U.S. patent application number 17/667684 was filed with the patent office on 2022-08-25 for formulations and methods for the treatment of cancers.
The applicant listed for this patent is Stemirna (Shanghai) Biotechnology Co. LTED.. Invention is credited to Xin CHEN, Dean G. TANG.
Application Number | 20220265610 17/667684 |
Document ID | / |
Family ID | 1000006330197 |
Filed Date | 2022-08-25 |
United States Patent
Application |
20220265610 |
Kind Code |
A1 |
TANG; Dean G. ; et
al. |
August 25, 2022 |
FORMULATIONS AND METHODS FOR THE TREATMENT OF CANCERS
Abstract
The present invention is directed to a formulation for treating
cancer comprising an androgen receptor signaling inhibitor and a
B-cell-lymphoma-2 inhibitor, which may further comprising a
Bromodomain-and-Extra-Terminal protein inhibitor or a
phosphoinositide 3-kinase inhibitor.
Inventors: |
TANG; Dean G.;
(Williamsville, NY) ; CHEN; Xin; (Wuhan,
CN) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Stemirna (Shanghai) Biotechnology Co. LTED. |
Shanghai |
|
CN |
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|
Family ID: |
1000006330197 |
Appl. No.: |
17/667684 |
Filed: |
February 9, 2022 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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16825104 |
Mar 20, 2020 |
11278524 |
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17667684 |
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15888962 |
Feb 5, 2018 |
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16825104 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 31/4709 20130101;
A61K 31/4166 20130101; A61K 31/437 20130101; A61K 31/551 20130101;
A61K 2300/00 20130101; A61P 35/00 20180101; A61K 31/496
20130101 |
International
Class: |
A61K 31/4166 20060101
A61K031/4166; A61P 35/00 20060101 A61P035/00 |
Claims
1. A formulation, comprising: an androgen receptor signaling
inhibitor; and a B-cell lymphoma 2 inhibitor.
2. The formulation of claim 1 wherein the androgen receptor
signaling inhibitor is enzalutamide.
3. The formulation of claim 2, wherein enzalutamide is contained in
said formulation in a concentration of about 5 mg/ml to about 10.0
mg/ml
4. The formulation of claim 1, wherein the B-cell lymphoma 2
inhibitor is venetoclax.
5. The formulation of claim 4, wherein venetoclax is contained in
said formulation in a concentration of about 10 mg/ml to about 15
mg/ml.
6. A pharmaceutical composition comprising the formulation of claim
1 and a pharmaceutically acceptable carrier.
7. The formulation of claim 1, further comprising: a Bromodomain
and Extra-Terminal protein inhibitor.
8. The formulation of claim 7, wherein the Bromodomain and
Extra-Terminal protein inhibitor is JQ1.
9. The formulation of claim 8, wherein JQ1 is contained in said
formulation in a concentration of about 10 mg/ml to about 15
mg/ml.
10. A pharmaceutical composition comprising the formulation of
claim 7 and a pharmaceutically acceptable carrier.
11. The formulation of claim 1, further comprising: a
phosphoinositide 3-kinase inhibitor.
12. The formulation of claim 11, wherein the phosphoinositide
3-kinase inhibitor is NVP-BEZ235.
13. A pharmaceutical composition comprising the formulation of
claim 11 and a pharmaceutically acceptable carrier.
14. A method for inhibiting growth of a tumor in a castration
resistant prostate cancer comprising the step of: contacting cells
in the tumor with a pharmacologically effective amount of the
formulation of claim 1.
15. A method for inhibiting growth of a tumor in a castration
resistant prostate cancer comprising the step of: contacting cells
in the tumor with a pharmacologically effective amount of the
formulation of claim 7.
16. A method for inhibiting growth of a tumor in a castration
resistant prostate cancer comprising the step of: contacting cells
in the tumor with a pharmacologically effective amount of the
formulation of claim 11.
17. A method for treating a prostate cancer in a subject in need of
such treatment, comprising the step of: administering to said
subject a pharmacologically effective amount of an androgen
receptor signaling inhibitor and a B-cell lymphoma 2 inhibitor.
18. The method of claim 17, wherein said inhibitors are
administered sequentially or simultaneously.
19. The method of claim 17, further comprising: administering to
said subject a pharmacologically effective amount of a Bromodomain
and Extra-Terminal protein inhibitor.
20. The method of claim 17, further comprising: administering to
said subject a pharmacologically effective amount of a
phosphoinositide 3-kinase inhibitor.
21. The method of claim 17, wherein said prostate cancer is an
androgen dependent prostate cancer, an androgen independent
prostate cancer, an androgen receptor negative prostate cancer, or
a castration resistant prostate cancer.
22. A method for treating a cancer in a subject in need of such
treatment, comprising the step of: administering either
sequentially or simultaneously to said subject a pharmacologically
effective amount of an androgen receptor signaling inhibitor, a
B-cell lymphoma 2 inhibitor, a Bromodomain and Extra-Terminal
protein inhibitor, and a phosphoinositide 3-kinase inhibitor.
23. The method of claim 22, wherein the androgen receptor signaling
inhibitor is enzalutamide.
24. The method of claim 22, wherein the B-cell lymphoma 2 inhibitor
is venetoclax.
25. The method of claim 22, wherein the Bromodomain and
Extra-Terminal protein inhibitor is JQ1.
26. The method of claim 22, wherein the phosphoinositide 3-kinase
inhibitor is NVP-BEZ235.
27. The method of claim 22, wherein the cancer is a prostate
cancer.
28. The method of claim 22, wherein the prostate cancer is a
castration resistant prostate cancer.
Description
BACKGROUND OF THE INVENTION
Field of the Invention
[0001] The present invention relates generally to methods for the
treatment of cancer. More specifically, the invention relates to
methods of treating cancers, such as, using a combination
therapy.
Description of the Related Art
[0002] Prostate cancer is an androgen-fueled malignancy and the
androgen receptor (AR) represents the prime therapeutic target.
However, androgen deprivation therapies (ADT) including chemical
castration and use of androgen receptor antagonists have had
limited success, since the targeted tumors become refractory to
treatment and progress into castration resistant prostate cancer.
Although many mechanisms have been reported to contribute to
castration resistance, the origins of the castration resistant
prostate cancer cell or the interactions between heterogeneous
subpopulations of prostate cancer cells contributing to castration
resistant prostate cancer remain poorly understood.
[0003] An analysis of >20 normal human prostate epithelial
strains aimed at elucidating the precursor cells to human prostate
cancer determined that primary, normal human prostate cells are AR-
PSA- CD44+ 2 1+ CK5+ CK18+ progenitors that express the stem cell
markers hTERT, p63, CD44, and Bcl-2, and regenerate prostate in
vivo. Significantly, these progenitor cells can be transformed
using a combination of androgen receptors, AKT, and ERG, suggesting
that these cells can function as the cell of origin for human
prostate cancer. On the other hand, lineage-tracing studies in the
mouse prostate indicate that luminal cells function as the
preferred transformation targets.
[0004] Cellular heterogeneity of human prostate cancer has been
examined using cell and xenograft models, and >220 primary human
prostate cancer (HPCa) derived cells and early-generation
xenografts (patient derived xenografts) have been identified. These
studies established that all prostate cancer cells are not
functionally equal. For example, a subpopulation of prostate cancer
cells, operationally termed prostate cancer stem cells (PCSCs),
having stem cell gene expression profiles are endowed with enhanced
tumorigenic and metastatic potential. Furthermore, prostate cancer
cell population that lacks the differentiation marker Prostate
Specific Antigen) (PSA.sup.-/lo), harbors long-term
castration-resistant tumor-propagating properties. Interestingly, a
fraction (5-18%) of these cells undergo asymmetric cell division
(ACD), a fundamental trait in adult stem cells. The PSA.sup.-/lo
PCa cells are further enriched in epigenetic profiles such as
bivalent chromatin domains, another well-established feature of
stem cells. This is important from a clinical perspective since,
PSA.sup.-/lo prostate cancer cells are heterogenous and can
initiate robust tumor regeneration in androgen ablated individuals
and mediate tumor recurrence during persistent castration, thereby
functioning as a cell of origin for castration resistant prostate
cancer.
[0005] Many of the PCSCs reported have low androgen receptor
(AR.sup.-/lo) expression, making them inherently resistant to
androgen deprivation therapies. For instance, the PSA.sup.-/lo PCSC
population is highly enriched in AR.sup.-/lo cells and readily
propagates castration resistant prostate cancer. Still others have
shown that the prostate cancer stem cell phenotype is induced by
STAT3 dependent loss of androgen receptors, implying that,
signaling through STAT3 and receptor tyrosine kinases, RAS/MAPK,
PI3K/PTEN/AKT/mTOR may all regulate prostate cancer and castration
resistance by reprogramming bulk prostate cancer stem cells.
[0006] Immunohistochemical studies (IHC) have revealed that; (1)
androgen receptor and PSA protein expression in prostate cancer is
both heterogeneous and discordant (that is, AR.sup.+PSA.sup.-,
AR.sub.+PSA.sup.+, AR.sup.-PSA.sup.-, and/or AR.sup.-PSA.sup.+
phenotypes); (2) AR.sup.-/lo and/or PSA.sup.-/lo prostate cancer
cells tend to increase in advanced, metastatic and recurrent
tumors; and (3) poorly differentiated prostate cancer often
completely lack PSA prostate cancer cells. These observations are
in agreement with other studies showing that high-grade and
metastatic prostate cancers have an attenuated androgen-signaling
signature, recurrence is associated with low PSA mRNA in prostate
cancer cells, and, discordant androgen receptorand/or PSA
expression manifest into differential sensitivities to ADT and
other therapeutics. Heterogeneity in androgen receptorexpression is
further emphasized by observations that .about.25% of the
castration resistant prostate cancer samples completely lacked
androgen receptor expression. The remainder AR.sup.+ tumor
population had highly heterogeneous androgen receptor expression;
with both AR.sup.+ and AR.sup.-/lo areas either intermingled with
or frequently separated from each other. Among the AR.sup.+
population, androgen receptor expression showed heterogeneity in
subcellular localization classified into, exclusively nuclear
(nuc-AR), exclusively cytoplasmic (cyto-AR), and a combination of
the above (nuc/cyto-AR).
[0007] For example, transitioning of the PCa xenograft models,
LAPC9, LAPC4, LNCaP, and VCaP, from androgen-dependent (AD) to
androgen-independent (AI) states during propagation in castrated
mice resulted in the androgen-independent tumors displaying nuc-AR
(LNCaP), cyto-AR (LAPC4), nuc/cyto-AR (VCaP) and AR.sup.-/lo
(LAPC9) phenotypic patterns, reminiscent of the clinical castration
resistant prostate cancer manifestation. Since different androgen
receptor-expressing patterns are likely to confer differential
sensitivities to endocrine therapy in prostate cancer cells, a
personalized approach that considers androgen receptor-expression
status described above, would be central for successful
treatment.
[0008] Androgen deprivation therapy remains the primary therapeutic
approach in the treatment of prostate cancer. There are currently
two approaches to androgen deprivation therapy--chemical
"castration" using GnRH agonists such as goserelin to block
testicular androgens or anti-androgens such as
4-(3-(4-Cyano-3-(trifluoromethyl)phenyl)-5,5-dimethyl-4-oxo-2-thioxoimida-
zolidin-1-yl)-2-fluoro-N-methylbenzamide (Enzalutamide) to block
androgen receptor functions. Unfortunately, these approaches have
not led to clinically acceptable remission since, many prostate
cancer patients undergoing androgen deprivation therapy eventually
become refractory to the treatment, progressing into advanced
castration resistant prostate cancer, which is insensitive to this
line of treatment. There is hence the need for new, patient
centric, personalized treatment approaches in castration resistant
prostate cancer.
[0009] Overall, there is a deficiency in the art for combinatorial
and/or synergistic approaches to treat cancers. The present
invention fulfills this longstanding need and desire in the
art.
SUMMARY OF THE INVENTION
[0010] The present invention is directed to a formulation for
treating cancer comprising an androgen receptor signaling inhibitor
and a B-cell-lymphoma-2 inhibitor. The present invention is
directed to a related formulation further comprising a
Bromodomain-and-Extra-Terminal protein inhibitor. The present
invention is directed to another related formulation further
comprising a phosphoinositide 3-kinase inhibitor.
[0011] The present invention also is directed to a method for
inhibiting tumor growth in a castration resistant prostate cancer.
The method comprises contacting cells in the tumor with a
pharmacologically effective amount of one of the formulations
described herein.
[0012] The present invention is directed further to a method for
treating a prostate cancer in a subject in need of such treatment.
The method comprises administering to the subject a
pharmacologically effective amount of one of the formulations
described herein.
[0013] The present invention is further directed to a method for
treating a cancer in a subject in need of such treatment. The
method comprises administering either sequentially or
simultaneously to the subject a pharmacologically effective amount
of an androgen receptor signaling inhibitor, a B-cell lymphoma 2
inhibitor, a Bromodomain and Extra-Terminal protein inhibitor, and
a phosphoinositide 3-kinase inhibitor.
[0014] Other and further aspects, features, benefits, and
advantages of the present invention will be apparent from the
following description of the presently preferred embodiments of the
invention given for the purpose of disclosure.
BRIEF DESCRIPTION OF DRAWINGS
[0015] So that the matter in which the above-recited features,
advantages and objects of the invention, as well as others that
will become clear, are attained and can be understood in detail,
more particular descriptions of the invention briefly summarized
above may be had by reference to certain embodiments thereof that
are illustrated in the appended drawings. These drawings form a
part of the specification. It is to be noted, however, that the
appended drawings illustrate preferred embodiments of the invention
and therefore are not to be considered limiting in their scope.
[0016] FIGS. 1A-1C shows androgen receptor heterogeneity in
prostate cancer cells. FIG. 1A shows heterogeneous expression of
androgen receptor in a patient castration resistant prostate cancer
(representative patient #13553). * marks denote androgen
receptor-negative (neg) areas. Inset on top right shows negative
control. Lower panels show 4 patterns of androgen receptor
expression. FIG. 1B shows a schematic for xenograft model used to
generate primary and secondary castration resistant prostate
cancer. FIG. 1C shows androgen receptor immunohistochemistry for
androgen dependent and androgen independent xenograft models. Note
the four androgen receptor staining patterns (nuc, cyto, nuc/cyto,
and neg).
[0017] FIGS. 2A-2B shows western blot analysis of molecular changes
in castration resistant prostate cancer induced by surgical
castration. FIG. 2A shows western blotting analysis of whole cell
lysates from the LAPC9 AI (PC9) and LAPC4 (PC4) models. LNCaP cells
were used as control for androgen receptor and PSA FIG. 2B shows
western blotting analysis of whole cell lysates from VCaP and LNCaP
models. LAPC9 AD tumor was run for comparison. Asterisk denotes,
non-specific bands.
[0018] FIGS. 3A-3F show molecular changes in secondary LNCaP
castration resistant prostate cancer in response to Enzalutamide.
FIG. 3A shows the experimental timeline. FIG. 3B shows treatment
response of the androgen-independent LNCaP tumors to Enzalutamide.
The arrow indicates the time when Enzalutamide-resistant secondary
castration resistant prostate cancer began to emerge. FIG. 3C shows
WB analysis of the molecules indicated in 3-4 representative tumors
treated with vehicle (corn oil) or Enzalutamide (both .beta.-actin
and GAPDH were used as loading controls). Asterisk denotes the
androgen receptor and GR splice variants. FIG. 3D shows
representative IHC images of androgen receptor, PSA, and GR in one
each vehicle- and Enzalutamide-treated endpoint tumors. FIG. 3E
shows timeline for a pilot secondary castration resistant prostate
cancer treatment. FIG. 3F shows the endpoint tumor images with
tumor incidence and tumor weights indicated on the right. The two
small tumors in the rectangle box were harvested two weeks later
than the rest.
[0019] FIGS. 4A-4G shows molecular changes in castration- and
Enzalutamide-resistant LAPC9 tumors. FIG. 4A shows the experimental
timeline. FIG. 4B shows treatment response of the primary
androgen-independent LAPC9 tumors to Enzalutamide. FIG. 4C shows
western blot analysis of the molecules indicated in 4
representative tumors treated with vehicle (corn oil) or
Enzalutamide (both .beta.-actin and GAPDH were used as loading
controls). FIG. 4D shows Representative IHC images of androgen
receptor, PSA, and GR in AD, vehicle- and Enzalutamide-treated
endpoint tumors. FIG. 4E shows the timeline for a pilot secondary
castration resistant prostate cancer treatment. FIG. 4F shows JQ1
and JQ1+.alpha.2.beta.1 inhibitor inhibited the LAPC9 AI tumor
growth. FIG. 4G shows endpoint tumor images with tumor incidence
and mean tumor weights indicated on the right.
[0020] FIGS. 5A-SE shows molecular changes in castration- &
Enzalutamide-resistant LAPC4 tumors. FIG. 5A shows subcellular
fractionation shows mainly cytosolic localization of androgen
receptors in LAPC4 androgen-independent tumors (compare lanes 1 and
2) and cytosolic androgen receptors can be immunoprecipitated down
by an anti-AR N-terminus Ab. Bracket and * indicate low M.W species
AR variants. Cyto, cytosol; NE, nuclear extract. FIG. 5B shows
timeline of therapeutic experiments. FIG. SC shows treatment
response of the LAPC4 primary androgen-independent tumors to
Enzalutamide. FIG. 5D shows representative IHC images of androgen
receptor and PSA in AD, vehicle- and Enzalutamide-treated endpoint
tumors. FIG. 5E shows western blot analysis of the molecules
indicated in 4 representative tumors (each) treated with vehicle
(corn oil) or Enzalutamide (both .beta.-actin and GAPDH were used
as loading controls).
DETAILED DESCRIPTION OF THE INVENTION
[0021] As used herein in 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.
[0022] As used herein "another" or "other" may mean at least a
second or more of the same or different claim element or components
thereof. Similarly, the word "or" is intended to include "and"
unless the context clearly indicates otherwise. "Comprise" means
"include."
[0023] As used herein, the term "about" refers to a numeric value,
including, for example, whole numbers, fractions, and percentages,
whether or not explicitly indicated. The term "about" generally
refers to a range of numerical values (e.g., +/-5-10% of the
recited value) that one of ordinary skill in the art would consider
equivalent to the recited value (e.g., having the same function or
result). In some instances, the term "about" may include numerical
values that are rounded to the nearest significant figure.
[0024] As used herein, the term "contacting" refers to any suitable
method of bringing the formulation described herein or one or more
of its components into contact with a tumor or a cell comprising
the same. In vitro or ex vivo this is achieved by exposing the
comprising the same to the formulation or components thereof in a
suitable medium. For in vivo applications, any known method of
administration is suitable as described herein.
[0025] In one embodiment of the present invention, there is
provided a formulation comprising, an androgen receptor signaling
inhibitor and a B-cell-lymphoma-2 inhibitor. Androgen receptor
signaling inhibitors may include, but are not limited to,
enzalutamide, or other equivalent inhibitors, drugs or agents. In
one aspect of this embodiment, the androgen receptor signaling
inhibitor is enzalutamide contained in the formulation in an amount
of about 5 mg/ml to about 10 mg/ml. B-cell-lymphoma-2 inhibitors
may include, but are not limited to, venetoclax or other such
inhibitors, drugs or agents. In one aspect of this embodiment, the
B-cell-lymphoma-2 inhibitor is venetoclax contained in the
formulation in an amount of about 10 mg/ml to about 15 mg/ml.
[0026] This embodiment also provides a formulation comprising, an
androgen receptor signaling inhibitor, a B-cell-lymphoma-2
inhibitor, and a Bromodomain-and-Extra-Terminal protein inhibitor.
Possible androgen receptor signaling inhibitors and
B-cell-lymphoma-2 inhibitors are as described supra.
Bromodomain-and-Extra-Terminal protein inhibitors may include, but
are not limited to, JQ1, or other equivalent protein inhibitors,
drugs or agents. In one aspect of this embodiment, enzalutamide and
venetoclax may be contained at concentrations described supra, and
JQ1 may be contained in the formulation in an amount of about 10
mg/ml to about 15 mg/ml.
[0027] This embodiment further provides a formulation comprising,
an androgen receptor signaling inhibitor, a B-cell-lymphoma-2
inhibitor, and a phosphoinositide 3-kinase inhibitor. Possible
androgen receptor signaling inhibitors and B-cell-lymphoma-2
inhibitors are as described supra. Phosphoinositide 3-kinase
inhibitors may include, but are not limited to, NVP-BEZ235, or
other equivalent inhibitors, drugs or agents. In one aspect of this
embodiment, enzalutamide, venetoclax and NVP-BEZ235 may be
contained at a concentration described supra.
[0028] This embodiment provides yet another formulation comprising,
an androgen receptor signaling inhibitor, a B-cell-lymphoma-2
inhibitor, a Bromodomain-and-Extra-Terminal protein inhibitor and a
phosphoinositide 3-kinase inhibitor. Representative examples of
inhibitors from each class that may comprise this formulation is
described supra. It is well within the skill of an artisan to
determine the dosage of each inhibitor in the formulation.
[0029] In this embodiment further provided is a pharmaceutical
composition comprising the formulation described supra and a
physiologically acceptable carrier and/or excipient. Representative
examples of pharmaceutical carriers include but are not limited to
oil, suspension, spray, solution, nanoparticle, liposome,
microcapsule, delivery device, or powder as known in the art. The
pharmaceutical composition may be administered via oral or
parenteral routes, such as subcutaneous, intravenous,
intraperitoneal, intradermal, intranasal, or intramuscular
routes.
[0030] The pharmaceutical composition may be administered one or
more times to achieve a therapeutic effect. It is well within the
skill of an artisan to determine dosage or whether a suitable
dosage comprises a single administered dose or multiple
administered doses. An appropriate dosage depends on the subject's
health, the progression or remission of the disease, the route of
administration and the formulation used.
[0031] In another embodiment there are provided methods for
inhibiting growth of a castration resistant prostate cancer. In one
aspect, the method comprises the step of contacting cells in the
tumor with a pharmacologically effective amount of the formulation
of an androgen receptor signaling inhibitor and a B-cell lymphoma 2
inhibitor. In another aspect of this embodiment, the method
comprises the step of contacting cells in the tumor with a
pharmacologically effective amount of the formulation of an
androgen receptor signaling inhibitor, a B-cell lymphoma 2
inhibitor and a Bromodomain and Extra-Terminal inhibitor. In yet
another aspect of this embodiment, the method comprises the step of
contacting cells in the tumor with a pharmacologically effective
amount of the formulation of an androgen receptor signaling
inhibitor, a B-cell lymphoma 2 inhibitor and a phosphoinositide
3-kinase inhibitor. In yet another aspect of this embodiment, the
method comprises the step of contacting cells in the tumor with a
pharmacologically effective amount of the formulation of an
androgen receptor signaling inhibitor, a B-cell lymphoma 2
inhibitor, a Bromodomain and Extra-Terminal inhibitor and a
phosphoinositide 3-kinase inhibitor. Representative examples of
inhibitors from each class are described supra. It is further well
within the skill of an artisan to determine the dosage of each
inhibitor in the formulation.
[0032] In yet another embodiment there are provided methods for
treating a prostate cancer in a subject in need of such treatment,
comprising the step of administering to the subject a
therapeutically effective amount of an androgen receptor signaling
inhibitor and a B-cell-lymphoma-2 inhibitor.
[0033] This embodiment also provides a method for treating a
prostate cancer in a subject in need of such treatment, comprising
the step of administering to the subject a therapeutically
effective amount of an androgen receptor signaling inhibitor, a
B-cell-lymphoma-2 inhibitor and a Bromodomain-and-Extra-Terminal
protein inhibitor.
[0034] This embodiment further provides a method for treating a
prostate cancer in a subject in need of such treatment, comprising
the step of administering to the subject a therapeutically
effective amount of an androgen receptor signaling inhibitor, a
B-cell-lymphoma-2 inhibitor and a phosphoinositide 3-kinase
inhibitor.
[0035] This embodiment provides further still a method for treating
a prostate cancer in a subject in need of such treatment,
comprising the step of administering to the subject a
therapeutically effective amount of an androgen receptor signaling
inhibitor, a B-cell-lymphoma-2 inhibitor, a
Bromodomain-and-Extra-Terminal protein inhibitor and a
phosphoinositide 3-kinase inhibitor.
[0036] In one aspect of this embodiment, the prostate cancer is
androgen dependent prostate cancer. In another aspect of this
embodiment, the cancer is androgen independent prostate cancer. In
yet another aspect of this embodiment, the cancer is androgen
receptor negative prostate cancer. In yet another aspect of this
embodiment, the cancer is castration resistant prostate cancer.
[0037] In yet another embodiment there is provided a method for
treating a cancer in a subject in need of such treatment comprising
the step of administering either sequentially or simultaneously to
the subject a pharmacologically effective amount of an androgen
receptor signaling inhibitor, a B-cell lymphoma 2 inhibitor, a
Bromodomain and Extra-Terminal protein inhibitor, and a
phosphoinositide 3-kinase inhibitor.
[0038] In this embodiment the androgen receptor signaling inhibitor
is enzalutamide. Also in this embodiment the B-cell lymphoma 2
inhibitor is venetoclax. In addition in this embodiment the
Bromodomain and Extra-Terminal protein inhibitor is JQ1. Further in
this embodiment the phosphoinositide 3-kinase inhibitor is
NVP-BEZ235.
[0039] In this embodment the cancer is a prostate cancer, such as,
but not limited to, a castration resistant prostate cancer. Dosages
or concentrations of these inhibitors may be as described supra or
may be other amounts as readily determined by one of ordinary skill
in the art.
[0040] A person having ordinary skill in this art would readily be
able to determine the appropriate dosage for the androgen receptor
signaling inhibitors, the B-cell-lymphoma-2 inhibitors,
Bromodomain-and-Extra-Terminal protein inhibitors and the
phosphoinositide 3-kinase inhibitors in the formulation described
supra, useful to treat the indicated prostate cancer. A person
having ordinary skill in this art would also be capable of
determining the most effective routes of administration, which
include but are not limited to, oral administration and intravenous
administration. A person having ordinary skill in this art would
further be able to determine the frequency of administration, which
may be adjusted from once a week, to several times a week to every
week day depending on the cancer being treated.
[0041] Thus, provided herein are formulations of inhibitors and
other compounds and pharmaceutical compositions thereof effective
for treating a cancer or inhibiting growth of cells or tumors
associated with the prostate cancer. Representative compounds
comprising the formulation may be:
TABLE-US-00001 Compound Chemical name Enzalutamide
-(3-(4-Cyano-3-(trifluoromethyl)phenyl)-5,5-dimethyl-4-oxo-2-thioxo
imidazolidin-1-yl)-2-fluoro-N-methylbenzamide Venetoclax
4-(4-{[2-(4-Chlorophenyl)-4,4-dimethyl-1-cyclohexen-1-yl]methyl-
}-1-
piperazinyl)-N-({3-nitro-4-[(tetrahydro-2H-pyran-4-ylmethyl)amino]phenyl}
sulfonyl)-2-(1H-pyrrolo[2,3-b]pyridin-5-yloxy) benzamide JQ1
(S)-tert-butyl
2-(4-(4-chlorophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-
- a][1,4]diazepin-6-yl)acetate NVP-BEZ235
2-Methyl-2-{4-[3-methyl-2-oxo-8-(quinolin-3-yl)-2,3-dihydro-1H--
imidazo [4,5-c]quinolin-1-yl]phenyl}propanenitrile
[0042] As is well known in the art, the methods of the present
invention may be administered to either human or non-human
subjects. For example, administration may be oral or parenteral. As
is well known in the art, the methods of the present invention may
be administered alone or in combination with one or more other
commonly used cancer chemotherapeutic agents to a subject to treat
a particular condition.
[0043] The following examples are given for the purpose of
illustrating various embodiments of the invention and are not meant
to limit the present invention in any fashion.
EXAMPLE 1
Patterns of AR Expression in Clinical Castration-Resistant Prostate
Cancer and Castration-Resistant (CR) Xenografts
[0044] About 100 castration resistant prostate cancer whole-mount
(WM) or tissue microarray (TMA) slides were assayed for androgen
receptor expression using two antibodies that recognize the
N-terminal epitopes of the androgen receptor, which could detect
both full-length androgen receptor and all c-terminal truncated
androgen receptor variants. The results revealed interesting and
striking heterogeneous patterns of androgen receptor expression. Of
the tested clinical samples, 26 cases (25%) completely lacked
androgen receptor expression. The remaining AR.sup.+ tumors showed
highly heterogeneous androgen receptor expression with both
AR.sup.+ and AR.sup.-/lo areas either inter-mingled with or
frequently separated from each other (FIG. 1A). When `zoomed` in,
the androgen receptor-positive PCa cells showed 3 expression
patterns: substantially nuclear (nuc-AR), substantially cytoplasmic
(cyto-AR), and both nuclear and cytoplasmic (nuc/cyto-AR) (FIG. 1A,
lower panel). When the PCa xenograft models--LAPC9, LAPC4, LNCaP,
and VCaP, transitioned from androgen-dependent (AD) to
androgen-independent (AI) state during propagation in castrated
mice (FIG. 1B), the androgen-independent tumors displayed the 4 AR
patterns resembling those observed in clinical castration resistant
prostate cancer discussed above. Specifically, the LNCaP
androgen-independent tumors showed substantially nuc-AR, LAPC4
showed substantially cyto-AR, VCaP showed nuc/cyto-AR, and LAPC9
showed substantially AR.sup.-/lo phenotypes (FIG. 1C).
[0045] Molecular Changes in Castration Resistant Prostate Cancer
Induced by Surgical Castration
[0046] The xenograft models--LAPC9, LAPC4, LNCaP, and VCaP were
serially passaged in castrated male NOD/SCID--.gamma. mice (NSG;
for LNCaP or NOD/SCID for the other models). The tumors were then
harvested for western blot analysis for androgen receptor, androgen
receptor-target molecules, cancer stem cell (CSC) and castration
resistance (CR) markers. PC3 cells were used as a negative control
for androgen receptors. 60 .mu.g of whole cell lysate was loaded in
each lane. GADPH was run as the loading controls (top and bottom
panels). FIG. 2A shows that castration led to a significant
reduction of androgen receptors and its two targets PSA and FKBPS
in LAPC9 androgen-independent tumors. On the other hand, the LAPC9
primary androgen-independent tumors showed significant upregulation
of 5 molecules/pathways: N-cadherin, Bcl-2, integrin .alpha.2,
c-Myc, and p-ERK1/2. In contrast, markers, GR, pAkt, ALDH7A1 (57),
and p-Stat3 did not show any difference between androgen-dependent
and androgen-independent tumors (FIG. 2A).
[0047] This enrichment of AR.sup.-/lo/PSA.sup.-/lo PCa cells in
LAPC9 androgen-independent suggests that they can function as the
cell-of-origin for castration resistant prostate cancer.
[0048] In the LNCaP model, castration led to increased androgen
receptor expression (FIG. 2B), which was localized mostly in the
nuclei (see FIG. 1C). The 3 androgen receptor target molecules,
PSA, FKBPS, and NKX3.1, were expressed and no androgen receptor
splice variants including AR-V7 were observed (FIG. 2B). Among the
CSC and CR markers, N-cadherin was not detected; GR (glucocorticoid
receptor) showed only transient and minimal changes; and c-Myc,
E-cadherin, and p-AKT decreased slightly (FIG. 2B). In contrast,
integrin .alpha.2 (CSC marker) and pERK1/2 showed initial increase
followed by decline, whereas Bcl-2 and pStat3 demonstrated
sustained increases (FIG. 2B).
EXAMPLE 2
Molecular Changes in Secondary LNCaP Castration Resistant Prostate
Cancer in Response to Enzalutamide Therapy
[0049] To test the efficacy of enzalutamide in reducing
androgen-independent PCa tumor burden, therapy experiments were
performed by treating mice bearing LNCaP AI-tumors with
enzalutamide, administered by i.p. route. FIG. 4B revealed that
enzalutamide suppressed growth of androgen-independent LNCaP for
the first 6.5 weeks, suggesting that the upregulated nuc-AR (see
FIG. 1C) is causally mediating the primary castration resistant
prostate cancer in the LNCaP androgen-independent-tumor model.
However, at about 7 weeks no further response to enzalutamide was
observed (arrow, FIG. 4B) suggesting the emergence of
enzalutamide-resistant tumors. These secondary castration resistant
prostate cancer tumors were resistant to both surgical castration
and enzalutamide as evidenced by western blotting analysis (FIG.
4C) and immune-histochemical analysis (FIG. 4D) for markers,
androgen receptor, GR, including. GR splice variants (FIG. 4C).
Among the 3 androgen receptor targets, PSA (FIG. 4C-4D) and FKBP5
(FIG. 4C) decreased, suggesting that in enzalutamide resistant
secondary LNCaP CR tumors, androgen receptor binding to the genome
is shifting away from conventional targets. The observed decrease
in PSA is consistent with earlier observations that persistent
castration leads to an enrichment in phenotypically
undifferentiated PSA.sup.-lo PCa cells.
[0050] Molecular changes in secondary LNCaP castration resistant
prostate cancer in response to Enzalutamide/Venetoclax combination
therapy
[0051] Among the CSCs and castration resistance markers examined,
Bcl-2 showed the most prominent upregulation in response to
enzalutamide (FIG. 4C). To determine if therapeutic targeting of
Bcl-2 in combination with Enzalutamide a targeted therapy
experiment was performed by treating mice bearing LNCaP primary
androgen-independent tumors simultaneously with enzalutamide and a
Bcl-2 antagonist, venetoclax (FIG. 4E). The results shown in FIG.
4F demonstrate a clear and striking suppression of emergence
(incidence) of LNCaP secondary castration resistant prostate cancer
(P<0.0001, .chi..sup.2 test) in animals administered
Enzalutamide+Venetoclax combination therapy. In contrast, control
combination experiments in which venetoclax was substituted with
the GR antagonist, mifepristone (RU486), inhibited only inhibited
tumor growth without reducing incidence (FIG. 4F).
EXAMPLE 3
Castration of LAPC9 AD Tumors Leads to Decreased AR and
Upregulation of Many CSC and Castration-Associated Molecules and
Pathways
[0052] To determine the sensitivity of LAPC9 androgen-independent
tumors to enzalutamide, therapy experiments were performed as
described for the LNCaP primary AI tumors (FIG. 3). FIG. 4 shows
that in sharp contrast to the observed reductions in tumor volume
following Enzalutamide therapy, LNCaP primary androgen-independent
tumors were refractory to enzalutamide administration (FIG. 4B).
Western blotting (FIG. 4C) and immuno-histochemical (FIG. 4D)
analysis revealed no expression of androgen receptors and PSA.
FKBPS levels were greatly diminished in both vehicle and treatment
groups. On the other hand Enzalutamide treatment continued to
express high levels of integrin .alpha.2, c-Myc, N-cadherin, Bcl-2,
and p-ERK1/2 (FIG. 4C). To determine whether blocking alternate
pathways regulated by any of these high expressing proteins, in the
Enzalutamide resistant LAPC9 androgen-independent tumors, affects
tumor burden, therapy experiments were performed with the
Bromodomain and Extra-Terminal protein (a upstream regulator of Myc
transcription) inhibitor, JQ1. The integrin .alpha.2.beta.1
inhibitor, Compound 15 (142,143) was used as a positive control.
FIGS. 4F-4G shows that JQ1 significantly inhibited tumor growth but
not tumor incidence (compare FIG. 4F with FIG. 3F). These studies
show that Myc in the AR.sup.-/lo LAPC9 androgen-independent tumors
is causally important for tumor growth under androgen-independent
conditions, and further suggest the likely benefits of making a
combinatorial formulation comprising Enzalutamide and JQ1 in the
treatment of androgen-independent prostate cancer and castration
resistant prostate cancer.
EXAMPLE 4
Response to Enzalutamide Treatment in LAPC4 AI Tumors Having
Cytoplasmic AR Localization
[0053] As shown in FIG. 1C, immuno-histochemistry analysis revealed
significant cytoplasmic localization of ARs in LAPC4
androgen-independent tumors. This is further confirmed by
subcellular fractionation followed by western blot analysis (FIG.
5A), which shows that a majority of androgen receptor was in the
cytosol, with only a minor signal being attributed to the nuclear
fraction (FIG. 5A lanes 1 vs. 2). To determine whether targeting
cyt-AR in these tumors would be effective in reducing tumor burden,
castrated mice bearing LAPC4 androgen-independent tumors were
administered enzalutamide as indicated (FIG. 5B), and the tumor
volume measured.
[0054] FIG. 5C shows that LAPC4 androgen-independent tumors in the
enzalutamide treatment group grew significantly slower in the first
2-3 weeks compared to control groups that were administered only
the vehicle, corn oil (FIG. 5C). However, starting from
.about.6.5-7 weeks, tumors resumed growth, indicative of resistance
to enzalutamide. Interestingly, unlike the vigorous growth pattern
of enzalutamide-resistant nuc-AR.sup.+ LNCaP tumors (FIG. 3C), the
enzalutamide-resistant cyto-AR.sup.+ LAPC4 tumors grew relatively
slow even up to 13 weeks (FIG. 5C). This slow growth was not due to
relocation of androgen receptor to the nucleus, or resurgence in AD
nu-AR.sup.+ tumors since immuno-histochemistry showed that androgen
receptor was still largely in the cytoplasm (FIG. 5D). Western blot
analysis revealed that the enzalutamide-resistant LAPC4 tumors
continued to express high levels of GR, N-cadherin, Bcl-2, and
integrin .alpha.2, with slight increases in E-cadherin expression
FIG. 5E). This opens the possibility of employing combinatorial
approaches that target androgen receptor (Enzalutamide) together
molecules such as Bcl2 (Venetoclax).
[0055] The present invention is well adapted to attain the ends and
advantages mentioned as well as those that are inherent therein.
The particular embodiments disclosed above are illustrative only,
as the present invention may be modified and practiced in different
but equivalent manners apparent to those skilled in the art having
the benefit of the teachings herein. Furthermore, no limitations
are intended to the details of construction or design herein shown,
other than as described in the claims below. It is therefore
evident that the particular illustrative embodiments disclosed
above may be altered or modified and all such variations are
considered within the scope and spirit of the present invention.
Also, the terms in the claims have their plain, ordinary meaning
unless otherwise explicitly and clearly defined by the
patentee.
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