U.S. patent application number 14/477512 was filed with the patent office on 2015-04-30 for method for detecting and controlling cancer.
The applicant listed for this patent is George Mason Research Foundation, Inc., Istituto Superiore di Sanita. Invention is credited to Valerie Calvert, Lance A. Liotta, Emanuel F. Petricoin, III, Mariaelena Pierobon.
Application Number | 20150118246 14/477512 |
Document ID | / |
Family ID | 39365024 |
Filed Date | 2015-04-30 |
United States Patent
Application |
20150118246 |
Kind Code |
A1 |
Petricoin, III; Emanuel F. ;
et al. |
April 30, 2015 |
METHOD FOR DETECTING AND CONTROLLING CANCER
Abstract
Disclosed herein are: 1) methods for determining the appropriate
drug therapy for a patient with colorectal cancer that has
metastasized to the patient's liver and/or lung; 2) methods for
treating such a patient; and 3) pharmaceutical compositions for
such treatment.
Inventors: |
Petricoin, III; Emanuel F.;
(Gainesville, VA) ; Pierobon; Mariaelena;
(Fairfax, VA) ; Calvert; Valerie; (Arlington,
VA) ; Liotta; Lance A.; (Bethesda, MD) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
George Mason Research Foundation, Inc.
Istituto Superiore di Sanita |
Fairfax
Roma |
VA |
US
IT |
|
|
Family ID: |
39365024 |
Appl. No.: |
14/477512 |
Filed: |
September 4, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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12513040 |
Apr 30, 2009 |
8834873 |
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PCT/US2007/022863 |
Oct 31, 2007 |
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14477512 |
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60855751 |
Nov 1, 2006 |
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Current U.S.
Class: |
424/142.1 ;
506/9 |
Current CPC
Class: |
A61K 31/506 20130101;
C07K 16/22 20130101; A61K 39/39558 20130101; G01N 33/57438
20130101; A61P 43/00 20180101; G01N 33/57419 20130101; A61P 35/00
20180101; A61P 35/04 20180101; G01N 2800/52 20130101 |
Class at
Publication: |
424/142.1 ;
506/9 |
International
Class: |
G01N 33/574 20060101
G01N033/574; A61K 39/395 20060101 A61K039/395; A61K 31/506 20060101
A61K031/506 |
Claims
1-30. (canceled)
31. A method for determining the appropriate drug therapy for a
patient with colorectal cancer that has metastasized to the
patient's liver and/or lung comprising the steps of: (a) analyzing
a sample of the metastatic cancer from the patient to determine if
two or more of the following proteins are elevated: pAKT, pSTAT3,
pVEGFR, pFKHR-FKHRL1, pcKit, pShc, pSMAD2, pSrc, pSTAT5, pAbl,
pJak1, pPyk2, pPDGFRb, pPRAS40, and pEGFR; and (b) selecting one or
more drugs for treating the patient that target at least two of any
of the proteins found to be elevated, wherein the drugs are
selected from the group consisting of Imatinib mesylate,
Bevacizumab, Erlotinib hydrochloride, Panitumumab, Lapatinib, an
akt inhibitor, and a PI3 kinase inhibitor.
32. The method of claim 31, wherein the step of analyzing the
sample comprises: isolating epithelial cells from the sample;
lysing the epithelial cells; and analyzing the lysate.
33. The method of claim 32, wherein the cells are isolated by laser
capture microdissection and the lysate is distributed onto a
reverse phase microarray and then analyzed by an immunoassay.
34. The method of claim 31, wherein the proteins determined to be
elevated are at least two of pAKT, pAbl, pcKit, pPDGFRb, pVEGFR,
and pEGFR.
35. The method of claim 34, wherein the selected drug is Imatinib
mesylate if pAbl, pcKit, and/or pPDGFRb are elevated, Bevacizumab
if pVEGFR is elevated, and Erlotinib hydrochloride, and/or
Lapatinib if pEGFR is elevated.
36. The method of claim 31, wherein the protein or proteins
determined to be elevated are at least two of pAbl, pcKit, and
pPDGFRb.
37. The method of claim 36, wherein the selected drug is Imatinib
mesylate.
38. The method of claim 31, wherein the proteins are pAKT S473,
pSTAT3 S727, pVEGFR Y951 and Y996, pFKHR-FKHRL1 T24-T32, pcKit
Y703, pShc Y317, pSMAD2 S465-467, pSrc Y527, pSTAT5 Y694, pAbl
Y245, pJak1 Y1022-1023, pPyk2 Y402, pPDGFRb Y751, pPRAS40, and
pEGFR Y992 and Y1173.
39. A method for determining the appropriate drug therapy for a
patient with colorectal cancer that has metastasized to the
patient's liver and/or lung comprising the steps of: (a) analyzing
a sample of the metastatic cancer from the patient to determine if
two or more of the following proteins are elevated: pAKT S473,
pSTAT3 S727, pVEGFR Y951 and Y996, pFKHR-FKHRL1 T24-T32, pcKit
Y703, pShc Y317, pSMAD2 S465-467, pSrc Y527, pSTAT5 Y694, pAbl
Y245, pJak1 Y1022-1023, pPyk2 Y402, pPDGFRb Y751, pPRAS40, and
pEGFR Y992 and Y1173; and (b) selecting one or more drugs for
treating the patient that target at least two of any of the
proteins found to be elevated, wherein the drug is selected from
the group consisting of Imatinib mesylate, Bevacizumab, Erlotinib
hydrochloride, Panitumumab, Lapatinib, and akt inhibitor, and a PI3
kinase inhibitor.
40. The method of claim 39, wherein the proteins determined to be
elevated are at least two of pAKT S473, pAbl Y245, pcKit Y703,
pPDGFRb Y751, pVEGFR Y951 and Y996, and pEGFR Y992 and Y1173.
41. The method of claim 40, wherein the selected drug is Imatinib
mesylate if pAbl Y245, pcKit Y703, and/or pPDGFRb Y751 are
elevated, Bevacizumab if pVEGFR Y951 and/or Y996 is elevated, and
Erlotinib hydrochloride and/or Lapatinib if pEGFR Y992 and/or Y1173
is elevated.
42. The method of claim 40, wherein the protein or proteins
determined to be elevated are at least two of pAbl Y245, pcKit
Y703, and pPDGFRb Y751 and the selected drug is Imatinib
mesylate.
43. The method of claim 39, wherein the step of analyzing the
sample comprises: isolating epithelial cells from the sample;
lysing the epithelial cells; and analyzing the lysate.
44. The method of claim 43, wherein the cells are isolated by laser
capture microdissection and the lysate is distributed onto a
reverse phase microarray and then analyzed by an immunoassay.
45. A pharmaceutical composition, comprising an effective amount
of: (a) two or more inhibitors of pAKT, pSTAT3, pVEGFR,
pFKHR-FKHRL1, pFAK, pcKit, pShc, pSMAD2, pSrc, pSTAT5, pAbl, pJak1,
pPyk2, pPDGFRb, pPRAS40, and pEGFR; and (b) a pharmaceutically
acceptable carrier. wherein the inhibitors are selected from the
group consisting of Imatinib mesylate, Bevacizumab, Erlotinib
hydrochloride, Panitumumab, Lapatinib, an akt inhibitor, and a PI3
kinase inhibitor.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to U.S. Provisional
Application No. 60/855,751, filed Nov. 1, 2006, which is hereby
incorporated by reference.
BACKGROUND
[0002] Cancer is a complex and devastating group of diseases that
kills one in five adults in developing countries. Although cancers
arise from a wide variety of cells and tissues in the body, there
are unifying features of this group of diseases. Cancer is
predominantly a genetic disease, resulting from the accumulation of
mutations that promote clonal selection of cells that exhibit
uncontrolled growth and division. The result of such uncontrolled
growth of tumor cells is the formation of disorganized tissue that
compromises the function of normal organs, ultimately threatening
the life of a patient.
[0003] Basic research designed to unravel the mechanics of
carcinogenesis have revolutionized our understanding of the
molecular nature of genetic changes that initiate tumor formation.
Notably, specific genes have been identified that are frequently
mutated in tumor cells. These genes regulate, for example, DNA
damage repair, homologous recombination, cell cycle control, growth
factor signaling, apoptosis, differentiation, angiogenesis, immune
response, cell migration, and telomere maintenance. Thus, based on
mutations in certain genes, it is possible to distinguish cancer
cells from normal cells.
[0004] However, despite advances in our understanding of the
genetic basis underlying cancer cell phenotypes, effective methods
for treating cancer remain largely undiscovered. While
chemotherapeutic agents are designed to kill or block tumor cell
proliferation, chemotherapeutic agents are frequently unable to
exclusively suppress upregulated activity of a particular protein
in a tumor cell without deleteriously affecting necessary levels of
protein activity in normal cells. In the case of metastasized
cancer, current cancer treatments, such as chemotherapy and
radiation, are generally ineffective.
SUMMARY
[0005] In one aspect, the invention provides a method of treating a
patient with liver metastasis, comprising administering to the
patient an effective amount of at least one therapeutic selected
from the group consisting of GLEEVEC, AVASTIN, TARCEVA, Vectibix,
and Lapatinib.
[0006] In one embodiment, the therapeutic is GLEEVEC. In another
embodiment, the therapeutic is AVASTIN. In another embodiment, the
therapeutic is TARCEVA. In another embodiment, the therapeutic is
Lapatinib. In another embodiment, the therapeutic is Vectibix. In
another embodiment, the therapeutic is GLEEVEC and AVASTIN. In
another embodiment, the therapeutic is AVASTIN and TARCEVA. In
another embodiment, the therapeutic is TARCEVA and Lapatinib. In
another embodiment, the therapeutic is GLEEVAC and TARCEVA. In
another embodiment, the therapeutic is GLEEVAC and Lapatinib. In
another embodiment, the therapeutic is GLEEVAC and Vectibix. In
another embodiment, the therapeutic is AVASTIN and Lapatinib.
[0007] In another embodiment, the method may further comprise (a)
procuring a biopsy from the patient; (b) subjecting the biopsy to
epithelial cell enrichment; (c) lysing the epithelal cells to
produce a lysate; (d) analyzing the lysate by immunoassay, wherein
the immunoassay is selected from the group consisting of an ELISA,
reverse phase array, suspension bean array, and immunohistochemical
detection; (e) comparing the intensity value to a high and low
reference value; and (f) reporting the value to a physician.
[0008] In another aspect, the invention provides a method of
treating a patient with pulmonary metastasis, comprising
administering to the patient an effective amount of at least one
therapeutic selected from the group consisting of GLEEVEC, AVASTIN,
TARCEVA, Vectibix, and lapatinib.
[0009] In one embodiment, the therapeutic is GLEEVEC. In another
embodiment, the therapeutic is AVASTIN. In another embodiment, the
therapeutic is TARCEVA. In another embodiment, the therapeutic is
Lapatinib. In another embodiment, the therapeutic is Vectibix. In
another embodiment, the therapeutic is GLEEVEC and AVASTIN. In
another embodiment, the therapeutic is AVASTIN and TARCEVA. In
another embodiment, the therapeutic is TARCEVA and Lapatinib. In
another embodiment, the therapeutic is GLEEVAC and TARCEVA. In
another embodiment, the therapeutic is GLEEVAC and Lapatinib. In
another embodiment, the therapeutic is GLEEVAC and Vectibix. In
another embodiment, the therapeutic is AVASTIN and Lapatinib.
[0010] In one aspect, the invention provides a method of treating a
patient with liver metastasis, comprising administering to the
patient an effective amount of an akt inhibitor.
[0011] In another aspect, the invention provides a method of
treating a patient with pulmonary metastasis, comprising
administering to the patient an effective amount of an akt
inhibitor.
[0012] In another aspect, the invention provides a method of
treating a patient with liver metastasis, comprising administering
to the patient an effective amount of a PI3kinase inhibitor.
[0013] In another aspect, the invention provides a method of
treating a patient with pulmonary metastasis, comprising
administering to the patient an effective amount of a PI3kinase
inhibitor.
[0014] In another aspect, the invention provides a method of
treating a patient with liver metastasis, comprising administering
to the patient an effective amount of an akt inhibitor and at least
one of GLEEVEC, AVASTIN, TARCEVA, vectibix, and lapatinib.
[0015] In another aspect, the invention provides a method of
treating a patient with pulmonary metastasis, comprising
administering to the patient an effective amount of an akt
inhibitor and at least one of GLEEVEC, AVASTIN, TARCEVA, vectibix,
and lapatinib.
[0016] Other objects, features and advantages will become apparent
from the following detailed description. The detailed description
and specific examples are given for illustration only since various
changes and modifications within the spirit and scope of the
invention will become apparent to those skilled in the art from
this detailed description. Further, the examples demonstrate the
principle of the invention and cannot be expected to specifically
illustrate the application of this invention to all the examples
where it will be obviously useful to those skilled in the prior
art.
BRIEF DESCRIPTION OF THE FIGURES
[0017] FIG. 1 shows phosphoendpoints differentially elevated in
liver metastasis compared with the primary colon.
[0018] FIG. 2 shows elevated drug targets and pathways observed in
colorectal carcinoma that metastasized to the lung.
[0019] FIG. 3 depicts phosphorylation of pTEN and phospho akt.
[0020] FIG. 4 provides a three-way map of GLEEVEC.RTM. targets.
[0021] FIG. 5 shows a scatter plot of patient values.
DETAILED DESCRIPTION
[0022] Mapping tumor cell protein networks revealed altered
signaling pathways in cancer progression. In particular, network
analysis of expected kinase substrate cascades revealed differences
between primary and metastatic lesions. Moreover, the elevated
phosphorylation events fell into only a few major signaling
pathways, known to be involved in pro-survival, growth receptor and
motility related events. Targeting the associated kinases would
disrupt aberrant signal pathways and thwart metastasis. Thus,
methods for treating liver or lung metastasis involve administering
therapeutics that target the c-kit, PDGFr, abl family of kinases,
the VEGFr family of kinases, the EGFr family of kinases and the
AKT/mTOR pathway.
[0023] Metastasis refers to a complex series of steps in which
cancer cells migrate from the original tumor site to other parts of
the body via the bloodstream or lymph system. In doing so,
malignant cells detach from the primary tumor and attach to and
degrade proteins comprising the surrounding extracellular matrix
(ECM), which separates the tumor from adjoining tissue. By
degrading these proteins, cancer cells are able to breach the ECM
and migrate.
[0024] The most common places for the metastases to occur are the
adrenals, liver, brain, and the bones. There is also a propensity
for certain tumors to seed in particular organs. For example,
prostate cancer usually metastasizes to the bones. Similarly, colon
cancer has a tendency to metastasize to the liver. Stomach cancer
often metastasizes to the ovary in women, where it forms a
Krukenberg tumor.
[0025] It is this ability to migrate or metastasize to other
tissues and organs that makes cancer a potentially life-threatening
disease. Accordingly, the present invention provides methods for
treating cancer metastasis.
[0026] The present invention uses terms and phrases that are well
known to those practicing the art. Unless defined otherwise, all
technical and scientific terms used herein have the same meaning as
commonly understood by one of ordinary skill in the art to which
this invention belongs. Generally, the nomenclature used herein and
the laboratory procedures in cell culture, molecular genetics, and
nucleic acid chemistry and hybridization described herein are those
well known and commonly employed in the art. Standard techniques
are used for recombinant nucleic acid methods, polynucleotide
synthesis, microbial culture, cell culture, tissue culture,
transformation, transfection, transduction, analytical chemistry,
organic synthetic chemistry, chemical syntheses, chemical analysis,
and pharmaceutical formulation and delivery. Generally, enzymatic
reactions and purification and/or isolation steps are performed
according to the manufacturers' specifications. The techniques and
procedures are generally performed according to conventional
methodology (Molecular Cloning, A Laboratory Manual, 3rd. edition,
edited by Sambrook & Russel Cold Spring Harbor Laboratory
Press, Cold Spring Harbor, N.Y., 2001).
I. Mapping Tumor Cell Networks
[0027] The sequencing of the human genome prompted a revolution in
technologies that facilitate molecular profiling in disease
research. In the field of oncology, for example, array technologies
have been used for profiling tumors at the DNA, RNA, and protein
levels. Protein microarrays provide an especially powerful
technology for drug discovery, biomarker identification, and signal
transduction profiling of cellular material. Specifically, and as
contemplated in the present invention, protein microarrays can be
used for profiling cellular signaling pathways by monitoring
changes in phosphorylation over time, for example, before and after
a particular treatment, between disease and non-disease states, and
between responders and nonresponders. Reviewed in Wulflcuhle, J. et
al. Nat Clin Pract Oncol. 3:5:256-68 (2006), which is hereby
incorporated by reference in its entirety. Based on protein
phosphorylation activities, particular pathways can be identified
and used for tailoring patient-specific treatment. Liotta, L.
Cancer Cell 3:317-325 (2003), which is hereby incorporated by
reference in its entirety.
[0028] Protein microarrays are classified depending on whether the
analytes are captured from solution phase or bound to the solid
phase. In forward-phase arrays (FPAs), the capture molecules are
immobilized onto the substratum and act as a bait molecule. In
contrast, reverse-phase arrays (RPAs) immobilize an individual test
sample in each array spot such that an array is comprised of
hundreds of different samples. Because human tissues comprise
hundreds of interacting cell populations, RPAs afford the ability
to identify changes in the cellular proteome. Methods for preparing
protein microarrays are routine and well-known in the art and are
disclosed, for example, in Nishizuka, S. et al. Proc Natl Acad Sci
USA 100: 14299:14234; Paweletz et al. Oncogene 20:1981-1989
(2001).
[0029] Based on changes in phosphorylation levels, it is possible
to identify signaling pathways that are known to be involved in
pro-survival, growth receptor, and motility related events.
Targeting the associated kinases disrupt aberrant signal pathways
and thereby thwart metastasis. Accordingly, methods for treating
liver or lung metastasis involve administering therapeutics that
target the c-kit, PDGFr, abl family of kinases, the VEGFr family of
kinases, the EGFr family of kinases, and the AKT/mTOR pathway.
II. Therapeutics for Targeting Kinase Pathways
[0030] There are several known and readily available drugs that can
be used for targeting a kinase pathway and thereby used as a means
for treating metastasized cancer, such as liver or lung
metsastasis.
[0031] Imatinib mesylate, also called Gleevec.RTM. or STI571, is
approved by the U.S. Food and Drug Administration (FDA) for the
treatment of some forms of adult and pediatric chronic myelogenous
leukemia (CML), and for the treatment of a rare form of cancer
called gastrointestinal stromal tumor (GIST). Gleevec.RTM. is the
first approved drug to directly turn off the signal of a protein
known to cause a cancer. Gleevec.RTM. inhibits the receptor
tyrosine kinases for platelet-derived growth factor (PDGF) and stem
cell factor (SCF)/c-kit; the SCF/c-kit receptor tyrosine kinase.
Other molecular-targeting drugs previously approved by the FDA
interfere with proteins associated with other cancers, but not with
proteins that directly cause the disease.
[0032] Avastin.RTM., or Bevacizumab, is approved by the FDA for use
with other drugs to treat colorectal cancer that has spread to
other parts of the body. Avastin.RTM. is also approved for use with
other drugs to treat non-small cell lung cancer that cannot be
removed by surgery, has spread to other parts of the body, or has
recurred. Avastin.RTM. is a recombinant humanized monoclonal
antibody directed against the vascular endothelial growth factor
(VEGF), a pro-angiogenic cytokine. Avastin.RTM. binds to VEGF and
inhibits VEGF receptor binding, thereby preventing the growth and
maintenance of tumor blood vessels.
[0033] Tarceva.RTM., also known as erlotinib hydrochloride or
N-(3-Ethynylphenyl)-6,7-bis(2-methoxyethoxy)-4-quinazolinamine
Monohydrochloride, is approved by the FDA for treating forms of
non-small cell lung cancer that have not responded well with
chemotherapy. Competing with adenosine triphosphate, Tarceva.RTM.
reversibly binds to the intracellular catalytic domain of epidermal
growth factor receptor (EGFR) tyrosine kinase, thereby reversibly
inhibiting EGFR phosphorylation and blocking the signal
transduction events and tumorigenic effects associated with EGFR
activation.
[0034] Lapatinib (Tykerb.RTM.) is an experimental drug that blocks
the activity of the HER2 protein, a protein frequently found in
breast cancers. Tumors that overexpress HER2 (called HER2-positive)
tend to grow faster and are more likely to come back than tumors
that don't overexpress the protein. Lapatinib binds to the part of
the HER2 protein found inside breast cancer cells.
[0035] Several signaling pathway inhibitors are known and can be
used as a cancer metastasis therapeutic. Such inhibitors include
but are not limited to akt and PI3kinase inhibitors.
III. Administering Therapeutics
[0036] Each therapeutic drug, or a combination of drugs, may be
administered by a variety of methods. As used herein, therapeutic
drug includes proteins, such as antibodies, and small molecules.
For effective treatment, the present invention contemplates
administering each drug, either alone or in combination with
another drug, by topical, oral, anal, ocular, buccal, nasal,
intramuscular, subcutaneous, intravenous, or parenteral routes. The
ultimate choice of route, formulation, and dose is made by the
attending physician and is based upon a patient's unique
condition.
[0037] Where the administration is by bolus injection, including
bolus injection of a slow release formulation, dosing schedules
also can be continuous in that the drug is administered each day,
or may be discontinuous. Discontinuous bolus injection dosing
schedules, for example, include on periods (days on which an
injection is given) selected from 1, 2, 3, 4, 5, 6, or 7 days, 1,
2, 3, 4, or more weeks, or any combination thereof, and off periods
(days on which an injection is not given) selected from 1, 2, 3, 4,
5, 6, or 7 days, 1, 2, 3, 4, or more weeks.
[0038] As will be apparent to one of skill in the art, intranasal
and inhalant administration is generally more convenient for the
subject as it does not involve the use of injections, catheters or
transdermal infusion devices. Intranasal and inhaled doses may thus
be smaller, but given more frequently, than doses given by other
parenteral routes. Accordingly, intranasal and inhalant dosing
schedules can include a single dose per "on" day, or can involve
multiple doses (e.g., 2, 3, 4, 5 or more doses per day). Dosing
schedules may be continuous or discontinuous, with discontinuous
schedules utilizing on periods (e.g., days on which the drug is
administered at least once) selected from 1, 2, 3, 4, 5, 6, or 7
days, 1, 2, 3, 4, or more weeks, or any combination thereof, and
off periods (e.g., days on which the drug is not administered)
selected from 1, 2, 3, 4, 5, 6, or 7 days, 1, 2, 3, 4, or more
weeks.
[0039] Continuous and discontinuous administration schedules by any
method also include dosing schedules in which the dose is modulated
throughout the "on" period, such that, for example, at the
beginning of the on period, the dose is low and increased until the
end of the on period, the dose is initially high and decreased
during the on period, the dose is initially low, increased to a
peak level, then reduced towards the end of the on period, and any
combination thereof.
[0040] A drug, or combination thereof, may be administered by
parenteral route such as, but not limited to, intravenous (IV),
intramuscular (IM), subcutaneous (SC), intraperitoneal (IP),
transdermal, intranasal, and inhalant routes. IV, IM, SC, and IP
administration can be by bolus injection or bolus infusion or
continuous infusion, and in the case of SC or IM, also can be by
slow release implantable device, including, but not limited to,
pumps, slow release formulations and mechanical devices. For
example, the invention contemplates administering at least one of
GLEEVEC, AVASTIN, TARCEVA, Lapatinib, which of course allows
various drug combinations.
[0041] In general, a dose that is administered by subcutaneous or
intramuscular injection will be greater than the
therapeutically-equivalent dose given intravenously. Preferably,
the drug is dissolved in physiologically compatible carriers such
as normal saline, or phosphate buffered saline solution.
[0042] For potential administration, compositions of the drug may
be semi-solid or liquid preparations, such as liquids, suspensions,
and the like. Physiologically compatible carriers include, but are
not limited to, normal saline, serum albumin, 5% dextrose, and
plasma preparations. For intranasal and inhalant administration, a
powdered formulation, such as a freeze-dried powder, may be useful.
Optionally, the carrier also can include anti-microbial agents,
preservatives, detergents, or surfactants.
[0043] A "therapeutically effective" amount may be determined by
prevention or amelioration of adverse conditions or symptoms of
diseases, injuries, or disorders being treated. In the context of
the present invention, a "therapeutically effective" amount would
reduce metastasis in liver and lung tissues. The dose of a
particular drug to be administered can be determined readily by
those skilled in the art, based on the condition to be treated, the
severity of the condition, the type of cytotoxic agent or radiation
used, and the patient's medical history. Preferably, when the drug
is administered daily, the subcutaneous or intramuscular dose for a
human is about 0.05 mg/kg to 50 mg/kg of body weight per day. More
preferably, the daily subcutaneous or intramuscular dose for a
human is about 0.5 mg/kg to 20 mg/kg. Most preferably, the daily
subcutaneous or intramuscular dose for a human is about 0.5 mg/kg
to 5 mg/kg. For subcutaneous or intramuscular administration, the
dose is preferably greater than the therapeutically equivalent dose
given intravenously.
[0044] For example, the FDA recommended dosage of Gleevec is 400 mg
per day for patients in chronic phase CML and 600 mg per day for
patients in accelerated phase or blast crisis. Based on these
guidelines, and depending on the severity of the cancer metastasis
and the health of the patient, the dosage could be 400-420,
420-440, 440-460, 460-480, 480-500, 500-520, 520-540, 540-560,
560-580, and 580-600 mg per day. The prescribed dose should be
administered orally, once daily with a meal and a large glass of
water. Treatment should be continued as long as the patient
continues to benefit. The formulation, route and method of
administration, and dosage will depend on the medical history of
the patient, including the severity of the disease.
[0045] A therapeutic drug can be combined with another drug, along
with inert pharmaceutical excipients such as lactose, oil,
mannitol, and starch to form pharmaceutical compositions. Such
compositions can be formulated into dosage forms such as elixirs,
liquids, ointments, lotions, IV fluids, alcohol, tablets, capsules,
and the like. For parenteral, intramuscular, subcutaneous and
intravenous administration, a drug can be formulated with an inert,
parenterally acceptable vehicle such as water, saline, sesame oil,
ethanol buffered aqueous medium, propylene glycol and the like. For
topical and oral administration, a drug can be formulated with
waxes, oils, buffered aqueous medium, and the like. These various
pharmaceutical dosage forms are compounded by methods well-known in
the art.
[0046] The drugs of the present invention also can be prepared as
pharmaceutically acceptable salts. A pharmaceutically acceptable
salt means a salt formed between any one or more of the charged
groups in a peptide and any one or more pharmaceutically
acceptable, non-toxic cations or anions. Organic and inorganic
salts include, for example, those prepared from acids such as
hydrochloric, sulfuric, sulfonic, tartaric, fumaric, hydrobromic,
glycolic, citric, maleic, phosphoric, succinic, acetic, nitric,
benzoic, ascorbic, p-toluenesulfonic, benzenesulfonic,
naphthalenesulfonic, propionic, carbonic, and the like.
Pharmaceutically acceptable salts may also contain cations
including, but not limited to, ammonium, sodium, potassium,
calcium, or magnesium.
[0047] Specific examples are presented below for controlling cancer
metastasis. They are meant to be exemplary and not as limitations
on the present invention.
EXAMPLES
Example 1
Mapping Tumor Cell Networks
[0048] Using laser capture microdissection, 68 cases of
patient-matched (from 34 patients) colorectal cancer and hepatic
metastasis and 15 cases of pulmonary metastasis from the colon were
obtained. Pure cancer cell populations were lysed and subjected to
reverse-phase protein microarray technology as described in
Petricoin, E. et al. J Clin Oncol, 23:15:3614-20 (2003) and
incorporated by reference in its entirety.
[0049] Using this technique, the phosphorylation state of 70 kinase
substrates were measured and molecular network analysis was
performed using commercially available software (Microvigene,
VigeneTech, MA). Of the 70 phosphoendpoints analyzed, 21 were
statistically significant (Student t-test p<0.05) and were
expressed between the patient matched colorectal signature and the
hepatic metastasis.
[0050] As shown in FIG. 1, of the 21 statistically significant
phosphoendpoints, 14 were elevated in liver metastasis compared
with the primary colon. Unexpectedly, the 14 phosphorylation events
fell into only a few major signaling pathways. These pathways are
known to be involved in pro-survival, growth receptor, and motility
related events, and many of which are drug targets for specific
targeted therapies.
[0051] Moreover, these endpoints nucleated into well-known pathways
whereby levels of proteins that are known to be antagonists of the
signaling were also predictably down-regulated in liver metastasis.
This critical observation provides a large degree of confidence in
the final data insofar as entire pathways have been discovered to
be deranged. For example, up-regulating a growth factor receptor
(i.e. GLEEVEC, AVASTIN, and TARCEVA targets) concomitant with
up-regulating downstream prosurvival components pAK, pRAS, FKHRL,
STAT5, pSRC, and down-regulation of the tumor suppressor pTEN
(controls AKT activity), cleaved caspase 9 and BAD phosphorylation
(controls apoptosis) as well as down-regulating PKCalpha and
zet-lambda (both shown to modulate AKT activation) were all
observed, and were entirely contained within the statistically
significant endpoints.
[0052] Furthermore, and as shown in FIG. 2, elevated drug targets
and pathways were also observed in colorectal carcinoma that
metastasized to the lung, indicating that these pathways could be
targeted in both liver and pulmonary metastasis.
[0053] FIG. 3 provides a two-way plot using two of the evaluated
phosphoendpoints from FIG. 1. In measuring the phosphorylation of
pTEN, which regulates akt, and phospho akt, it demonstrates
activation of the signaling cascade. When pTEN is phosphorylated,
it becomes destabilized and degraded, which prompts akt
phosphorylation. From these data, it is apparent that the signaling
pathway is dysregulated and in doing so, a cancer cell in the liver
would selectively die, thereby thwarting metastasis.
[0054] FIG. 4 provides a three-way map for three GLEEVEC targets,
each of which are independently activated, thereby providing
evidence that dysregulation of a signaling pathway activates cell
death.
Example 2
Selecting and Stratifying Patients for Treatment
[0055] A predefined cut-value of combined PDGFr/c-kit and c-abl
phosphorylation levels (normalized and compared to A431+/-EGF
reference material), herein referred to as phospho-Gleevec drug
target score (PGDT) can be obtained and used to select and stratify
patients for Gleevec therapy. The PGDT is determined by the
combination of the phosphorylation of c-abl, c-kit and PDGFr for
each patient. The relative phosphorylation of each has been
determined based on a reference standard, A431 cells treated with
100 ng/ml EGF for 15 minutes, that is printed and quantified on
every assay. The inter and intra assay CV for the PGDT of the
A431+EGF reference material was less than 5%, well within clinical
diagnostic standard analyte testing determiniant, and within our
previously published results (Paweletz et al, Oncogene, 2001).
[0056] For example, a 28 day run using Gleevec alone prior to
administration of panitumumab may be used. Based on preliminary
data, a PGDT cut-point of 2.3 relative intensity units (RIU) may be
used, since this value would provide 100% specificity: the
rationale being that 0/34 primary colorectal cancers would have a
value above this cut-point (P<0.00001), as shown below in FIG.
5. A PGDT cut point of 2.3 RIU theoretically would have stratified
the top third of all patients within the study set (11/34 patients
(33%). Based on the highly statistical significance of the
separation obtained between primary and metastatic disease, it is
likely that one-third of the enrollment population will be eligible
for the Gleevec arm of the study. Importantly, a PGDT of 2.3
corresponds to a value slightly higher than the average of
quintuplicate replicates of the A431 control cell values
(2.15+/-0.09). A431 cells are well known to have extraordinarily
high EGFR levels and fairly high constitutive signaling in the
absence of EGF ligand. Thus, the data provide evidence that using a
2.3 cut-point can be used to select those patients whose metastatic
signaling is above the highest signal from any of the primary
colorectal cancer levels and higher than the unstimulated A431 cell
line.
Example 3
GLEEVEC+Vectibix Treatment
[0057] Gleevec either alone or in combination with Vectibix may be
effective in the treatment of metastatic colorectal cancer as
measured by reduction in tumor size by imaging of the hepatic
metastasis. Additionally, Gleevec and Vectibix efficacy may
correlate with phosphorylation levels of PDGFr (Y751), cKIT (Y703),
cAbl (Y245), AKT (S473), AKT (T308), mTOR (S2448), mTOR (S2481),
4EBP1 (S65), 4EBP1 (T37/46), 4EBP1 (T70), total EGFR, phospho
EGFR(Y1173, Y1068, Y1045, Y845, Y992) using liver biopsy specimens
obtained before and after therapy.
[0058] For example, patients with a PGDT score >2.3, as defined
above in Example 2, will receive Gleevec monotherapy for a 28 day
period with a second biopsy on around day 28. Thereafter, all
patients will receive a combination of Gleevec and Vectibix (6
mg/kg). For patients with PGDT Score <2.3, each patient will
receive standard of care therapy (Vectibix containing regimen per
investigator's discretion). All protein determinants will be
measured using reverse phase protein microarray (RPMA) as described
above in Example 1.
TABLE-US-00001 TABLE 1 Clinical characterization of the 34 matched
patients affected by colorectal cancer and synchronous liver
metastases and the 16 cases of metachronous lung metastases from
colorectal cancer. METASTATIC COLON CANCER MATCHED SYNCHRONOUS
LIVER METASTASES MALE FEMALE TOTAL NUMBER OF PATIENTS: 21 13 34
MEDIAN AGE: 59.8 57.07 YEARS (RANGE): 37-84 33-66 STAGE: IV-DUKES
STAGE D 21 13 34 SIT OF THE PRIMARY TUMOR TOTAL CAECUM/ASCENDING 4
TRANSVERSE 1 DESCENDING/SIGMOID 10 RECTUM 18 UNKNOWN 1
Table 1 provides data describing the study set sample, which
represents a typical population distribution for patients with
colorectal cancer that had metastasized to the liver.
TABLE-US-00002 TABLE 2 Sample study of patients with lung cancer
that has metastasized to the liver. LUNG METASTASES FROM COLON
CANCER MALE FEMALE TOTAL NUMBER OF PATIENTS: 6 10 16 MEDIAN AGE:
60.7 53 YEARS (RANGE): 40-74 27-74
Table 2 provides data describing the study set sample, which
represents a typical population distribution for patients with lung
cancer that had metastasized to the liver.
TABLE-US-00003 TABLE 3 Statistically significant endpoints between
primary colorectal cancer and synchronous liver metastases. P TREND
IN THE LIVER VARIABLE VALUE METASTASES Cl Caspase9 D315 0.0066
.dwnarw. EGFR 0.0019 .uparw. p4EBP1 S65 0.0326 .uparw. pAbl Y245
0.0037 .uparw. pAKT S473 0.0001 .uparw. pAKT T308 0.0163 .uparw.
pBAD S136 0.0087 .dwnarw. pcAbl T735 0.0251 .dwnarw. pcKit Y703
0.005 .uparw. pEGFR Y 1448 0.0424 .dwnarw. peIF4G S1108 0.0416
.uparw. peNOS S1177 0.0476 .uparw. pFADDS194 0.0222 .uparw. pFAK
Y576-577 0.0001 .uparw. pFKHR-FKHRL1 T24-T32 0.0001 .uparw. pIKBa
S32 0.0341 .dwnarw. pIKBa S32-36 0.0212 .dwnarw. pmTOR S 2481
0.0436 .dwnarw. pP70 S6 S371 0.0006 .dwnarw. pPDGFR beta Y716
0.0262 .dwnarw. pPDGFRb Y751 0.0181 .uparw. pPDK1 S241 0.0051
.dwnarw. pPKC theta T538 0.0182 .uparw. pPKC zeta-lambda T410-403
0.0001 .dwnarw. pPKCa S657 0.0001 .dwnarw. pPKCa-b II T638-641
0.0017 .dwnarw. pPRAS40 0.0003 .uparw. pPyk2 Y402 0.0001 .uparw.
pShc Y317 0.0001 .uparw. pSMAD2 S465-467 0.0043 .uparw. pSrc Y527
0.0001 .uparw. pSTAT3 Y705 0.0225 .dwnarw. pSTAT5 Y694 0.0159
.uparw. pTEN S380 0.0001 .dwnarw. pVEGFR Y951 0.0001 .uparw.
pVEGFR2 Y1175 0.0481 .uparw.
Table 3 shows, out of the 80 proteins analyzed, the statistically
significant endpoints between primary colorectal cancer and
synchronous liver metastases. The right column shows the trend in
the hepatic lesions presented in comparison to the primary
cancer.
* * * * *