308] Any of the various embodiments described above can be
combined to provide further embodiments. All of the U.S. patents,
U.S. patent application publications, U.S. patent applications, PCT
application publications, foreign patents, foreign patent
applications and non-patent publications referred to in this
specification, are incorporated herein by reference, in their
entirety. Aspects of the embodiments can be modified, if necessary
to employ concepts of the various patents, applications and
publications to provide yet further embodiments. These and other
changes can be made to the embodiments in light of the
above-detailed description. In general, in the following claims,
the terms used should not be construed to limit the claims to the
specific embodiments disclosed in the specification and the claims,
but should be construed to include all possible embodiments along
with the full scope of equivalents to which such claims are
entitled. Accordingly, the claims are not limited by the
disclosure.
[0309] All patents, publications, scientific articles, web sites,
and other documents and materials referenced or mentioned herein
are indicative of the levels of skill of those skilled in the art
to which the invention pertains, and each such referenced document
and material is hereby incorporated by reference to the same extent
as if it had been incorporated by reference in its entirety
individually or set forth herein in its entirety. Applicants
reserve the right to physically incorporate into this specification
any and all materials and information from any such patents,
publications, scientific articles, web sites, electronically
available information, and other referenced materials or
documents.
[0310] The written description portion of this patent includes all
claims. Furthermore, all claims, including all original claims as
well as all claims from any and all priority documents, are hereby
incorporated by reference in their entirety into the written
description portion of the specification, and Applicants reserve
the right to physically incorporate into the written description or
any other portion of the application, any and all such claims.
Thus, for example, under no circumstances may the patent be
interpreted as allegedly not providing a written description for a
claim on the assertion that the precise wording of the claim is not
set forth in haec verba in written description portion of the
patent.
[0311] The claims will be interpreted according to law. However,
and notwithstanding the alleged or perceived ease or difficulty of
interpreting any claim or portion thereof, under no circumstances
may any adjustment or amendment of a claim or any portion thereof
during prosecution of the application or applications leading to
this patent be interpreted as having forfeited any right to any and
all equivalents thereof that do not form a part of the prior
art.
[0312] All of the features disclosed in this specification may be
combined in any combination. Thus, unless expressly stated
otherwise, each feature disclosed is only an example of a generic
series of equivalent or similar features.
[0313] It is to be understood that while the invention has been
described in conjunction with the detailed description thereof, the
foregoing description is intended to illustrate and not limit the
scope of the invention, which is defined by the scope of the
appended claims. Thus, from the foregoing, it will be appreciated
that, although specific nonlimiting embodiments of the invention
have been described herein for the purpose of illustration, various
modifications may be made without deviating from the spirit and
scope of the invention. Other aspects, advantages, and
modifications are within the scope of the following claims and the
present invention is not limited except as by the appended
claims.
[0314] The specific methods and compositions described herein are
representative of preferred nonlimiting embodiments and are
exemplary and not intended as limitations on the scope of the
invention. Other objects, aspects, and embodiments will occur to
those skilled in the art upon consideration of this specification,
and are encompassed within the spirit of the invention as defined
by the scope of the claims. It will be readily apparent to one
skilled in the art that varying substitutions and modifications may
be made to the invention disclosed herein without departing from
the scope and spirit of the invention. The invention illustratively
described herein suitably may be practiced in the absence of any
element or elements, or limitation or limitations, which is not
specifically disclosed herein as essential. Thus, for example, in
each instance herein, in nonlimiting embodiments or examples of the
present invention, the terms "comprising", "including",
"containing", etc. are to be read expansively and without
limitation. The methods and processes illustratively described
herein suitably may be practiced in differing orders of steps, and
that they are not necessarily restricted to the orders of steps
indicated herein or in the claims.
[0315] The terms and expressions that have been employed are used
as terms of description and not of limitation, and there is no
intent in the use of such terms and expressions to exclude any
equivalent of the features shown and described or portions thereof,
but it is recognized that various modifications are possible within
the scope of the invention as claimed. Thus, it will be understood
that although the present invention has been specifically disclosed
by various nonlimiting embodiments and/or preferred nonlimiting
embodiments and optional features, any and all modifications and
variations of the concepts herein disclosed that may be resorted to
by those skilled in the art are considered to be within the scope
of this invention as defined by the appended claims.
[0316] The invention has been described broadly and generically
herein. Each of the narrower species and subgeneric groupings
falling within the generic disclosure also form part of the
invention. This includes the generic description of the invention
with a proviso or negative limitation removing any subject matter
from the genus, regardless of whether or not the excised material
is specifically recited herein.
[0317] It is also to be understood that as used herein and in the
appended claims, the singular forms "a," "an," and "the" include
plural reference unless the context clearly dictates otherwise, the
term "X and/or Y" means "X" or "Y" or both "X" and "Y", and the
letter "s" following a noun designates both the plural and singular
forms of that noun. In addition, where features or aspects of the
invention are described in terms of Markush groups, it is intended,
and those skilled in the art will recognize, that the invention
embraces and is also thereby described in terms of any individual
member and any subgroup of members of the Markush group, and
applicants reserve the right to revise the application or claims to
refer specifically to any individual member or any subgroup of
members of the Markush group.
[0318] Other nonlimiting embodiments are within the following
claims. The patent may not be interpreted to be limited to the
specific examples or nonlimiting embodiments or methods
specifically and/or expressly disclosed herein. Under no
circumstances may the patent be interpreted to be limited by any
statement made by any Examiner or any other official or employee of
the Patent and Trademark Office unless such statement is
specifically and without qualification or reservation expressly
adopted in a responsive writing by Applicants.
VII. REFERENCES
[0319] The following references, to the extent that they provide
exemplary procedural or other details supplementary to those set
forth herein, are specifically incorporated herein by reference.
[0320] U.S. Pat. No. 8,992,458 [0321] U.S. Patent Publication
2013/0035660 [0322] U.S. Patent Publication 2013/0035574 [0323]
U.S. Patent Publication 2013/0035560 [0324] I. Mei, Z., et al.,
Tumour-infiltrating inflammation and prognosis in colorectal
cancer: systematic review and meta-analysis. Br J Cancer, 2014.
110(6): p. 1595-605. [0325] II. Salgado, R., et al., Harmonization
of the evaluation of tumor infiltrating lymphocytes (TILs) in
breast cancer: recommendations by an international TILs-working
group 2014. Ann Oncol, 2014 [0326] III. Schatton, T., et al.,
Tumor-infiltrating lymphocytes and their significance in melanoma
prognosis. Methods Mol Biol, 2014. 1102: p. 287-324. [0327] IV.
Gooden, M. J., et al., The prognostic influence of
tumour-infiltrating lymphocytes in cancer: a systematic review with
meta-analysis. Br J Cancer, 2011. 105(1): p. 93-103. [0328] V.
Schreiber, R. D., L. J. Old, and M. J. Smyth, Cancer immunoediting:
integrating immunity's roles in cancer suppression and promotion.
Science, 2011. 331(6024): p. 1565-70. [0329] VI. Bremnes, R. M., et
al., The role of tumor-infiltrating immune cells and chronic
inflammation at the tumor site on cancer development, progression,
and prognosis: emphasis on non-small cell lung cancer. J Thorac
Oncol, 2011. 6(4): p. 824-33. [0330] VII. Talmadge, J. E., Immune
cell infiltration of primary and metastatic lesions: mechanisms and
clinical impact. Semin Cancer Biol, 2011. 21(2): p. 131-8. [0331]
VIII. Shirabe, K., et al., Tumor-infiltrating lymphocytes and
hepatocellular carcinoma: pathology and clinical management. Int J
Clin Oncol, 2010. 15(6): p. 552-8. [0332] IX. Nosho, K., et al.,
Tumour-infiltrating T-cell subsets, molecular changes in colorectal
cancer, and prognosis: cohort study and literature review. J
Pathol, 2010. 222(4): p. 350-66. [0333] X. Bellati, F., et al.,
Immunology of gynecologic neoplasms: analysis of the prognostic
significance of the immune status. Curr Cancer Drug Targets, 2009.
9(4): p. 541-65. [0334] XI. Oble, D. A., et al., Focus on TILs:
prognostic significance of tumor infiltrating lymphocytes in human
melanoma. Cancer Immun, 2009. 9: p. 3. [0335] XII. Uppaluri, R., G.
P. Dunn, and J. S. Lewis, Jr., Focus on TILs: prognostic
significance of tumor infiltrating lymphocytes in head and neck
cancers. Cancer Immun, 2008. 8: p. 16. [0336] XIII. Dunn, G. P., I.
F. Dunn, and W. T. Curry, Focus on TILs: Prognostic significance of
tumor infiltrating lymphocytes in human glioma. Cancer Immun, 2007.
7: p. 12. [0337] XIV. Chang, W. J., et al., Inflammation-related
factors predicting prognosis of gastric cancer. World J
Gastroenterol, 2014. 20(16): p. 4586-96. [0338] X. V. Alonso M M,
Gomez-Manzano C, Bekele B N, Yung W K, Fueyo J. Adenovirus-based
strategies overcome temozolomide resistance by silencing the
O6-methylguanine-DNA methyltransferase promoter. Cancer research
2007; 67:11499-504. [0339] XVI. Alonso M M, Jiang H, Yokoyama T, et
al. Delta-24-RGD in combination with RAD001 induces enhanced
anti-glioma effect viac autophagic cell death. Molecular therapy:
the journal of the American Society of Gene Therapy 2008;
16:487-93. [0340] XVII. Fueyo J, Alemany R, Gomez-Manzano C, et al.
Preclinical characterization of the antiglioma activity of a
tropism-enhanced adenovirus targeted to the retinoblastoma pathway.
Journal of the National Cancer Institute 2003; 95:652-60. [0341]
XVIII. Fueyo J, Gomez-Manzano C, Alemany R, et al. A mutant
oncolytic adenovirus targeting the Rb pathway produces anti-glioma
effect in vivo. Oncogene 2000; 19:2-12. [0342] XIX. Fueyo J,
Gomez-Manzano C, Yung W K. Advances in translational research in
neuro-oncology. Archives of neurology 2011; 68:303-8. [0343] XX.
Wang M, Hemminki A, Siegal G P, et al. Adenoviruses with an RGD-4C
modification of the fiber knob elicit a neutralizing antibody
response but continue to allow enhanced gene delivery. Gynecologic
oncology 2005; 96:341-8. [0344] XXI. Wolchok J D, Hoos A, O'Day S,
et al. Guidelines for the evaluation of immune therapy activity in
solid tumors: immune-related response criteria. Clin Cancer Res.
2009; 15(23):7412-7420. [0345] XXII. Gomez-Manzano C, Fueyo J.
Oncolytic adenoviruses for the treatment of brain tumors. Current
opinion in molecular therapeutics 2010; 12:530-7. [0346] XXIII.
Jiang H, Conrad C, Fueyo J, Gomez-Manzano C, Liu T J. Oncolytic
adenoviruses for malignant glioma therapy. Frontiers in bioscience:
a journal and virtual library 2003; 8:d577-88. [0347] XXIV. Bilbao
G, Contreras J L, Dmitriev I, et al. Genetically modified
adenovirus vector containing an RGD peptide in the HI loop of the
fiber knob improves gene transfer to nonhuman primate isolated
pancreatic islets. American journal of transplantation: official
journal of the American Society of Transplantation and the American
Society of Transplant Surgeons 2002; 2:237-43. [0348] XXV. Poole,
R. M., Pembrolizumab: First Global Approval. Drugs, 2014. [0349]
XXVI. Disis, M. L., Immune regulation of cancer. J Clin Oncol,
2010. 28(29): p. 4531-8. [0350] XXVII. Preston, C. C., et al., The
ratios of CD8+ T cells to CD4+CD25+FOXP3+ and FOXP3- T cells
correlate with poor clinical outcome in human serous ovarian
cancer. PLoS One, 2013. 8(11): p. e80063. [0351] XXVIII. Yoon, H.
H., et al., Prognostic impact of FoxP3+ regulatory T cells in
relation to CD8+T lymphocyte density in human colon carcinomas.
PLoS One, 2012. 7(8): p. e42274. [0352] XXIX. Kim, S. T., et al.,
Tumor-infiltrating lymphocytes, tumor characteristics, and
recurrence in patients with early breast cancer. Am J Clin Oncol,
2013. 36(3): p. 224-31. [0353] XXX. Mathai, A. M., et al., Role of
Foxp3-positive tumor-infiltrating lymphocytes in the histologic
features and clinical outcomes of hepatocellular carcinoma. Am J
Surg Pathol, 2012. 36(7): p. 980-6. [0354] XXXI. Liu, F., et al.,
CD8(+) cytotoxic T cell and FOXP3(+) regulatory T cell infiltration
in relation to breast cancer survival and molecular subtypes.
Breast Cancer Res Treat, 2011. 130(2): p. 645-55. [0355] XXXII.
Kirk, R., Risk factors. CD8+:FOXP3+ cell ratio is a novel survival
marker for colorectal cancer. Nat Rev Clin Oncol, 2010. 7(6): p.
299. [0356] XXXIII. Pedoeem, A., et al., Programmed death-1 pathway
in cancer and autoimmunity. Clin Immunol, 2014. 153(1): p. 145-52.
[0357] XXXIV. Zhang, X., et al., Structural and functional analysis
of the costimulatory receptor programmed death-1. Immunity, 2004.
20(3): p. 337-47. [0358] XXXV. Lazar-Molnar, E., et al., Crystal
structure of the complex between programmed death-1 (PD-1) and its
ligand PD-L2. Proc Natl Acad Sci USA, 2008. 105(30): p. 10483-8.
[0359] XXXVI. Lin, D. Y., et al., The PD-1/PD-L1 complex resembles
the antigen-binding Fv domains of antibodies and T cell receptors.
Proc Natl Acad Sci USA, 2008. 105(8): p. 3011-6. [0360] XXXVII.
Cheng, X., et al., Structure and interactions of the human
programmed cell death 1 receptor. J Biol Chem, 2013. 288(17): p.
11771-85. [0361] XXXVIII. Sheppard, K. A., et al., PD-1 inhibits
T-cell receptor induced phosphorylation of the ZAP70/CD3zeta
signalosome and downstream signaling to PKCtheta. FEBS Lett, 2004.
574(1-3): p. 37-41. [0362] XXXIX. Ott, P. A., F. S. Hodi, and C.
Robert, CTLA-4 and PD-1/PD-L1 blockade: new immunotherapeutic
modalities with durable clinical benefit in melanoma patients. Clin
Cancer Res, 2013. 19(19): p. 5300-9. [0363] XL. Yao, S. and L.
Chen, PD-1 as an immune modulatory receptor. Cancer J, 2014. 20(4):
p. 26 [0364] XLI. Nishimura, H., et al., Developmentally regulated
expression of the PD-1 protein on the surface of double-negative
(CD4-CD8-) thymocytes. Int Immunol, 1996. 8(5): p. 773-80. [0365]
XLII. Huang, X., et al., PD-1 expression by macrophages plays a
pathologic role in altering microbial clearance and the innate
inflammatory response to sepsis. Proc Natl Acad Sci USA, 2009.
106(15): p. 6303-8. [0366] XLIII. Pena-Cruz, V., et al., PD-1 on
immature and PD-1 ligands on migratory human Langerhans cells
regulate antigen-presenting cell activity. J Invest Dermatol, 2010.
130(9): p. 2222-30. [0367] XLIV. Keir, M. E., et al., PD-1 and its
ligands in tolerance and immunity Annu Rev Immunol, 2008. 26: p.
677-704. [0368] XLV. Karim, R., et al., Tumor-expressed B7-H1 and
B7-DC in relation to PD-1+ T-cell infiltration and survival of
patients with cervical carcinoma. Clin Cancer Res, 2009. 15(20): p.
6341-7. [0369] XLVI. Taube, J. M., et al., Colocalization of
inflammatory response with B7-hl expression in human melanocytic
lesions supports an adaptive resistance mechanism of immune escape.
Sci Transl Med, 2012. 4(127): p. 127ra37. [0370] XLVII. Sanmamed,
M. F. and L. Chen, Inducible expression of B7-H1 (PD-L1) and its
selective role in tumor site immune modulation. Cancer J, 2014.
20(4): p. 256-61. [0371] XLVIII. Topalian, S. L., C. G. Drake, and
D. M. Pardoll, Targeting the PD-1/B7-H1(PD-L1) pathway to activate
anti-tumor immunity Curr Opin Immunol, 2012. 24(2): p. 207-12.
[0372] XLIX. Hirano, F., et al., Blockade of B7-H1 and PD-1 by
monoclonal antibodies potentiates cancer therapeutic immunity
Cancer Res, 2005. 65(3): p. 1089-96. [0373] L. Blank, C., et al.,
PD-L1/B7H-1 inhibits the effector phase of tumor rejection by T
cell receptor (TCR) transgenic CD8+ T cells. Cancer Res, 2004.
64(3): p. 1140-5. [0374] LI. Weber, J., Immune checkpoint proteins:
a new therapeutic paradigm for cancer--preclinical background:
CTLA-4 and PD-1 blockade. Semin Oncol, 2010. 37(5): p. 430-9.
[0375] LII. Strome, S. E., et al., B7-H1 blockade augments adoptive
T-cell immunotherapy for squamous cell carcinoma. Cancer Res, 2003.
63(19): p. 6501-5. [0376] LIII. Spranger, S., et al., Mechanism of
tumor rejection with doublets of CTLA-4, PD-1/PD-L1, or IDO
blockade involves restored IL-2 production and proliferation of
CD8(+) T cells directly within the tumor microenvironment. J
Immunother Cancer, 2014. 2: p. 3. [0377] LIV. Curran, M. A., et
al., PD-1 and CTLA-4 combination blockade expands infiltrating T
cells and reduces regulatory T and myeloid cells within B16
melanoma tumors. Proc Natl Acad Sci USA, 2010. 107(9): p. 4275-80.
[0378] LV. Pilon-Thomas, S., et al., Blockade of programmed death
ligand 1 enhances the therapeutic efficacy of combination
immunotherapy against melanoma. J Immunol, 2010. 184(7): p. 3442-9.
[0379] LVI. Nomi, T., et al., Clinical significance and therapeutic
potential of the programmed death-1 ligand/programmed death-1
pathway in human pancreatic cancer. Clin Cancer Res, 2007. 13(7):
p. 2151-7. [0380] LVII. Wick W, et al. Phase III Study of
Enzastaurin Compared With Lomustine in the Treatment of Recurrent
Intracranial Glioblastoma. J Clin Oncol 2010; 28:1168-1174. [0381]
LVIII. Lamborn K R, Yung W K A, Chang S M, et al. Progression-free
survival: an important end point in evaluating therapy for
recurrent high-grade gliomas. Neuro-Oncology. 2008; 10(2):
162-170.
* * * * *