WO2008121307A2 - Procédés et compositions pour identifier le cancer de la prostate ou une réponse immunitaire à médiation humorale contre le cancer de la prostate - Google Patents

Procédés et compositions pour identifier le cancer de la prostate ou une réponse immunitaire à médiation humorale contre le cancer de la prostate Download PDF

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Publication number
WO2008121307A2
WO2008121307A2 PCT/US2008/004016 US2008004016W WO2008121307A2 WO 2008121307 A2 WO2008121307 A2 WO 2008121307A2 US 2008004016 W US2008004016 W US 2008004016W WO 2008121307 A2 WO2008121307 A2 WO 2008121307A2
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Prior art keywords
measured
responsiveness
cancer therapy
increased
immune response
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PCT/US2008/004016
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English (en)
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WO2008121307A3 (fr
Inventor
Karin Jooss
Thomas Harding
Minh Nguyen
Kathryn E. Koprivnikar
Alan J. Korman
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Cell Genesys, Inc.
Medarex, Inc.
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Application filed by Cell Genesys, Inc., Medarex, Inc. filed Critical Cell Genesys, Inc.
Publication of WO2008121307A2 publication Critical patent/WO2008121307A2/fr
Publication of WO2008121307A3 publication Critical patent/WO2008121307A3/fr

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
    • G01N33/57407—Specifically defined cancers
    • G01N33/57434—Specifically defined cancers of prostate
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/52—Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/60—Complex ways of combining multiple protein biomarkers for diagnosis

Definitions

  • the present invention relates to prostate cancer markers, compositions comprising such markers, immunoglobulins specific for such markers, and methods of using such markers and/or immunoglobulins to assess an immune response against prostate cancer.
  • An immune response against the markers correlates with an immune response, in particular a humoral immune response, against prostate cancer cells which immune response is preferably associated with prophylaxis of prostate cancer, treatment of prostate cancer, and/or amelioration of at least one symptom associated with prostate cancer.
  • the immune system plays a critical role in the pathogenesis of a wide variety of cancers. When cancers progress, it is widely believed that the immune system either fails to respond sufficiently or fails to respond appropriately, allowing cancer cells to grow.
  • standard medical treatments for cancer including chemotherapy, surgery, radiation therapy and cellular therapy have clear limitations with regard to both efficacy and toxicity. To date, these approaches have met with varying degrees of success dependent upon the type of cancer, general health of the patient, stage of disease at the time of diagnosis, etc.
  • Improved strategies that combine specific manipulation of the immune response to cancer in combination with standard medical treatments may provide a means for enhanced efficacy and decreased toxicity.
  • cytokines which express cytokines locally at the vaccine site.
  • Activity has been demonstrated in tumor models using a variety of immunomodulatory cytokines, including IL-4, IL-2, TNF-alpha, G-CSF, IL-7, IL-6 and GM-CSF, as described in Golumbeck PT et al, Science 254:13-716, 1991; Gansbacher B et al, J. Exp. Med. 172:1217-1224, 1990; Fearon ER et al, Cell 60:397-403, 1990; Gansbacher B et al, Cancer Res.
  • the present invention provides prostate cancer markers, compositions comprising such markers, immunoglobulins specific for such markers, and methods of using such markers and/or immunoglobulins to assess an immune response against prostate cancer.
  • An immune response against the markers correlates with an immune response, in particular a humoral immune response against prostate cancer cells.
  • the immune response is preferably associated with prophylaxis of prostate cancer, treatment of prostate cancer, and/or amelioration of at least one symptom associated with prostate cancer.
  • the invention provides a method for identifying whether a subject is afflicted with prostate cancer, comprising detecting an immune response against an antigen identified in Table 1 or 2, wherein detection of the immune response indicates that the subject is afflicted with prostate cancer.
  • the subject is a mammal.
  • the subject is a human.
  • the immune response is a humoral immune response.
  • the immune response is a cellular immune response.
  • an immune response is detected against an antigen identified in Table 1.
  • an immune response is detected against an antigen identified in Table 2.
  • an immune response is detected against 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 of the antigens in Table 1.
  • the invention provides a method for determining whether a subject is likely to respond to prostate cancer therapy with a combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation- incompetent and have been genetically engineered to express GM-CSF, the method comprising detecting an immune response against an antigen listed in Table 1 or 2, wherein detecting the immune response indicates that the subject is likely to respond to said prostate cancer therapy.
  • the prostate cancer therapy can be other than a therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF; in such embodiments, the prostate cancer therapy can be any cancer immunotherapy known to one skilled in the art without limitation.
  • the subject is a mammal. In certain embodiments, the subject is a human.
  • the cancer cells are autologous. In certain embodiments, the cancer cells are allogeneic. In certain embodiments, the cancer cells are LnCaP cells or PC3 cells.
  • the anti-CTLA-4 monoclonal antibody is mammalian. In certain embodiments, the anti-CTLA-4 antibody is fully human. In certain embodiments, the anti-CTLA-4 antibody is humanized. In certain embodiments, the anti-CTLA-4 antibody is monoclonal. [0012] In certain embodiments, an immune response is detected against an antigen listed in Table 1. In certain embodiments, an immune response is detected against an antigen listed in Table 2. In certain embodiments, an immune response is detected against one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, or more of the antigens listed in Table 1.
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers, increased overall survival time, increased progression-free survival, decreased tumor size, decreased bone metastasis marker response, increased impact on minimal residual disease, increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent, increased induction of delayed-type- hypersensitivity (DTH) response to injections of autologous tumor, increased induction of T cell response to autologous tumor or candidate tumor-associated antigens, increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response, decreased concentrations of prostate- specific antigen (PSA), reduced slope of PSA doubling time, increased PSA doubling time, reduced metastisis as measured by bone scan, increased time to progression, increased survival time as compared to the Halabi nomogram, decreased serum concentrations of ICTP, or decreased concentrations of serum C-reactive protein. See Halabi et al., 2003. J Clin Oncol 21 :1232-7,
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers, e.g., prostate-specific antigen (PSA).
  • responsiveness to the combination cancer therapy is measured by increased overall survival time.
  • responsiveness to the combination cancer therapy is measured by increased progression-free survival.
  • responsiveness to the combination cancer therapy is measured by decreased tumor size.
  • responsiveness to the combination cancer therapy is measured by decreased bone metastasis marker response.
  • responsiveness to the combination cancer therapy is measured by increased impact on minimal residual disease.
  • responsiveness to the combination cancer therapy is measured by increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent.
  • responsiveness to the combination cancer therapy is measured by increased induction of delayed -type-hypersensitivity (DTH) response to injections of autologous tumor. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of T cell response to autologous tumor or candidate tumor-associated antigens. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response. [0015] In certain embodiments, the immune response is a humoral immune response. In certain embodiments, the immune response is a cellular immune response.
  • DTH delayed -type-hypersensitivity
  • the invention provides a computer-implemented method for determining whether a subject is likely to respond to prostate cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF, the method comprising inputting into a computer memory data indicating whether an immune response against an antigen listed in Table 1 or 2 is detected, inputting into the computer memory a correlation between an immune response against an antigen listed in Table 1 or 2 and a likelihood of responding to said therapy, and determining whether the subject is likely to respond to said therapy.
  • the subject is a mammal. In certain embodiments, the subject is a human.
  • the cancer cells are autologous. In certain embodiments, the cancer cells are allogeneic. In certain embodiments, the cancer cells are LnCaP cells or PC3 cells. In certain embodiments, the anti-CTLA-4 antibody is fully human. In certain embodiments, the anti-CTLA-4 antibody is humanized. In certain embodiments, the anti-CTLA-4 antibody is monoclonal. [0019] In certain embodiments, an immune response is detected against an antigen listed in Table 1. In certain embodiments, an immune response is detected against an antigen listed in Table 2.
  • an immune response is detected against one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, or more of the antigens listed in Table 1.
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers, increased overall survival time, increased progression-free survival, decreased tumor size, decreased bone metastasis marker response, increased impact on minimal residual disease, increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent, increased induction of delayed-type- hypersensitivity (DTH) response to injections of autologous tumor, increased induction of T cell response to autologous tumor or candidate tumor-associated antigens, increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response, decreased concentrations of prostate- specific antigen (PSA), reduced slope of PSA doubling time, increased PS A doubling time, reduced metastisis as measured by bone scan, increased time to progression, increased survival time
  • PSA prostate- specific antigen
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased overall survival time. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased progression-free survival. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased tumor size. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased bone metastasis marker response. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on minimal residual disease. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of antibody response to the cancer cells that have been rendered proliferation- incompetent.
  • responsiveness to the combination cancer therapy is measured by increased induction of delayed-type-hypersensitivity (DTH) response to injections of autologous tumor.
  • responsiveness to the combination cancer therapy is measured by increased induction of T cell response to autologous tumor or candidate tumor-associated antigens.
  • responsiveness to the combination cancer therapy is measured by increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response.
  • DTH delayed-type-hypersensitivity
  • the immune response is a humoral immune response. In certain embodiments, the immune response is a cellular immune response.
  • the invention provides a method for determining whether a subject is responding to prostate cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation- incompetent and have been genetically engineered to express GM-CSF, the method comprising administering an effective amount of the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF, and detecting an immune response against an antigen listed in Table 1 or 2, wherein detecting the immune response indicates that the subject is responding to said prostate cancer therapy.
  • the subject is a mammal. In certain embodiments, the subject is a human. In certain embodiments, the cancer cells are autologous. In certain embodiments, the cancer cells are allogeneic. In certain embodiments, the cancer cells are LnCaP cells or PC3 cells. In certain embodiments, the anti-CTLA-4 monoclonal antibody is fully human. In certain embodiments, the anti-CTLA-4 monoclonal antibody is humanized.
  • an immune response is detected against an antigen listed in Table 1. In certain embodiments, an immune response is detected against an antigen listed in Table 2. In certain embodiments, an immune response is detected against one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, or more of the antigens listed in Table 1.
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers, increased overall survival time, increased progression-free survival, decreased tumor size, decreased bone metastasis marker response, increased impact on minimal residual disease, increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent, increased induction of delayed-type- hypersensitivity (DTH) response to injections of autologous tumor, increased induction of T cell response to autologous tumor or candidate tumor-associated antigens, or increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response.
  • DTH delayed-type- hypersensitivity
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased overall survival time. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased progression-free survival. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased tumor size. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased bone metastasis marker response. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on minimal residual disease. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of antibody response to the cancer cells that have been rendered proliferation- incompetent.
  • responsiveness to the combination cancer therapy is measured by increased induction of delayed-type-hypersensitivity (DTH) response to injections of autologous tumor.
  • responsiveness to the combination cancer therapy is measured by increased induction of T cell response to autologous tumor or candidate tumor-associated antigens.
  • responsiveness to the combination cancer therapy is measured by increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response.
  • the immune response is a humoral immune response. In certain embodiments, the immune response is a cellular immune response.
  • the invention provides a computer-implemented method for determining whether a subject responding to prostate cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF, the method comprising administering an effective amount of the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation- incompetent and have been genetically engineered to express GM-CSF, inputting into a computer memory data indicating whether an immune response against an antigen listed in Table 1 or 2 is detected, inputting into the computer memory a correlation between an immune response against an antigen listed in Table 1 or 2, and responsiveness to said therapy, and determining whether the subject is responding to said therapy.
  • the subject is a mammal. In certain embodiments, the subject is a human.
  • the cancer cells are autologous. In certain embodiments, the cancer cells are allogeneic. In certain embodiments, the cancer cells are LnCaP cells or PC3 cells. In certain embodiments, the anti-CTLA-4 antibody is fully human. In certain embodiments, the anti-CTLA-4 antibody is humanized. In certain embodiments, the anti-CTLA-4 antibody is monoclonal. [0032] In certain embodiments, an immune response is detected against an antigen listed in Table 1. In certain embodiments, an immune response is detected against an antigen listed in Table 2.
  • an immune response is detected against one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, or more of the antigens listed in Table 1.
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers, increased overall survival time, increased progression-free survival, decreased tumor size, decreased bone metastasis marker response, increased impact on minimal residual disease, increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent, increased induction of delayed-type- hypersensitivity (DTH) response to injections of autologous tumor, increased induction of T cell response to autologous tumor or candidate tumor-associated antigens, or increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response.
  • DTH delayed-type- hypersensitivity
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased overall survival time. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased progression- free survival. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased tumor size. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased bone metastasis marker response. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on minimal residual disease. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of antibody response to the cancer cells that have been rendered proliferation- incompetent.
  • responsiveness to the combination cancer therapy is measured by increased induction of delayed-type-hypersensitivity (DTH) response to injections of autologous tumor.
  • responsiveness to the combination cancer therapy is measured by increased induction of T cell response to autologous tumor or candidate tumor-associated antigens.
  • responsiveness to the combination cancer therapy is measured by increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response.
  • DTH delayed-type-hypersensitivity
  • the immune response is a humoral immune response. In certain embodiments, the immune response is a cellular immune response.
  • the invention provides a method for determining whether a subject is responding to prostate cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation- incompetent and have been genetically engineered to express GM-CSF, the method comprising detecting an immune response against an antigen listed in Table 1 or 2 at a first time, administering an effective amount of the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF, and detecting an immune response against the antigen listed in Table 1 or 2 at a later second time, wherein an increase in the immune response detected at the later second time relative to the earlier first time indicates that the subject is responding to said prostate cancer therapy.
  • the subject is a mammal. In certain embodiments, wherein the subject is a human.
  • the cancer cells are autologous. In certain embodiments, the cancer cells are allogeneic. In certain embodiments, the cancer cells are LnCaP cells or PC3 cells.
  • the anti-CTLA-4 antibody is fully human. In certain embodiments, the anti-CTLA-4 antibody is humanized. In certain embodiments, the anti-CTLA-4 antibody is monoclonal.
  • an immune response is detected at the first and second times against an antigen listed in Table 1. In certain embodiments, an immune response is detected at the first and second times against an antigen listed in Table 2. In certain embodiments, an immune response is detected at the first and second times against one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, or more of the antigens listed in Table 1.
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers, increased overall survival time, increased progression-free survival, decreased tumor size, decreased bone metastasis marker response, increased impact on minimal residual disease, increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent, increased induction of delayed-type- hypersensitivity (DTH) response to injections of autologous tumor, increased induction of T cell response to autologous tumor or candidate tumor-associated antigens, increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response, decreased concentrations of PSA, reduced slope of PSA doubling time, increased PSA doubling time, reduced metastisis as measured by bone scan, increased time to progression, increased survival time as compared to the Halabi nomogram, decreased serum concentrations of ICTP, or decreased concentrations of serum C-reactive protein.
  • DTH delayed-type- hypersensitivity
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased overall survival time. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased progression- free survival. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased tumor size. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased bone metastasis marker response. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on minimal residual disease. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of antibody response to the cancer cells that have been rendered proliferation- incompetent.
  • responsiveness to the combination cancer therapy is measured by increased induction of delayed-type-hypersensitivity (DTH) response to injections of autologous tumor. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of T cell response to autologous tumor or candidate tumor-associated antigens. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased concentrations of PSA. In certain embodiments, responsiveness to the combination cancer therapy is measured by reduced slope of PSA doubling time. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased PSA doubling time.
  • DTH delayed-type-hypersensitivity
  • responsiveness to the combination cancer therapy is measured by reduced metastisis as measured by bone scan. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased time to progression. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased survival time as compared to the Halabi nomogram. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased serum concentrations of ICTP. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased concentrations of serum C-reactive protein
  • the immune response detected at the first and second times is a humoral immune response. In certain embodiments, the immune response detected at the first and second times is a cellular immune response.
  • the invention provides a computer-implemented method for determining whether a subject is responding to prostate cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF, the method comprising administering an effective amount of the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation- incompetent and have been genetically engineered to express GM-CSF, inputing into a computer memory data indicating whether an immune response against an antigen listed in Table 1 or 2 is detected at a first time prior to said step of administering and at a later second time subsequent to said step of administering, inputting
  • the subject is a mammal. In certain embodiments, the subject is a human. In certain embodiments, the cancer cells are autologous. In certain embodiments, the cancer cells are allogeneic. In certain embodiments, the cancer cells are LnCaP cells or PC3 cells. In certain embodiments, the anti-CTLA-4 antibody is fully human. In certain embodiments, the anti-CTLA-4 antibody is humanized. In certain embodiments, the anti-CTLA-4 antibody is monoclonal. [0044] In certain embodiments, an immune response is detected at said first time and said second time against an antigen listed in Table 1. In certain embodiments, an immune response is detected at said first time and said second time against an antigen listed in Table 2.
  • an immune response is detected at said first time and said second time against one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, or more of the antigens listed in Table 1.
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers, increased overall survival time, increased progression-free survival, decreased tumor size, decreased bone metastasis marker response, increased impact on minimal residual disease, increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent, increased induction of delayed-type- hypersensitivity (DTH) response to injections of autologous tumor, increased induction of T cell response to autologous tumor or candidate tumor-associated antigens, increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response, decreased concentrations of PSA, reduced slope of PSA doubling time, increased PSA doubling time, reduced metastisis as measured by bone scan, increased time to progression, increased
  • DTH delayed-type- hypersensitivity
  • responsiveness to the combination cancer therapy is measured by decreased serum concentrations of tumor specific markers. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased overall survival time. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased progression-free survival. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased tumor size. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased bone metastasis marker response. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on minimal residual disease. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of antibody response to the cancer cells that have been rendered proliferation- incompetent.
  • responsiveness to the combination cancer therapy is measured by increased induction of delayed-type-hypersensitivity (DTH) response to injections of autologous tumor. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased induction of T cell response to autologous tumor or candidate tumor-associated antigens. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased concentrations of PSA. In certain embodiments, responsiveness to the combination cancer therapy is measured by reduced slope of PSA doubling time. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased PSA doubling time.
  • DTH delayed-type-hypersensitivity
  • responsiveness to the combination cancer therapy is measured by reduced metastisis as measured by bone scan. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased time to progression. In certain embodiments, responsiveness to the combination cancer therapy is measured by increased survival time as compared to the Halabi nomogram. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased serum concentrations of ICTP. In certain embodiments, responsiveness to the combination cancer therapy is measured by decreased concentrations of serum C-reactive protein.
  • the immune response is a humoral immune response. In certain embodiments, the immune response is a cellular immune response.
  • the invention provides computer-readable media embedded with computer executable instructions for performing a method of the invention.
  • the invention provides a computer system configured to perform a method of the invention.
  • the present invention provides prostate cancer markers, compositions comprising such markers, immunoglobulins specific for such markers, and methods of using such markers and/or immunoglobulins to assess an immune response against prostate cancer.
  • the markers, compositions, immunoglobulins, and methods are useful, for example, for assessing an immune response, in particular a humoral immune response against prostate cancer cells, wherein the immune response is preferably associated with prophylaxis of prostate cancer, treatment of prostate cancer, and/or amelioration of at least one symptom associated with prostate cancer.
  • one aspect of the immune response induced by coadministration of genetically modified tumor cells that express a cytokine with an antibody that specifically binds to CTLA-4 is an immune response against certain polypeptides expressed by the genetically modified tumor cell and/or cells from the tumor afflicting the subject. It is also believed that this immune response plays an important role in the effectiveness of this therapy to treat, e.g., prostate cancer.
  • cytokine or grammatical equivalents, herein is meant the general class of hormones of the cells of the immune system, including lymphokines, monokines, and others.
  • the definition includes, without limitation, those hormones that act locally and do not circulate in the blood, and which, when used in accord with the present invention, will result in an alteration of an individual's immune response.
  • cytokine or "cytokines” as used herein refers to the general class of biological molecules, which affect cells of the immune system. The definition is meant to include, but is not limited to, those biological molecules that act locally or may circulate in the blood, and which, when used in the compositions or methods of the present invention serve to regulate or modulate an individual's immune response to cancer.
  • Exemplary cytokines for use in practicing the invention include, but are not limited to, interferon-alpha (IFN-alpha), IFN-beta, and IFN-gamma, interleukins (e.g., IL-I to IL-29, in particular, IL-2, IL-7, IL- 12, IL- 15 and IL- 18), tumor necrosis factors (e.g., TNF-alpha and TNF-beta), erythropoietin (EPO), MIP3a, ICAM, macrophage colony stimulating factor (M-CSF), granulocyte colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF).
  • IFN-alpha interferon-alpha
  • IFN-beta interleukins
  • interleukins e.g., IL-I to IL-29, in particular, IL-2, IL-7, IL- 12, IL- 15 and
  • cancer As used herein, the terms “cancer,” “cancer cells,” “neoplastic cells,” “neoplasia,” “tumor,” and “tumor cells” (used interchangeably) refer to cells that exhibit relatively autonomous growth, so that they exhibit an aberrant growth phenotype or aberrant cell status characterized by a significant loss of control of cell proliferation.
  • a tumor cell may be a hyperplastic cell, a cell that shows a lack of contact inhibition of growth in vitro or in vivo, a cell that is incapable of metastasis in vivo, or a cell that is capable of metastasis in vivo.
  • Neoplastic cells can be malignant or benign. It follows that cancer cells are considered to have an aberrant cell status.
  • Tumor cells may be derived from a primary tumor or derived from a tumor metastases.
  • the “tumor cells” may be recently isolated from a patient (a "primary tumor cell”) or may be the product of long term in vitro culture.
  • the term "primary tumor cell” is used in accordance with the meaning in the art.
  • a primary tumor cell is a cancer cell that is isolated from a tumor in a mammal and has not been extensively cultured in vitro.
  • tumor antigen from a tumor cell and “tumor antigen” and “tumor cell antigen” may be used interchangeably herein and refer to any protein, peptide, carbohydrate or other component derived from or expressed by a tumor cell which is capable of eliciting an immune response.
  • the definition is meant to include, but is not limited to, whole tumor cells, tumor cell fragments, plasma membranes taken from a tumor cell, proteins purified from the cell surface or membrane of a tumor cell, unique carbohydrate moieties associated with the cell surface of a tumor cell or tumor antigens expressed from a vector in a cell.
  • the definition also includes those antigens from the surface of the cell, which require special treatment of the cells to access.
  • the term "genetically modified tumor cell” as used herein refers to a composition comprising a population of cells that has been genetically modified to express a transgene, and that is administered to a patient as part of a cancer treatment regimen.
  • the genetically modified tumor cell vaccine comprises tumor cells which are "autologous” or “allogeneic” to the patient undergoing treatment or "bystander cells” that are mixed with tumor cells taken from the patient.
  • the genetically modified tumor cell is of the same general type of tumor cell as is afflicting the patient, e.g., if the patient is afflicted with metastatic prostate cancer, the genetically modified tumor cell is also a metastatic prostate cancer cell.
  • a GM-CSF- expressing genetically modified tumor cell vaccine may be referred to herein as "GV AX®".
  • GV AX® Autologous and allogeneic cancer cells that have been genetically modified to express a cytokine, e.g., GM-CSF, followed by readministration to a patient for the treatment of cancer are described in U.S. Pat. Nos. 5,637,483, 5,904,920, 6,277,368 and 6,350,445, each of which is expressly incorporated by reference herein.
  • a form of GM-CSF-expressing genetically modified cancer cells or a "cytokine-expressing cellular vaccine" for the treatment of pancreatic cancer is described in U.S. Pat. Nos.
  • Cells may be genetically modified to increase the expression of a cytokine, such as GM-CSF, or an antigen the immune response to which is enhanced following administration of a cytokine-expressing cellular vaccine, such as GV AX®.
  • the expression of an endogenous antigen may be increased using any method known in the art, such as genetically modifying promoter regions of genomic sequences or genetically altering cellular signaling pathways to increase production of the antigen.
  • cells can be transduced with a vector coding for the antigen or immunogenic fragment thereof.
  • systemic immune response or grammatical equivalents herein is meant an immune response which is not localized, but affects the individual as a whole, thus allowing specific subsequent responses to the same stimulus.
  • proliferation-incompetent or “inactivated” refers to cells that are unable to undergo multiple rounds of mitosis, but still retain the capability to express proteins such as cytokines or tumor antigens. This may be achieved through numerous methods known to those skilled in the art. Embodiments of the invention include, but are not limited to, treatments that inhibit at least about 95%, at least about 99% or substantially 100% of the cells from further proliferation.
  • the cells are irradiated at a dose of from about 50 to about 200 rads/min or from about 120 to about 140 rads/min prior to administration to the mammal.
  • the levels required are 2,500 rads, 5,000 rads, 10,000 rads, 15,000 rads or 20,000 rads.
  • the cells produce beta-f ⁇ lamin or immunogenic fragment thereof, two days after irradiation, at a rate that is at least about 10%, at least about 20%, at least about 50% or at least about 100% of the pre-irradiated level, when standardized for viable cell number.
  • cells are rendered proliferation incompetent by irradiation prior to administration to the subject. [0060]
  • individual by the term "subject” or grammatical equivalents thereof is meant any one individual mammal.
  • cytotoxic T lymphocyte-associated antigen-4 and "CTLA-4,” are synonomous with one another, and include variants, isoforms, species homologs of human CTLA-4, and analogs having at least one common epitope with CTLA-4 (see, e.g., Balzano (1992) Int. J. Cancer Suppl. 7:28-32).
  • CTLA-4 The complete cDNA sequence of human CTLA-4 has the Genbank accession number Ll 5006.
  • the region of amino acids 1-37 is the leader peptide; 38-161 is the extracellular V-like domain; 162-187 is the transmembrane domain; and 188-223 is the cytoplasmic domain.
  • Variants of the nucleotide sequence have been reported, including a G to A transition at position 49, a C to T transition at position 272, and an A to G transition at position 439.
  • the complete DNA sequence of mouse CTLA-4 has the EMBL accession number X05719 (Brunet et al. (1987) Nature 328:267-270).
  • the region of amino acids 1-35 is the leader peptide.
  • An intact "antibody” comprises at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds.
  • Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as HCVR or VH) and a heavy chain constant region.
  • the heavy chain constant region is comprised of three domains, CHl, CH2 and CH3.
  • Each light chain is comprised of a light chain variable region (abbreviated herein as LCVR or VL) and a light chain constant region.
  • the light chain constant region is comprised of one domain, CL.
  • VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR).
  • CDR complementarity determining regions
  • FR framework regions
  • Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxyl-terminus in the following order: FRl, CDRl, FR2, CDR2, FR3, CDR3, FR4.
  • the variable regions of the heavy and light chains contain a binding domain that interacts with an antigen.
  • the constant regions of the antibodies may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (CIq) of the classical complement system.
  • antibody includes antigen-binding portions of an intact antibody that retain capacity to bind CTLA-4.
  • binding include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CHl domains; (ii) a F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting of the VH and CHl domains; (iv) a Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment (Ward et al., (1989) Nature 341 :544-546), which consists of a VH domain; and (vi) an isolated complementarity determining region (CDR).
  • CDR complementarity determining region
  • the two domains of the Fv fragment, VL and VH are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv (scFv); See, e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883).
  • single chain Fv single chain Fv
  • Such single chain antibodies are included by reference to the term "antibody” Fragments can be prepared by recombinant techniques or enzymatic or chemical cleavage of intact antibodies.
  • Specific binding refers to antibody binding to a predetermined antigen.
  • the phrase “specifically (or selectively) binds” to an antibody refers to a binding reaction that is determinative of the presence of the protein in a heterogeneous population of proteins and other biologies.
  • the antibody binds with an association constant (K.sub.a) of at least about IxIO 6 M '1 or 10 7 M “1 , or about 10 8 M “1 to 10 9 M “1 , or about 10 10 M “ 'to 10 11 IVf 1 Or higher, and binds to the predetermined antigen with an affinity that is at least two-fold greater than its affinity for binding to a non-specific antigen (e.g., BSA, casein) other than the predetermined antigen or a closely-related antigen.
  • a non-specific antigen e.g., BSA, casein
  • reversal of an established tumor or grammatical equivalents herein is meant the suppression, regression, or partial or complete disappearance of a pre-existing tumor. The definition is meant to include any diminution in the size, potency or growth rate of a pre-existing tumor.
  • treatment shall refer to any and all uses of the claimed compositions which remedy a disease state or symptom, or otherwise prevent, hinder, retard, or reverse the progression of disease or other undesirable symptoms in any way whatsoever.
  • administered refers to any method that introduces the cells of the invention (e.g. cancer vaccine) to a mammal. This includes, but is not limited to, intradermal, parenteral, intramuscular, subcutaneous, intraperitoneal, intranasal, intravenous (including via an indwelling catheter), intratumoral, via an afferent lymph vessel, or by another route that is suitable in view of the patient's condition.
  • the compositions of this invention may be administered to the subject at any site. For example, they can be delivered to a site that is "distal” to or “distant” from the primary tumor.
  • the term "increased immune response" as used herein means that a detectable increase of a specific immune activation is detectable (e.g. an increase in B-cell and/or T-cell response).
  • An example of an increased immune response is an increase in the amount of an antibody that binds an antigen which is not detected or is detected a lower level prior to administration of a cytokine-expressing cellular vaccine of the invention.
  • Another example is an increased cellular immune response.
  • a cellular immune response involves T cells, and can be observed in vitro (e.g. measured by a Chromium release assay) or in vivo.
  • An increased immune response is typically accompanied by an increase of a specific population of immune cells.
  • tumor growth refers to any measurable decrease in tumor mass, tumor volume, amount of tumor cells or growth rate of the tumor. Measurable decreases in tumor mass can be detected by numerous methods known to those skilled in the art. These include direct measurement of accessible tumors, counting of tumor cells (e.g. present in blood), measurements of tumor antigens (e.g. Prostate Specific Antigen (PSA), Alphafetoprotein (AFP) and various visualization techniques (e.g.
  • terapéuticaally effective amount refers to an amount of an agent, e.g., a cytokine-expressing cellular vaccine of the invention, that is sufficient to modulate, either by stimulation or suppression, the immune response of an individual. This amount may be different for different individuals, different tumor types, and different preparations.
  • the "therapeutically effective amount” is determined using procedures routinely employed by those of skill in the art such that an "improved therapeutic outcome" results.
  • nucleic acid refers to deoxyribonucleotides or ribonucleotides and polymers thereof ("polynucleotides”) in either single- or double-stranded form.
  • nucleic acids containing known analogues of natural nucleotides that have similar binding properties as the reference nucleic acid and are metabolized in a manner similar to naturally occurring nucleotides.
  • a particular nucleic acid molecule/polynucleotide also implicitly encompasses conservatively modified variants thereof (e.g. degenerate codon substitutions) and complementary sequences and as well as the sequence explicitly indicated.
  • degenerate codon substitutions may be achieved by generating sequences in which the third position of one or more selected (or all) codons is substituted with mixed-base and/or deoxyinosine residues (Batzer et al., Nucleic Acid Res.
  • Nucleotides are indicated by their bases by the following standard abbreviations: adenine (A), cytosine (C), thymine (T), and guanine (G).
  • “Stringent hybridization conditions” and “stringent wash conditions” in the context of nucleic acid hybridization experiments such as Southern and Northern hybridizations are sequence dependent, and are different under different environmental parameters. Longer sequences hybridize at higher temperatures.
  • the T m is the temperature (under defined ionic strength and pH) at which 50% of the target sequence hybridizes to a perfectly matched probe.
  • Very stringent conditions are selected to be equal to the T n , for a particular probe.
  • An example of stringent hybridization conditions for hybridization of complementary nucleic acids that have more than 100 complementary residues on a filter in a Southern or northern blot is 50% formamide with 1 mg of heparin at 42° C, with the hybridization being carried out overnight.
  • An example of highly stringent wash conditions is 0.1 5M NaCl at 72° C for about 15 minutes.
  • An example of stringent wash conditions is a 0.2 x SSC wash at 65° C for 15 minutes ⁇ see, Sambrook, infra, for a description of SSC buffer). Often, a high stringency wash is preceded by a low stringency wash to remove background probe signal.
  • An example medium stringency wash for a duplex of, e.g., more than 100 nucleotides, is 1 x SSC at 45° C for 15 minutes.
  • An example low stringency wash for a duplex of, e.g., more than 100 nucleotides, is 4-6 x SSC at 40° C for 15 minutes.
  • stringent conditions typically involve salt concentrations of less than about 1.0M Na ion, typically about 0.01 to 1.0 M Na ion concentration (or other salts) at pH 7.0 to 8.3, and the temperature is typically at least about 30° C.
  • Stringent conditions can also be achieved with the addition of destabilizing agents such as formamide.
  • a signal to noise ratio of 2 x (or higher) than that observed for an unrelated probe in the particular hybridization assay indicates detection of a specific hybridization
  • the terms "identical” or percent “identity” in the context of two or more nucleic acid or protein sequences refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues or nucleotides that are the same, when compared and aligned for maximum correspondence, as measured using one of the sequence comparison algorithms described herein or by visual inspection
  • sequence comparison typically one sequence acts as a reference sequence to which test sequences are compared.
  • test and reference sequences are input into a computer, subsequence coordinates are designated if necessary, and sequence algorithm program parameters are designated.
  • sequence comparison algorithm then calculates the percent sequence identity for the test sequence(s) relative to the reference sequence, based on the designated program parameters.
  • Optimal alignment of sequences for comparison can be conducted, e.g., by the local homology algorithm of Smith & Waterman, Adv. Appl. Math. 2:482 (1981), by the homology alignment algorithm of Needleman & Wunsch, J. MoI. Biol. 48: 443 (1970), by the search for similarity method of Pearson & Lipman, Proc. Nat'l. Acad. Sci. USA 85:2444 (1988), by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, Wis.), by the BLAST algorithm, Altschul et al., J. MoI. Biol.
  • a "peptide” refers to an amino acid polymer containing between about 8 and about 12 amino acids linked together via peptide bonds.
  • a peptide according to the present invention can comprise additional atoms beyond those of the 8 to twelve amino acids, so long as the peptide retains the ability to bind an MHC I receptor, e.g., an HLA-A2 receptor, and form a ternary complex with the T-cell receptor, the MHC I receptor, and the peptide.
  • Conservative substitution refers to the substitution in a polypeptide of an amino acid with a functionally similar amino acid.
  • the following six groups each contain amino acids that are conservative substitutions for one another:
  • the term “about 5 ⁇ g/kg” means a range of from 4.5 ⁇ g/kg to 5.5 ⁇ g/kg.
  • “about 1 hour” means a range of from 48 minutes to 72 minutes.
  • the modified value should be rounded to the nearest whole number.
  • “about 12 amino acids” means a range of 11 to 13 amino acids.
  • physiological conditions refers to the salt concentrations normally observed in human serum.
  • physiological conditions need not mirror the exact proportions of all ions found in human serum, rather, considerable adjustment can be made in the exact concentration of sodium, potassium, calcium, chloride, and other ions, while the overall ionic strength of the solution remains constant.
  • the invention provides methods that comprise assessing immune responses against antigens associated with a likelihood of responsiveness to treatment with the combination of proliferation- incompetent tumor cells that express cytokines, e.g., GM-CSF, and an anti-CLTA-4 antibody.
  • the combination therapies are predicted to result in an improved therapeutic outcome for the subject, for example, a reduction in the level of PSA in the patient's serum, a decrease in cancer-associated pain or improvement in the condition of the patient according to any clinically acceptable criteria, including but not limited to a decrease in metastases, an increase in life expectancy or an improvement in quality of life.
  • the antigens may be expressed endogenously by cells native to the subject or may be exogenously provided to the subject by, e.g., the administered engineered tumor cells.
  • PSMD2 proteasome 26S subunit, non-ATPase, 2
  • S2P97, TRAP2, MGC 14274 is a 100200 Da protein of 908 amino acids (SEQ ID NO: 1) encoded on chromosome 3 (Ensembl cytogenetic band: 3q27.1).
  • a representative nucleotide sequence is NM_002808 (SEQ ID NO: 2).
  • PSMD2 acts as a regulatory subunit of the 26 proteasome which is involved in the ATP-dependent degradation of ubiquitinated proteins (Coux et al. Annu. Rev. Biochem. 65, 801-847 (1996).
  • An essential function of a modified proteasome, the immunoproteasome, is the processing of class I MHC peptides.
  • this subunit may also participate in the TNF signalling pathway since it interacts with the tumor necrosis factor type 1 receptor. See e.g., Tsunami et al, Eur. J. Biochem. 239 (3), 912-921 (1996); and Song et al, Biochem. J. 309 (PT 3), 825- 829 (1995).
  • Interphase cytoplasmic protein 45 (alias tRNA-histidine guanylyltransferase 1 -like (THGlL), FLJl 1601, FLJ20546) is a 20157 Da protein of 173 amino acids (SEQ ID NO: 3) encoded on chromosome 5 (Ensembl cytogenetic band: 5q33.3).
  • a representative nucleotide sequence is NM_017872 (SEQ ID NO: 4).
  • ICF45 is a highly conserved protein, which is expressed in a cell cycle-dependent manner and appears to be involved in cell cycle progression and cell proliferation. See e.g., Guo et al, J. Biol. Chem.
  • Aldolase A fructose-bisphosphate (ALDOA) (alias aldolase A, fructose- bisphosphate, ALDA 5 MGC 10942, MGC 17716, MGC 17767) encodes aldolase A (amino acid reference sequence NP_000025; SEQ ID NO: 5), a 39289 Da protein of 363 amino acids) on chromosome 16 (Ensembl cytogenetic band: 16pl 1.2).
  • a representative nucleotide sequence is NM_000034 (SEQ ID NO:6).
  • Aldolase A (fructose-bisphosphate aldolase) is a glycolytic enzyme that catalyzes the reversible conversion of fructose- 1,6-bisphosphate to glyceraldehyde 3-phosphate and dihydroxyacetone phosphate (Esposito et al, Biochem. J. 380 (PT 1), 51-56 (2004).
  • Three aldolase isozymes (A, B, and C), encoded by three different genes, are differentially expressed during development. Aldolase A is found in the developing embryo and is produced in even greater amounts in adult muscle.
  • Aldolase A expression is repressed in adult liver, kidney and intestine and similar to aldolase C levels in brain and other nervous tissue (Rehbein-Thoner and Pfleiderer 1977). Aldolase A deficiency has been associated with myopathy and hemolytic anemia. Alternative splicing of this gene results in multiple transcript variants which encode the same protein. See Hoppe-Seyler, Physiol. Chem. 358 (2), 169-180 (1977).
  • Cyclin E2 (CCNE2) (alias: cyclin E2, CYCE2) (representative amino acid sequence NP_477097; SEQ ID NO:7) is a 46757 Da protein of 404 amino acids encoded on chromosome 8 (Ensembl cytogenetic band: 8q22.1). A representative nucleotide sequence is NM 057749 (SEQ ID NO:8).
  • CCNE2 belongs to the highly conserved cyclin family, whose members are characterized by a dramatic periodicity in protein abundance through the cell cycle. Cyclins function as regulators of cyclin dependent kinases.
  • CCNE2 forms a complex with and functions as a regulatory subunit of cyclin dependent kinase 2 (CDK2) and plays a role in cell cycle Gl /S transition (Zariwala et al, Oncogene 17 (21), 2787-2798 (1998)).
  • CDK2 cyclin dependent kinase 2
  • the expression of this gene peaks at the Gl-S phase and exhibits a pattern of tissue specificity distinct from that of cyclin El (Lauper, et al, Oncogene 17 (20), 2637-2643 (1998)). Two transcript variants encoding different isoforms have been found for this gene.
  • Dihydrolipoamide S-acetyltransferase (alias: DLTA, PDCE2, PDC-E2614) is a 65781 Da protein of 614 amino acids (SEQ ID NO: 9) encoded on chromosome 11 (Entrez Gene cytogenetic band: I lq23.1). A representative nucleotide sequence is NM_001931 (SEQ ID NO: 10).
  • the DLAT gene encodes dihydrolipoamide acetyltransferase, the E2 subunit of the mammalian pyruvate dehydrogenase complex (PDC) of the inner mitochondrial membrane.
  • PDC mammalian pyruvate dehydrogenase complex
  • the pyruvate dehydrogenase complex catalyzes the overall conversion of pyruvate to acetyl-CoA and CO(2). It contains multiple copies of three enzymatic components: pyruvate dehydrogenase (El), dihydrolipoamide acetyltransferase (E2) and lipoamide dehydrogenase. See Hiromasa et al, J. Biol. Chem. 279 (8), 6921-6933 (2004).
  • GHR Growth hormone receptor
  • GHBP growth hormone receptor
  • NP_000154 reference amino acid sequence NP_000154; SEQ ID NO:11
  • NP_000154 reference amino acid sequence NP_000154; SEQ ID NO:11
  • NP_000154 reference amino acid sequence NP_000154; SEQ ID NO:11
  • NP_000154 reference amino acid sequence NP_000154; SEQ ID NO:11
  • NM_000163 SEQ ID NO:12
  • the GHR gene encodes a protein that is a transmembrane receptor for growth hormone. Binding of growth hormone to the receptor leads to receptor dimerization and the activation of an intra- and intercellular signal transduction pathway leading to growth (Jorgensen et al, Am. J. Physiol Endocrinol. Metab.
  • High density lipoprotein binding protein (vigilin), (HDLBP) (alias, HBP, VGL, FLJ16432) (representative amino acid sequence NP_005327; SEQ ID NO: 13) is a 141440 Da protein of 1268 amino acids on chromosome 2 (Ensembl cytogenetic band: 2q37.3). A representative nucleotide sequence is NM_005336 (SEQ ID NO: 14).
  • High density lipoprotein-binding protein also known as vigilin, is a 110-kD protein that specifically binds HDL molecules and may function in the removal of excess cellular cholesterol. See McKnight, eg., J. Biol. Chem.
  • KIAA0692 (alias FLJ22280, FLJ36132) encodes protein KIAA0692 (reference sequence Q86XL3; SEQ ID NO: 15), a 104165 Da protein of 938 amino acids on chromosome 12 (Ensembl cytogenetic band: 12q24.33).
  • a representative nucleotide sequence is XM_925991 (SEQ ID NO: 16).
  • PHD finger protein 17 (alias PHD finger protein 17; synonyms: JADEl, FLJ22479, KIAA 1807) encodes PHD finger protein 17 (reference amino acid sequence NP_955352; SEQ ID NO: 17), a 79568 Da protein of 702 amino acids on chromosome 4 (Ensembl cytogenetic band: 4q28.2).
  • a representative nucleotide sequence is NM_199320 (SEQ ID NO: 18).
  • PHFl 7 encodes a short-lived candidate transcription factor with PEST (proline, glutamate, serine, and threonine) protein degradation and plant homeodomain motifs (Zhou et al, J. Biol. Chem. 277, 39887- 39898; Panchenko et al., . J. Biol. Chem. 279, 56032-5604 (2004)).
  • Jade-1 is the first member of a small family of proteins (Tzouanacou et al. Development. MoI. Cell. Biol. 23, 8553-8562; and Zhou et al, Cancer Res. 64, 1278-1286).
  • Jade-1 is a strong pVHL interactor that is also stabilized by pVHL and may represent a novel candidate regulatory factor in Von Hippel Lindau-mediated renal tumor suppression.
  • RANBP2-like and GRIP domain containing 5 (RGPD5) (alias: RGP5, BS- 63, DKFZp686I1842) encodes the protein RANBP2-like and GRIP domain containing 5 (representative amino acid sequence NP_005045; SEQ ID NO: 19) a 198740 Da protein of 1765 amino acids on chromosome 2 (Ensembl cytogenetic band: 2ql3).
  • a representative nucleotide sequence is NM_005054 (SEQ ID NO:20).
  • RGPD5 is a small GTP-binding protein of the RAS superfamily that is associated with the nuclear membrane and is thought to control a variety of cellular functions through its interactions with other proteins (Cai et al, MoI. Reprod. Dev. 61 (1), 126- 134 (2002)). This gene shares a high degree of sequence identity with RANBP2, a large RAN-binding protein localized at the cytoplasmic side of the nuclear pore complex. Alternative splicing has been observed for this locus and two variants are described. Additional splicing is suggested but complete sequence for further transcripts has not been determined. (Wang et al, Arch. Androl. 42 (2), 71-84 (1999); Nothwang et al, Genomics 47 (3), 383-392 (1998)).
  • Ubiquitin specific peptidase 25 (alias: USP21) encodes Ubiquitin specific peptidase 25 (representative amino acid sequence NP_037528; SEQ ID NO:21), a 125750 Da protein of 1056 amino acids on chromosome 21 (Ensembl cytogenetic band: 21q21.1). A representative nucleotide sequence is NM_013396 (SEQ ID NO: 22). USP25 produces three protein isoforms by alternative splicing. While two of the isoforms are expressed nearly ubiquituously, the expression of the longer USP25 isoform (USP25m) is restricted to muscular tissues and is upregulated during myogenesis.
  • USP25m interacts with three sarcomeric proteins: actin alpha- 1 (ACTAl), filamin C (FLNC), and myosin binding protein Cl (MyBPCl), which are critically involved in muscle differentiation and maintenance, and have been implicated in the pathogenesis of severe myopathies.
  • actin alpha- 1 ACTAl
  • FLNC filamin C
  • MyBPCl myosin binding protein Cl
  • PDZ domain containing 8 (PDZD8) (alias: PDZK8, FLJ25412) encodes PDZ domain containing 8 (reference sequence NP_776152; SEQ ID NO:23), a 128563 Da protein of 1154 amino acids encoded on chromosome 10 (Ensembl cytogenetic band: 10q26.1 1).
  • a representative nucleotide sequence is NM_173791 (SEQ ID NO:24).
  • PDZD8 was identified in a multiinstitutional effort to identify and sequence a cDNA clone containing a complete ORF for each human and mouse gene.
  • ESTs were generated from libraries enriched for full-length cDNAs and analyzed to identify candidate full-ORF clones, which then were sequenced to high accuracy (See Strausberg et al, Proc Natl Acad Sci USA. 2002 Dec 24;99(26): 16899-903).
  • PDZD8 represents one of these clones.
  • Triosephosphate isomerase 1 (TPIl) (alias ITPI, MGC88108) encodes triosephosphate isomerase 1 (representative amino acid sequence NP_000356; SEQ ID NO:25), a 26538 Da protein of 248 amino acids on chromosome 12 (Ensembl cytogenetic band: 12pl3.31). A representative nucleotide sequence is NM_000365 (SEQ ID NO:26). TPI protein is responsible for the conversion of dihydroxyacetone phosphate into glyceraldehyde-3 -phosphate in glycolysis.
  • TPI is considered a near perfect enzyme due to its catalytic efficiency; the rate of catalysis is diffusion controlled, suggesting the presence of strong selective pressure throughout the gene's evolution .
  • TPI is known to exist functionally as a homodimer, and the dimer interaction sites have been well characterized.
  • several mutations have been reported that result in TPI deficiency, a progressive disease that eventuates in neuromuscular failure, hemolytic anemia, increased susceptibility to infection, and premature death. See Daar et al, Proc. Natl. Acad. Sci. U.S.A. 83 (20), 7903-7907 (1986); and Maquat et al, J. Biol. Chem. 260 (6), 3748-3753 (1985).
  • Glutamic-oxaloacetic transaminase 2 (alias FLJ40994) encodes aspartate aminotransferase 2 precursor (representative amino acid sequence NP_002071 ; SEQ ID NO:27), a 47476 Da of 430 amino acids on chromosome 16 (Ensembl cytogenetic band: 16q21).
  • a representative nucleotide sequence is NM_002080 (SEQ ID NO:28).
  • Glutamic-oxaloacetic transaminase is a pyridoxal phosphate-dependent enzyme which exists in cytoplasmic and inner-membrane mitochondrial forms, GOTl and GOT2, respectively.
  • GOT plays a role in amino acid metabolism and the urea and tricarboxylic acid cycles.
  • the two enzymes are homodimeric and show close homology. See Lain et al, J. Biol. Chem. 270 (42), 24732-24739 (1995); Pol et al, Hum. Genet. 83 (2), 159-164 (1989); and Pol et al, Biophys. Res. Commun. 157 (3), 1309-1315 (1988).
  • CNDP dipeptidase 2 (metallopeptidase M20 family) (CNDP2) (alias CN2, CPGL, PEPA, HsT2298, FLJl 0830) encodes Cytosolic nonspecific dipeptidase (reference amino acid sequence Q96KP4; SEQ ID NO:29), a 52878Da protein of 475 amino acids on chromosome 18 (Ensembl cytogenetic band: 18q22.3).
  • a representative nucleotide sequence is NM_018235 (SEQ ID NO:30).
  • CNDP2 also known as tissue carnosinase and peptidase A (EC 3.4.13.18), is a nonspecific dipeptidase rather than a selective carnosinase. See Teufel et al., J. Biol. Chem. 278 (8), 6521-6531 (2003).
  • GPI-anchored membrane protein 1 (GPIAP 1 ) (alias MI lSl ,GPIP 137, pl37GPI) encodes membrane component chromosome 11 surface marker 1 isoform 1 (representative amino acid sequence NP 005889; SEQ ID NO:31), a 79477 Da protein of 709 amino acids on chromosome 11 (Ensembl cytogenetic band: 1 IpI 3).
  • a representative nucleotide sequence is NM_005898 (SEQ ID NO:32).
  • GPIAPl belongs to a novel family of proteins that are highly conserved in vertebrates (Wang et al, J. Immunol. 175 (7), 4274-4282 (2005)).
  • GPIAPl may serve a different function
  • levels of GPIAPl correlate with cellular proliferation, with high levels occurring in thymus and spleen, and low levels in kidney, liver, and muscle.
  • Levels of GPIAPl proteins increased when resting splenic T or B lymphocytes were stimulated to divide. See Grill et al, J. Immunol. 172 (4), 2389- 2400 (2004).
  • Apoptosis inhibitor 5 (alias AACl 1, AAC-11, API5L1) encodes apoptosis inhibitor 5 (representative amino acid sequence NP_006586; SEQ ID NO:33) a 57561 Da protein of 510 amino acids on chromosome 11 (Ensembl cytogenetic band: 1 IpI 2).
  • a representative nucleotide sequence is NM_006595 (SEQ ID NO:34).
  • API5 was initially isolated as a gene whose expression promoted cell survival following serum deprivation. Studies have shown that the API5 mRNA transcript is strongly expressed in transformed cell lines (Tewari et al, Cancer Res 57: 4063-4069 (1997)).
  • Orthologs of the API5 gene family are highly conserved in species as diverse as plants and humans, but there are no obvious family members in worms or yeast.
  • Api5 proteins share a number of conserved domains including a putative transactivation-domain flanked by two acidic domains, an LxxLL motif, a putative leucine zipper domain, and a nuclear localization sequence. The presence of these motifs suggests that Api5 family proteins might be transcriptional regulators.
  • Various deletion mutants of Api5 possess strong transactivation activity when fused to the DNA binding domain of Gal4. See Van den Berghe et al MoI Endocrinol 14: 1709-1724 (2000)).
  • PHD finger protein 14 (alias KIAA0783) encodes PHD finger protein 14 (representative amino acid sequence NP_001007158; SEQ ID NO: 35) a 106843 Da protein of 948 amino acids on chromosome 7 (Ensembl cytogenetic band: 7p21.3). A representative nucleotide sequence is NM_001007157 (SEQ ID NO:36). PHF 14 was identified in a large-scale analysis of HeLa nuclear phosphoproteins (Beausoleil et al, Proc. Natl. Acad. ScL U.S.A. 101 (33), 12130-12135 (2004)).
  • PHF 14 contiains a PHD finger domain; a common structural motif found in all eukaryotic genomes. It is a Zn(2+)-binding domain that binds to specific nuclear protein partners, apparently through the same surface that is used by RING domains to bind their cognate E2 ligases. New evidence also suggests that some PHD fingers bind to nucleosomes, raising the possibility that chromatin might be a common nuclear ligand of PHD fingers.
  • Ferritin, heavy polypeptide 1 (alias FTH, PLIF, FTHL6, PIGl 5, MGC 104426) encodes ferritin, heavy polypeptide 1 (representative protein sequence NP_002023; SEQ ID NO:37) a 21094 Da protein of 182 amino acids encoded on chromosome 11 (Ensembl cytogenetic band: I lql2.2).
  • a representative nucleotide sequence is NM_002032(SEQ ID NO:38).
  • FTHl encodes the heavy subunit of ferritin, the major intracellular iron storage protein in prokaryotes and eukaryotes.
  • ferritin subunit composition may affect the rates of iron uptake and release in different tissues.
  • a major function of ferritin is the storage of iron in a soluble and nontoxic state. Defects in ferritin proteins are associated with several neurodegenerative diseases. This gene has multiple pseudogenes. Several alternatively spliced transcript variants have been observed, but their biological validity has not been determined (Hentze et al, Proc. Natl. Acad. ScL U.S.A. 83 (19), 7226-7230 (1986)).
  • Microtubule-associated protein 4 (alias MGC8617, DKFZp779A1753) encodes microtubule-associated protein 4 (representative amino acid sequence NP_002366; SEQ ID NO:39), a 121019 Da protein of 1152 amino acids on chromosome 3 (Ensembl cytogenetic band: 3p21.31).
  • a representative nucleotide sequence is NM_002375 (SEQ ID NO:40).
  • MAP4 is a major non- neuronal microtubule-associated protein which contains a domain similar to the microtubule-binding domains of neuronal microtubule-associated protein (MAP2) and microtubule-associated protein tau (MAPT/TAU; Chapin, et al., J. Cell. ScL 98 (PT 1), 27-36 (1991)).
  • MAP4 promotes microtubule assembly, and has been shown to counteract destabilization of interphase microtubule catastrophe promotion. Cyclin B was found to interact with this protein, which targets cell division cycle 2 (CDC2) kinase to microtubules (Ookata et al, J. Cell Biol.
  • KIAA0406 encodes hypothetical protein LOC9675 (representative amino acid sequence NP_055472; SEQ ID NO:41) a 122069 Da protein of 1089 aa encoded on chromosome 20 (Ensembl cytogenetic band: 20ql 1.23).
  • LOC9675 representsative amino acid sequence NP_055472; SEQ ID NO:41
  • a 122069 Da protein of 1089 aa encoded on chromosome 20 Ensembl cytogenetic band: 20ql 1.23.
  • a representative nucleotide sequence is NM_014657 (SEQ ID NO:42).
  • KIAA0406 was identified in 2 large-scale studies as a hypothetical coding region/protein. See Strausberg et al., Proc. Natl. Acad. ScL U.S.A. 99 (26), 16899-16903 (2002); and Ishikawa et al, DNA Res. 4 (5), 307-313 (1997).
  • SlOO calcium binding protein Al 1 (calgizzarin) (SlOOAl 1) (alias MLN70, SlOOC) encodes SlOO calcium binding protein Al 1 (reference protein sequence NP_005611; SEQ ID NO: 43), a 11740 Da protein of 105 amino acids encoded on chromosome 1 (Ensembl cytogenetic band: Iq21.3).
  • a representative nucleotide sequence is NM_005620 (SEQ ID NO:44).
  • the protein encoded by this gene is a member of the SlOO family of proteins containing 2 EF-hand calcium-binding motifs.
  • SlOO proteins are localized in the cytoplasm and/or nucleus of a wide range of cells, and involved in the regulation of a number of cellular processes such as cell cycle progression and differentiation(Wicki et al, Cell Calcium 20 (6), 459-464 (1996); Mailliard et al, J. Biol. Chem. 271 (2), 719-725 (1996)).
  • SlOO genes include at least 13 members which are located as a cluster on chromosome Iq21 (Schafer et al, Genomics 25 (3), 638-643 (1995)). This protein may function in motility, invasion, and tubulin polymerization.
  • Hypothetical protein (DKFZP686A01247) (alias MGC72127,DKFZp434I0312, DKFZp686B2470, DKFZp686G2094, DKFZp781C1754, DKFZp781I1455, DKFZp686G 18243) encodes hypothetical protein LOC22998 (representative amino acid sequence NP_055803; SEQ ID NO:45), a 122467 Da protein of 1089 amino acids encoded on chromosome 4 (Ensembl cytogenetic band: 4pl4).
  • a representative nucleotide coding sequence is NM_014988 (SEQ ID NO: 46).
  • DKFZP686A01247 was identified in large-scale gene identification studies as a hypothetical coding region/protein (Beausoleil et al, Proc. Natl Acad. Sci. U.S.A. 101 (33), 12130-12135 (2004); Simpson et al, EMBO Rep. 1 (3), 287-292 (2000)).
  • SET domain containing 3 (alias FLJ23027, MGC87236, C14orfl54, DKFZp761E1415) encodes SET domain containing 3 (reference amino acid sequence NP_115609; SEQ ID NO:47) a 67913 Da protein of 595 amino acids on chromosome 14 (Ensembl cytogenetic band: 14q32.2).
  • a representative nucleotide sequence is NM_032233 (SEQ ID NO:48).
  • SETD3 was identified in large-scale gene identification studies. See Tao et al, Nat. Methods 2 (8), 591-598 (2005); and Harrington et al, Nat. Biotechnol. 19 (5), 440-445 (2001).
  • NudC domain containing 3 NUDCD3 (alias: NudCL,KIAAl 068) encodes NudC domain containing 3 (reference amino acid NP_056147; SEQ ID NO:49), a 24958 Da protein of 221 amino acids encoded on chromosome 7 (Ensembl cytogenetic band: 7pl3).
  • a representative nucleotide coding sequence is NM_015332 (SEQ ID NO:50).
  • NUDCD3 appears to influence the stabilization of dynein intermediate chain. Cytoplasmic dynein, a minus-end-directed microtubule motor, has been implicated in many fundamental cellular processes. The expression and phosphorylation of NudCL are increased during mitosis.
  • PCYTlB phosphate cytidylyltransferase 1, choline, beta encodes phosphate cytidylyltransferase 1 , choline, beta (reference amino acid sequence NP_004836; SEQ ID NO:51) a 41940 Da protein of 369 amino acids on chromosome 22 (Ensembl cytogenetic band: Xp22.11).
  • a reference nucleotide coding sequence is NM_004845 (SEQ ID NO:52).
  • CCT Phosphocholine cytidylyltransferase 1
  • beta is a key regulator of phosphatidylcholine biosynthesis along with CCT-alpha.
  • CCTbeta transcripts have been detected in human adult and fetal tissues, and very high transcript levels were found in placenta and testis. The amino terminus of CCTbeta bears no resemblance to the amino terminus of CCTalpha, and CCTbeta protein was localized to the cytoplasm as detected by indirect immunofluorescent microscopy. See Lykidis et al, J. Biol. Chem. 273 (22), 14022-14029 (1998).
  • the antigens of the invention find use in a variety of methods, including methods for determining whether an immune response against cancer cells has been induced in a subject, methods for determining whether an immune response effective to treat, prevent, or ameliorate a symptom of prostate cancer in a subject has been induced in the subject, methods for determining whether a subject afflicted with prostate cancer is likely to respond to treatment with genetically modified tumor cells that produce GM-CSF, and methods for assessing the effectiveness of prostate cancer therapy with a composition comprising an antibody that specifically binds to human CTLA-4 and genetically modified tumor cells that express GM-CSF, to treat or ameliorate a symptom of prostate cancer of a subject in need thereof.
  • the invention provides a method for determining whether an immune response effective to treat, prevent, or ameliorate a symptom of prostate cancer in a subject has been induced in the subject, comprising detecting an immune response against an antigen listed in Table 1 or 2, detecting said antigen indicates that an immune response effective to treat, prevent, or ameliorate a symptom of prostate cancer has been induced in the subject.
  • an immune response against 1, 2, 3 ,4 5, 6, 7, 8, 9,10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, or more of the antigens is detected.
  • the immune response that has been induced is . effective to prevent prostate cancer in the subject. In certain embodiments, the immune response that has been induced is effective to treat prostate cancer in the subject. In certain embodiments, the immune response that has been induced is effective to ameliorate a symptom of prostate cancer in the subject. In certain embodiments, the symptom of prostate cancer that is ameliorated is selected from the group consisting of a reduction in the level of prostate specific antigen (PSA) level in the subject's serum, cancer-associated pain, and metastasis.
  • PSA prostate specific antigen
  • the immune response is effective to result in decreased serum concentrations of tumor specific markers, increased overall survival time, increased progression-free survival, decreased tumor size, decreased bone metastasis marker response, increased impact on minimal residual disease, increased induction of antibody response to the cancer cells that have been rendered proliferation-incompetent, increased induction of delayed-type-hypersensitivity (DTH) response to injections of autologous tumor, increased induction of T cell response to autologous tumor or candidate tumor- associated antigens, or increased impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response.
  • DTH delayed-type-hypersensitivity
  • T cell response to autologous tumor or candidate tumor- associated antigens
  • the immune response is detected by western blot.
  • the immune response is detected by ELISA.
  • the immune response is detected by protein array analysis.
  • Clinical datasets of immune responses with clinical outcome data can be used to correlate immune responses with the likelihood of responding to cancer therapy or with responsiveness to cancer therapy.
  • Any method known in the art can be used to assess the immune response of a subject administered a cancer therapy, e.g., a combined cell- based/antibody cancer immunotherapy such as, e.g., GV AX® + anti-CTLA-4 therapy.
  • a cancer therapy e.g., a combined cell- based/antibody cancer immunotherapy such as, e.g., GV AX® + anti-CTLA-4 therapy.
  • immune responses can be assessed by western blot, by ELISA, by protein array analysis, and the like.
  • any method known in the art can be used to determine whether an immune response is correlated with responsiveness to the cancer therapy.
  • P values are used to determine the statistical significance of the correlation, such that the smaller the P value, the more significant the measurement.
  • the P values will be less than 0.05 (or 5%). More preferably, P values will be less than 0.01.
  • P values can be calculated by any means known to one of skill in the art. For the purposes of correlating an immune response with responsiveness to cancer therapy, P values can be calculated using Fisher's Exact Test. See, e.g., David Freedman, Robert Pisani & Roger Purves, 1980, STATISTICS, W. W. Norton, New York.
  • immune responses are measured from biological samples obtained from a subject.
  • Biological samples from a subject include, for example and without limitation, blood, blood plasma, serum, urine, saliva, tissue swab and the like.
  • the present invention provides a method of constructing an algorithm that correlates immune response data with responsiveness to cancer therapy, e.g., a combined cell-based/antibody cancer immunotherapy such as, e.g., GV AX® + anti-CTLA-4 therapy.
  • the method of constructing the algorithm comprises creating a rule or rules that correlate immune response data with responsiveness to cancer therapy, e.g., a combined cell-based/antibody cancer immunotherapy such as, e.g., GV AX® + anti-CTLA-4 therapy.
  • a data set comprising immune response data and clinical outcome data about each subject in a set of subjects is assembled. Any method known in the art can be used to collect immune response data. Examples of methods of collecting such data are provided above. Any method known in the art can be used for collecting clinical outcome data.
  • the data set comprises immune responses against one or more antigens as described herein. In some embodiments, the data set comprises immune responses against 1, 2, 3 ,4 5, 6, 7, 8, 9,10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, or more antigens.
  • the clinical outcome data comprises information regarding the level of prostate specific antigen (PSA) level in the subject's serum, cancer-associated pain, and/or metastasis.
  • the clinical outcome data comprises information regarding the serum concentrations of tumor specific markers, e.g., PSA, overall survival time, progression-free survival, tumor size, bone metastasis marker response, impact on minimal residual disease, induction of antibody response to the cancer cells that have been rendered proliferation- incompetent, induction of delayed-type-hypersensitivity (DTH) response to injections of autologous tumor, induction of T cell response to autologous tumor or candidate tumor-associated antigens, and/or impact on circulating T cell and dendritic cell numbers, phenotype, and function, cytokine response
  • DTH delayed-type-hypersensitivity
  • the immune response and clinical outcome data in the data set can be represented or organized in any way known in the art.
  • the data are displayed in the form of a graph.
  • the immune response and clinical outcome data in the data set are displayed in the form of a chart.
  • an algorithm is formulated that correlates the immune response with the clinical outcome data in the data set.
  • a clinical outcome cutoff point is defined.
  • the clinical outcome cutoff point is determined relative to a reference subject, and the cutoff point is the value above or below which a subject is defined as responsive to the cancer therapy and below or above which a virus or population of viruses is defined nonresponsive to the cancer therapy.
  • a clinical outcome cutoff point is defined.
  • the clinical outcome cutoff point is determined relative to a reference subject, and the cutoff point is the value above or below which a subject is defined as responsive to the cancer therapy and below or above which a virus or population of viruses is defined nonresponsive to the cancer therapy.
  • an increase in the clinical indicator indicates responsiveness
  • other clinical indicators e.g., tumor size
  • the upper or lower clinical cutoff point is used to define the level of immune responsiveness.
  • the number of antigens against which an immune response and/or the concentration of antibodies against an antigen against which an immune response is raised is correlated with the clinical outcome data.
  • An immune response cutoff point can be selected such that most subjects having an immune response against more than that number of antigens or with a concentration of antibodies higher than the cutoff concentration in the data set are immunologically responsive to treatment (IR-R), and most subjects having fewer or less than that number are immunologically not responsive (IR-N).
  • a subject in the data set with clinical outcome data more or less than, as appropriate, the clinical outcome cutoff is clinically responsive ("CL-R") to the cancer treatment
  • a subject in the data set with fewer or more than, as appropriate, the clinical outcome cutoff is clinically nonresponsive (“CL-N") to the treatment.
  • a immune response cutoff point is selected that produces the greatest percentage of subject in the data set that are either clinically and immunologically responsive (“IR-R, CL-R”), or immunologically responsive and clinically nonresponsive (“IR-N, CL-N”).
  • the percentage of discordant results is reduced by assigning differential weight values to immune responses against one or more antigens observed in the data set.
  • An algorithm that does not include this step assumes that each immune response in the data set contributes equally to the overall clinical outcome. In many cases this will not be true.
  • immune responses against such antigens are "weighted," e.g., assigned an increased score.
  • An immune response can be assigned a weight of, for example, two, three, four, five, six, seven, eight or more. For example, an immune response assigned a weight of 2 can be counted as two immune responses in a subject. Fractional weighting values can also be assigned. In certain embodiments, a value between zero and one can be assigned when an immune response is weakly associated with a clinical outcome. In another embodiment, values of less than zero can be assigned, wherein an immune response is associated with a negative clinical outcome to the immune response. [00126] One of skill in the art will appreciate that there is a tradeoff involved in assigning an increased weight to certain immune responses. As the weight of the immune response is increased, the number of IR-R, CL-N discordant results may increase.
  • a weight is assigned to an immune response that balances the reduction in IR-N, CL-R results with the increase in IR-R, CL-N results.
  • the interaction of different immune responses in the data set with each other is also factored into the algorithm.
  • two or more immune responses behave synergistically, i.e., that the coincidence of the immune responses in a subject contributes more significantly to the clinical outcome than would be predicted based on the effect of each immune response independent of the other.
  • the coincidence of two or more immune responses in a subject contributes less significantly to the clinical outcome than would be expected from the contributions made to resistance by each immune response when it occurs independently.
  • two or more immune responses may be found to occur more frequently together than as independent immune responses.
  • immune responses occurring together are weighted together.
  • the immune response cutoff point can be used to define a clinical outcome cutoff point by correlating the concentrations of antibody induced as well as the antigens against which immune responses are induced in the data set with the clinical outcome.
  • an algorithm is constructed that factors in the requirement for a certain concentration of antibody to be induced. [00130] By using, for example, the methods discussed above, the algorithm can be designed to achieve any desired result. In one embodiment, the algorithm is designed to maximize the overall concordance (the sum of the percentages of the IR-R, CL-R and the IR-N, CL-N groups, or 100 - (percentage of the IR-N, CL-R + IR-R, CL-N groups). In some embodiments, the overall concordance is greater than 75%, 80%, 85%, 90% or 95%. In one embodiment, the algorithm is designed to minimize the percentage of IR-R, CL-N results.
  • the algorithm is designed to minimize the percentage of IR-N, CL-R results. In another embodiment, the algorithm is designed to maximize the percentage of IR-R, CL-R results. In another embodiment, the algorithm is designed to maximize the percentage of IR-N, CL-N results.
  • the second data set consists of subjects that are not included in the data set, i.e., the second data set is a naive data set.
  • the second data set contains one or more subjects that were in the data set and one or more subjects that were not in the data set.
  • Use of the algorithm on a second data set, particularly a na ⁇ ve data set allows the predictive capability of the algorithm to be assessed.
  • the accuracy of an algorithm is assessed using a second data set, and the rules of the algorithm are modified as described above to improve its accuracy.
  • an iterative approach is used to create the algorithm, whereby an algorithm is tested and then modified repeatedly until a desired level of accuracy is achieved.
  • the present invention also provides a method for using an algorithm of the invention to predict the responsiveness of a subject to a cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSFbased on the immune responses of the subject.
  • the method comprises detecting, in the subject or derivative of the subject, the presence or absence of an immune response against one or more antigens associated with responsiveness to the combination cancer therapy, applying the rules of the algorithm to the detected immune responses, wherein a subject that satisfies the rules of the algorithm is responsive or partially responsive to the treatment, and a subject that does not satisfy the rules of the algorithm is nonresponsive to the treatment.
  • the method comprises detecting, in the subject or derivative of the subject, the presence or absence of an immune response against one or more antigens associated with responsiveness to a cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF, the method comprising applying the rules of the algorithm to the detected mutations, wherein a score equal to, or greater than the immune response cutoff score indicates that the subject is responsive or partially responsive to the treatment, and a score less than the immune response cutoff score indicates that the subject is nonresponsive to the treatment.
  • the method comprises detecting, in the subject or derivative of the subject, the presence or absence of an immune response against one or more antigens associated with responsiveness to a combination cancer therapy, applying the rules of the algorithm to the detected immune responses, wherein a score less than zero indicates that the subject is not likely to respond to the cancer treatment.
  • the present invention relates, in part, to methods relating to the effectiveness of a cancer therapy with the combination of a composition comprising an antibody that specifically binds to human CTLA-4 and a composition comprising cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF.
  • the methods relate to the effectiveness of a cancer therapy with a composition comprising an antibody that specifically binds to human CTLA-4 and cancer cells that have been rendered proliferation-incompetent and have been genetically engineered to express GM-CSF.
  • the method of treating prostate cancer in a subject comprises administering genetically modified cytokine-expressing cells to the subject, in combination with an antibody that specifically binds to human CTLA-4, as part of a therapeutic treatment for cancer.
  • the method can be carried out by genetically modifying (transducing) a first population of tumor cells to produce a cytokine, e.g., GM-CSF, and administering the first population of tumor cells alone or with a second population of tumor cells, in combination with administering an antibody that specifically binds to CTLA-4, to the subject.
  • the tumor cells may be tumor cells from the same individual (autologous), from a different individual (allogeneic) or bystander cells (further described below).
  • the tumor cells are from a tumor cell line of the same type as the tumor or cancer being treated, e.g., the modified cells are prostate or prostate cancer cells and the patient has prostate cancer.
  • the genetically modified tumor cells are rendered proliferation incompetent prior to administration.
  • the genetically modified tumor cells are administered before administration of the anti-CTLA-4 antibody.
  • the genetically modified tumor cells are co-administered with the anti-CTLA-4 antibody.
  • the genetically modified tumor cells are administered after administration of the anti-CTLA-4 antibody.
  • the genetically modified tumor cells are co-administered with the anti- CTLA-4 antibody in separate compositions.
  • the genetically modified tumor cells are co-administered with the anti-CTLA-4 antibody in the same composition.
  • the subject is a human who harbors prostate tumor cells of the same type as the genetically modified cytokine-expressing tumor cells.
  • an improved therapeutic outcome is evident following administration of both the genetically modified cytokine-expressing tumor cells and the anti-CTLA-4 antibody to the subject.
  • Any of the various parameters of an improved therapeutic outcome for a prostate cancer patient known to those of skill in the art may be used to assess the efficacy of the combination therapy described herein, e.g., a reduction in the serum level of PSA.
  • the method is effective to stimulate a systemic immune response in a prostate cancer patient, comprising administering a therapeutically effective amount of proliferation incompetent genetically modified cytokine-expressing cells in combination with an antibody that specifically binds to CTLA-4 to the subject.
  • the systemic immune response to the therapeutic composition may result in tumor regression or inhibit the growth of the tumor.
  • the prostate cancer is metastatic prostate cancer.
  • the prostate cancer is refractory to hormone therapy.
  • the primary prostate tumor has been treated, e.g., by ablation or recission and metasteses of the primary prostate cancer are treated by immunotherapy as described herein.
  • a viral or nonviral vector is utilized to deliver a human GM-CSF transgene (coding sequence) to a human tumor cell ex vivo. After transduction, the cells are irradiated to render them proliferation incompetent. The proliferation incompetent GM-CSF expressing tumor cells are then re-administered to the patient (e.g., by the intradermal or subcutaneous route) and thereby function as a cancer vaccine.
  • the human tumor cell may be a primary tumor cell or derived from a tumor cell line.
  • the genetically modified tumor cells include one or more of autologous tumor cells, allogeneic tumor cells and tumor cell lines (i.e., bystander cells).
  • the tumor cells may be transduced in vitro, ex vivo or in vivo.
  • Autologous and allogeneic cancer cells that have been genetically modified to express a cytokine, e.g., GM-CSF, followed by readministration to a patient for the treatment of cancer are described in U.S. Pat. Nos. 5,637,483, 5,904,920 and 6,350,445, expressly incorporated by reference herein.
  • GM-CSF-expressing genetically modified tumor cells or a "cytokine-expressing cellular vaccine” ("GV AX”®), for the treatment of pancreatic cancer is described in U.S. Pat. Nos. 6,033,674 and 5,985,290, expressly incorporated by reference herein.
  • GV AX® cytokine-expressing cellular vaccine
  • a universal immunomodulatory genetically modified bystander cell line is described in U.S. Pat. No. 6,464,973, expressly incorporated by reference herein.
  • GV AX® wherein the cellular vaccine comprises one or more prostate tumor cell lines selected from the group consisting of DU145, PC-3, and LNCaP is described in WO/0026676, expressly incorporated by reference herein.
  • LNCaP is a PSA-producing prostate tumor cell line, while PC-3 and DU- 145 are non-PSA-producing prostate tumor cell lines (Pang S. et al, Hum Gene Ther. 1995 November; 6(1 1):1417-1426).
  • GM-CSF-expressing cellular vaccines have been undertaken for treatment of prostate cancer, melanoma, lung cancer, pancreatic cancer, renal cancer, and multiple myeloma.
  • a number of clinical trials using GV AX® cellular vaccines have been described, most notably in melanoma, and prostate, renal and pancreatic carcinoma (Simons J W et al. Cancer Res. 1999; 59:5160-5168; Simons J W et al. Cancer Res 1997; 57:1537-1546; Soiffer R et al Proc. Natl. Acad. Sci USA 1998; 95:13141-13146; Jaffee, et al.
  • genetically modified GM-CSF expressing tumor cells are provided as an allogeneic or bystander cell line and one or more additional cancer therapeutic agents is included in the treatment regimen.
  • one or more additional transgenes are expressed by an allogeneic or bystander cell line while a cytokine (i.e., GM-CSF) is expressed by autologous or allogeneic cells.
  • GM-CSF cytokine
  • the GM-CSF coding sequence is introduced into the tumor cells using a viral or non-viral vector and routine methods commonly employed by those of skill in the art.
  • the preferred coding sequence for GM-CSF is the genomic sequence described in Huebner K. et al, Science 230(4731): 1282-5, 1985, however, in some cases the cDNA form of GM-CSF finds utility in practicing the methods (Cantrell et al, Proc. Natl. Acad. Sci., 82, 6250-6254, 1985).
  • the genetically modified tumor cells can be cryopreserved prior to administration.
  • the genetically modified tumor cells are irradiated at a dose of from about 50 to about 200 rads/min, even more preferably, from about 120 to about 140 rads/min prior to administration to the patient.
  • the cells are irradiated with a total dose sufficient to inhibit substantially 100% of the cells from further proliferation.
  • the cells are irradiated with a total dose of from about 10,000 to 20,000 rads, optimally, with about 15,000 rads.
  • cytokine e.g., GM-CSF
  • more than one administration of cytokine (e.g., GM-CSF) producing cells is delivered to the subject in a course of treatment.
  • multiple injections may be given at a single time point with the treatment repeated at various time intervals.
  • an initial or “priming” treatment may be followed by one or more ' " booster” treatments.
  • Such "priming” and “booster” treatments are typically delivered by the same route of administration and/or at about the same site.
  • the first immunization dose may be higher than subsequent immunization doses.
  • a 5 x 10 6 prime dose may be followed by several booster doses of 10 6 to 3 x 10 6 GM-CSF producing cells.
  • a single injection of cytokine-producing cells is typically between about 10 6 to 10 8 cells, e.g., 1 x 10 6 , 2 x 10 6 , 3 x 10 6 , 4 x 10 6 , 5 x 10 6 , 6 x 10 6 , 7 x 10 6 , 8 x 10 6 , 9 x 10 6 , 10 7 , 2 x 10 7 , 5 x 10 7 , or as many as 10 8 cells.
  • the number of cytokine-producing cells may be adjusted according, for example, to the level of cytokine produced by a given cytokine producing cellular vaccine.
  • cytokine-producing cells are administered in a dose that is capable of producing at least 500 ng of GM-CSF per 24 hours per one million cells. Determination of optimal cell dosage and ratios is a matter of routine determination and within the skill of a practitioner of ordinary skill, in light of the disclosure provided herein.
  • the attending physician may administer lower doses of the cytokine-expressing tumor cell vaccine and observe the patient's response. Larger doses of the cytokine- expressing tumor cell vaccine may be administered until the an improved therapeutic outcome is evident.
  • Cytokine-producing cells of the invention are processed to remove most additional components used in preparing the cells.
  • fetal calf serum, bovine serum components, or other biological supplements in the culture medium are removed.
  • the cells are washed, such as by repeated gentle centrifugation, into a suitable pharmacologically compatible excipient.
  • Compatible excipients include various cell culture media, isotonic saline, with or without a physiologically compatible buffer, for example, phosphate or hepes, and nutrients such as dextrose, physiologically compatible ions, or amino acids, particularly those devoid of other immunogenic components.
  • Carrying reagents, such as albumin and blood plasma fractions and inactive thickening agents, may also be used.
  • the method of treating prostate cancer comprises: (a) obtaining tumor cells from a mammalian subject harboring a prostate tumor; (b) genetically modifying the tumor cells to render them capable of producing an increased level of GM-CSF relative to unmodified tumor cells; (c) rendering the modified tumor cells proliferation incompetent; and (d) readministering the genetically modified tumor cells to the mammalian subject from which the tumor cells were obtained or to a mammal with the same MHC type as the mammal from which the tumor cells were obtained, in comination with administering an antibody that specifically binds CTLA-4.
  • the administered tumor cells are autologous and MHC- matched to the host.
  • the tumor cells are administered intradermally, subcutaneously or intratumorally to the mammalian subject.
  • a single autologous tumor cell may express GM-CSF alone or GM-CSF plus one or more additional transgenes.
  • GM-CSF and the one or more additional transgenes may be expressed by different autologous tumor cells.
  • an autologous tumor cell is modified by introduction of a vector comprising a nucleic acid sequence encoding GM-CSF, operatively linked to a promoter and expression/control sequences necessary for expression thereof.
  • the same autologous tumor cell or a second autologous tumor cell can be modified by introduction of a vector comprising a nucleic acid sequence encoding at least one additional transgene operatively linked to a promoter and expression/control sequences necessary for expression thereof.
  • the nucleic acid sequence encoding the one or more transgenes can be introduced into the same or a different autologous tumor cell using the same or a different vector.
  • the nucleic acid sequence encoding the transgene(s) may or may not further comprise a selectable marker sequence operatively linked to a promoter.
  • the autologous tumor cell expresses high levels of GM-CSF. 4.7.1.2 Allogeneic Cells
  • the bone marrow-derived APCs take up the whole cellular protein of the tumor for processing, and then present the antigenic peptide(s) on their MHC class I and II molecules, thereby priming both the CD4+ and the CD8+ T cell arms of the immune system, resulting in a systemic tumor-specific anti-tumor immune response.
  • these results suggest that it may not be necessary or optimal to use autologous or MHC-matched cells in order to elicit an anti-cancer immune response and that the transfer of allogeneic MHC genes (from a genetically dissimilar individual of the same species) can enhance tumor immunogenicity.
  • the rejection of tumors expressing allogeneic MHC class I molecules has resulted in enhanced systemic immune responses against subsequent challenge with the unmodified parental tumor. See, e.g., Jaffee et al, supra, and Huang et al , supra.
  • a tumor cell line comprises cells that were initially derived from a tumor. Such cells typically exhibit indefinite growth in culture.
  • the method for treating prostate cancer comprises: (a) obtaining a tumor cell line; (b) genetically modifying the tumor cell line to render the cells capable of producing an increased level of a cytokine, e.g., GM-CSF, relative to the unmodified tumor cell line; (c) rendering the modified tumor cell line proliferation incompetent; and (d) administering the tumor cell line to a mammalian subject (host) having at least one tumor that is of the same type of tumor as that from which the tumor cell line was obtained.
  • a mammalian subject host having at least one tumor that is of the same type of tumor as that from which the tumor cell line was obtained.
  • the administered tumor cell line is allogeneic and is not MHC-matched to the host.
  • allogeneic lines provide the advantage that they can be prepared in advance, characterized, aliquoted in vials containing known numbers of transgene (e.g., GM-CSF) expressing cells and stored (i.e. frozen) such that well characterized cells are available for administration to the patient.
  • transgene e.g., GM-CSF
  • Methods for the production of genetically modified allogeneic cells are described for example in WO 00/72686, expressly incorporated by reference herein.
  • a nucleic acid sequence (transgene) encoding GM-CSF alone or in combination with the nucleic acid coding sequence for one or more additional transgenes is introduced into a cell line that is an allogeneic tumor cell line (i.e., derived from an individual other than the individual being treated).
  • a nucleic acid sequence (transgene) encoding GM-CSF alone or in combination with the nucleic acid coding sequence for one or more additional transgenes is introduced into separate allogeneic tumor cell lines.
  • two or more different genetically modified allogeneic GM-CSF expressing cell lines e.g.
  • LNCAP and PC-3) are administered in combination, typically at a ratio of 1 : 1.
  • the cell or population of cells is from a tumor cell line of the same type as the tumor or cancer being treated, e.g. prostate cancer.
  • the nucleic acid sequence encoding the transgene(s) may be introduced into the same or a different allogeneic tumor cell using the same or a different vector.
  • the nucleic acid- sequence encoding the transgene(s) may or may not further comprise a selectable marker sequence operatively linked to a promoter.
  • the allogeneic cell line expresses high levels of GM-CSF.
  • one or more genetically modified GM-CSF expressing allogeneic cell lines can be exposed to an antigen, such that the patient's immune response to the antigen is increased in the presence of GM-CSF, e.g., an allogeneic or bystander cell that has been genetically modified to express GM-CSF.
  • an antigen is a peptide comprising an amino acid sequence obtained from filamin-B, as described extensively above.
  • the filamin-B peptide can be provided by (on) cells that are administered to the subject or may be provided by cells native to the patient.
  • the composition can be rendered proliferation-incompetent, typically by irradiation, wherein the allogeneic cells are plated in a tissue culture plate and irradiated at room temperature using a Cs source, as further described herein.
  • An allogeneic cellular vaccine composition of the invention may comprise allogeneic cells plus other cells, i.e. a different type of allogeneic cell, an autologous cell, or a bystander cell that may or may not be genetically modified. If genetically modified, the different type of allogeneic cell, autologous cell, or bystander cell may express GM-CSF or another transgene. The ratio of allogeneic cells to other cells in a given administration will vary dependent upon the combination.
  • any suitable route of administration can be used to introduce an allogeneic cell line composition into the patient, for example, the composition is administered intradermally, subcutaneously or intratumorally.
  • allogeneic cell lines in practicing the present invention provides the therapeutic advantage that administration of a genetically modified GM-CSF expressing cell line to a patient with cancer, together with an autologous cancer antigen, paracrine production of GM-CSF results in an effective immune response to a tumor. This obviates the need to culture and transduce autologous tumor cells for each patient.
  • a universal immunomodulatory genetically modified transgene-expressing bystander cell that expresses at least one transgene can be used in the immunotherapies described herein.
  • the same universal bystander cell line may express more than one transgene or individual transgenes may be expressed by different universal bystander cell lines.
  • the universal bystander cell line comprises cells which either naturally lack major histocompatibility class I (MHC-I) antigens and major histocompatibility class II (MHC-II) antigens or have been modified so that they lack MHC-I antigens and MHC-II antigens.
  • MHC-I major histocompatibility class I
  • MHC-II major histocompatibility class II
  • a universal bystander cell line can be modified by introduction of a vector wherein the vector comprises a nucleic acid sequence encoding a transgene, e.g., a cytokine such as GM-CSF, operably linked to a promoter and expression control sequences necessary for expression thereof.
  • a transgene e.g., a cytokine such as GM-CSF
  • the same universal bystander cell line or a second universal bystander cell line is modified by introduction of a vector comprising a nucleic acid sequence encoding at least one additional transgene operatively linked to a promoter and expression control sequences necessary for expression thereof.
  • the nucleic acid sequence encoding the transgene(s) may be introduced into the same or a different universal bystander cell line using the same or a different vector.
  • the nucleic acid sequence encoding the transgene(s) may or may not further comprise a selectable marker sequence operatively linked to a promoter. Any combination of transgene(s) that stimulate an anti-tumor immune response can be used.
  • the universal bystander cell line preferably grows in defined, i.e., serum-free medium, preferably as a suspension.
  • the universal bystander cell line expresses high levels of the transgene, e.g. a cytokine such as GM-CSF.
  • the one or more universal bystander cell lines can be incubated with an autologous cancer antigen, e.g., provided by an autologous tumor cell (which together comprise a universal bystander cell line composition), then the universal bystander cell line composition can be administered to the patient. Any suitable route of administration can be used to introduce a universal bystander cell line composition into the patient. In certain embodiments, the composition is administered intradermally, subcutaneously or intratumorally.
  • the autologous cancer antigen can be provided by a cell of the cancer to be treated, i.e., an autologous cancer cell.
  • the composition is rendered proliferation-incompetent by irradiation, wherein the bystander cells and cancer cells are plated in a tissue culture plate and irradiated at room temperature using a Cs source, as detailed above.
  • the ratio of bystander cells to autologous cancer cells in a given administration will vary dependent upon the combination. With respect to GM-CSF- producing bystander cells, the ratio of bystander cells to autologous cancer cells in a given administration should be such that a therapeutically effective level of GM-CSF is produced. In addition to the GM-CSF threshold, the ratio of bystander cells to autologous cancer cells should not be greater than 1 :1. Appropriate ratios of bystander cells to tumor cells or tumor antigens can be determined using routine methods known in the art.
  • the use of bystander cell lines in practicing the present invention provides the therapeutic advantage that, through administration of a cytokine-expressing bystander cell line and at least one additional cancer therapeutic agent (expressed by the same or a different cell) to a patient with cancer, together with an autologous cancer antigen, paracrine production of an immunomodulatory cytokine, results in an effective immune response to a tumor. This obviates the need to culture and transduce autologous tumor cells for each patient.
  • a minimum dose of about 3500 rads is sufficient to inactivate a cell and render it proliferation-incompetent, although doses up to about 30,000 rads are acceptable.
  • the cells are irradiated at a dose of from about 50 to about 200 rads/min or from about 120 to about 140 rads/min prior to administration to the mammal.
  • the levels required are 2,500 rads, 5,000 rads, 10,000 rads, 15,000 rads or 20,000 rads.
  • a dose of about 10,000 rads is used to inactivate a cell and render it proliferation-incompetent.
  • irradiation is but one way to render cells proliferation-incompetent, and that other methods of inactivation which result in cells incapable of multiple rounds of cell division but that retain the ability to express transgenes (e.g. cytokines) are included in the present invention (e.g., treatment with mitomycin C, cycloheximide, and conceptually analogous agents, or incorporation of a suicide gene by the cell).
  • transgenes e.g. cytokines
  • a "cytokine” or grammatical equivalent includes, without limitation, those hormones that act locally and do not circulate in the blood, and which, when used in accordance with the present invention, will result in an alteration of an individual's immune response. Also included in the definition of cytokine are adhesion or accessory molecules which result in an alteration of an individual's immune response.
  • cytokines include, but are not limited to, IL-I (a or P), IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL- 12, GM-CSF, M-CSF, G-CSF, LIF, LT, TGF-P, ⁇ -IFN, a-EFN, P-IFN, TNF- ⁇ , BCGF, CD2, or ICAM. Descriptions of the aforementioned cytokines as well as other applicable immunomodulatory agents may be found in "Cytokines and Cytokine Receptors," A. S. Hamblin, D.
  • the cytokines will preferably be substantially similar to the human form of the protein or will have been derived from human sequences (i.e., of human origin).
  • the transgene is a cytokine, such as GM-CSF.
  • cytokines of other mammals with substantial structural homology and/or amino acid sequence identity to the human forms of a given cytokine will be useful when demonstrated to exhibit similar activity on the human immune system.
  • proteins that are substantially analogous to any particular cytokine, but have conservative changes of protein sequence can also be used.
  • conservative substitutions in protein sequence may be possible without disturbing the functional abilities of the protein molecule, and thus proteins can be made that function as cytokines in the present invention but have amino acid sequences that differ slightly from currently known sequences.
  • Such conservative substitutions typically include substitutions within the following groups: glycine, alanine, valine, isoleucine, leucine; aspartic acid, glutamic acid, asparagine, glutamine; serine, threonine; lysine, arginine; and phenylalanine, tyrosine.
  • Granulocyte-macrophage colony stimulating factor is a cytokine produced by fibroblasts, endothelial cells, T cells and macrophages. This cytokine has been shown to induce the growth of hematopoetic cells of granulocyte and macrophage lineages. In addition, it also activates the antigen processing and presenting function of dendritic cells, which are the major antigen presenting cells (APC) of the immune system. Results from animal model experiments have convincingly shown that GM-CSF producing cells are able to induce an immune response against parental, non-transduced cells.
  • APC antigen presenting cells
  • GM-CSF augments the antigen presentation capability of the subclass of dendritic cells (DC) capable of stimulating robust anti-tumor responses (Gasson et al Blood 1991 Mar. 15;77(6):1131-45; M&ch et al. Cancer Res. 2000 Jun. 15;60(12):3239-46; reviewed in Mach and Dranoff, Curr Opin Immunol. 2000 October; 12(5):571-5). See, e.g., Boon and Old, Curr Opin Immunol. 1997 Oct. 1; 9(5):681-3). Presentation of tumor antigen epitopes to T cells in the draining lymph nodes is expected to result in systemic immune responses to tumor metastases.
  • DC dendritic cells
  • irradiated tumor cells expressing GM-CSF have been shown to function as potent vaccines against tumor challenge.
  • Localized high concentrations of certain cytokines, delivered by genetically modified cells, have been found to lead to tumor regression (Abe et al, J. Cane. Res. Clin. Oncol. 121 : 587-592 (1995); Gansbacher et al, Cancer Res. 50: 7820-7825 (1990); Formi et al, Cancer and Met. Reviews 7: 289-309 (1988).
  • PCT publication WO200072686 describes tumor cells expressing various cytokines.
  • the cellular immunogenic composition comprises a GM-CSF coding sequence operatively linked to regulatory elements for expression in the cells of the vaccine.
  • the GM-CSF coding sequence may code for a murine or human GM-CSF and may be in the form of genomic DNA (SEQ ID NO: NO.:53; disclosed as SEQ ID NO: NO.:1 in US Patent Publication NO. 2006/0057127, which is hereby incorporated by reference in its entirety) or cDNA (SEQ ID NO: NO.:54; disclosed as SEQ ID NO: NO.:2 in US Patent Publication NO. 2006/0057127, which is hereby incorporated by reference in its entirety).
  • the coding sequence for GM-CSF does not contain intronic sequences to be spliced out prior to translation.
  • the coding sequence contains at least one native GM-CSF intron that is spliced out prior to translation.
  • the GM-CSF coding sequence encodes the amino acid sequence presented as SEQ ID NO: NO.:55 (disclosed as SEQ ID NO.: 3 in US Patent Publication NO. 2006/0057127, which is hereby incorporated by reference in its entirety).
  • Other examples of GM-CSF coding sequences are found in Genbank accession numbers: AF373868, AC034228, AC034216, M 10663 and NM000758.
  • a GM-CSF coding sequence can be a full-length complement that hybridizes to the sequence shown in SEQ ID NO: NO:53 or SEQ ID NO: 54 under stringent conditions.
  • hybridizing to refers to the binding, duplexing, or hybridizing of a molecule to a particular nucleotide sequence under stringent conditions when that sequence is present in a complex mixture (e.g., total cellular) DNA or RNA.
  • Bod(s) substantially refers to complementary hybridization between a probe nucleic acid and a target nucleic acid and embraces minor mismatches that can be accommodated by reducing the stringency of the hybridization media to achieve the desired detection of the target nucleic acid sequence.
  • the coding sequence for a cytokine such as GM- CSF, can have at least 80, 85, 87, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99% or more % identity over its entire length to a native GM-CSF coding sequence.
  • a GM-CSF coding sequence can have at least 80, 85, 87, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99% or more sequence identity to a sequence presented as SEQ ID NO: NO:53 or SEQ ID NO:54, when compared and aligned for maximum correspondence, as measured a sequence comparison algorithm (as described above) or by visual inspection.
  • the given % sequence identity exists over a region of the sequences that is at least about 50 nucleotides in length. In another embodiment, the given % sequence identity exists over a region of at least about 100 nucleotides in length.
  • the given % sequence identity exists over a region of at least about 200 nucleotides in length. In another embodiment, the given % sequence identity exists over the entire length of the sequence.
  • the GM- CSF has authentic GM-CSF activity, e.g., can bind the GM-CSF receptor.
  • the amino acid sequence for a cytokine such as GM-CSF has at least 80, 85, 87, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99% or more sequence identity to the sequence presented as SEQ ID NO: NO:55, when compared and aligned for maximum correspondence.
  • an antibody that specifically binds to human CTLA-4 is administered in combination with tumor cells genetically altered to express cytokines to a subject.
  • CTLA-4 is a T cell surface molecule that was originally identified by differential screening of a murine cytolytic T cell cDNA library (Brunet et al., Nature 328:267-270(1987)).
  • CTLA-4 is also a member of the immunoglobulin (Ig) superfamily.
  • CTLA-4 comprises a single extracellular Ig domain.
  • CTLA-4 transcripts have been found in T cell populations having cytotoxic activity, suggesting that CTLA-4 might function in the cytolytic response (Brunet et al., supra; Brunet et al., Immunol. Rev.
  • CTLA-4 has an analogous function as a secondary costimulator (Linsley et al., J Exp. Med. 176:1595-1604 (1992); Wu et al., J. Exp. Med. 185:1327-1335 (1997) Lindsley, P. et al. U.S. Pat. Nos. 5,977,318; 5,968,510; 5,885,796; and 5,885,579).
  • CTLA-4 has an opposing role as a dampener of T cell activation (Krummel (1995) J. Exp. Med. 182:459-465); Krummel et al., Int'l Immunol.
  • CTLA-4 deficient mice suffer from massive lymphoproliferation (Chambers et al., supra). It has also been reported that CTLA-4 blockade augments T cell responses in vitro (Walunas et al., Immunity. 1 :405-413 (1994)) and in vivo (Kearney (1995) J. Immunol. 155:1032-1036), exacerbates antitumor immunity (Leach (1996) Science. 271 :1734-1736), and enhances an induced autoimmune disease (Luhder (1998) J Exp. Med. 187:427-432).
  • CTLA-4 has an alternative or additional impact on the initial character of the T cell immune response (Chambers (1997) Curr. Opin. Immunol. 9:396404; Bluestone (1997) J. Immunol. 158:1989- 1993; Thompson (1997) Immunity 7:445-450). This is consistent with the observation that some autoimmune patients have autoantibodies to CTLA-4. It is possible that CTLA-4 blocking antibodies have a pathogenic role in these patients (Matsui (1999) J. Immunol. 162:4328-4335).
  • the antibody administered in combination with genetically altered tumor cells is a polyclonal, monoclonal, single chain monoclonal, recombinant, chimeric, humanized, mammalian, or fully human antibody that specifically binds to CTLA-4.
  • Exemplary anti-CTLA-4 antibodies, and methods for their use in the treatment of human diseases, infections and other conditions are described in U.S. Pat. Nos. 6,682,736, 7,109,003, 7,132,281 and 6,984,720, and U.S. Patent App. Nos.
  • anti-CTLA-4 antibodies and methods for their use are described in PCT application publications WO2006/048749, WO2006/101691, WO2006/101692, and WO2007/008463, each of which are expressly incorporated by reference herein.
  • anti- CTLA-I antibodies and methods for their use are described in PCT application numbers PCT/US06/61753.
  • the anti-CTLA-4 antibody is a fully human monoclonal antibody specific for human CTLA-4, e.g. MDX-010 (ipilimumab).
  • the anti-CTLA-4 antibody may be administered, for example, in an amount of at least about 0.3 mg/kg, at least about 1 mg/kg, at least about 5 mg/kg, at least about 10 mg/kg, at least about 15 mg/kg or greater.
  • a single dose or multiples doses of the antibody may be administered.
  • at least one dose, or at least three, six, 12 doses or greater may be administered.
  • the doses may be administered, for example, every week, every two weeks, every 4 weeks, monthly, every three months, every six months or yearly.
  • Doses may be administered simultaneous with the administration of the genetically altered tumor cells, for example with the first administration of the therapeutic composition and with subsequent administrations of the therapeutic composition.
  • Exemplary methods for producing recombinant viral vectors useful for making genetically altered tumor cells that express GM-CSF, methods for using the genetically altered tumor cells that express GM-CSF in cancer therapies, particularly prostate cancer therapies, are extensively described in U.S. Patent Application Publication No. 20060057127, which is incorporated ' by reference in its entirety.
  • One such therapy that has been and is being evaluated in clinical trials for treatment of prostate cancer is GV AX® therapy combined with ipilimumab (MDX-OlO) therapy.
  • Humoral patient immune responses to the combined cell-based/anti- CTLA-4 prostate cancer immunotherapy have been evaluated to specifically identify which antigens may be specifically recognized by the patients' immune system following the therapy.
  • Two differing methods have been used to characterize this response using patients' sera: i) serological analysis of gene expression libraries (SEREX) and ii) defined prostate cancer antigen screening. From these two techniques, multiple antibody responses to proteins derived from the combination immunotherapy have been identified that are specifically induced or augmented following immunization.
  • SEREX allows the systematic cloning of tumor antigens recognized by the autoantibody repertoire of cancer patients (Sahin et al. 1995; McNeel et al. 2000; Wang et al. 2005; Dunphy et al. 2005; Qin et al. 2006).
  • This procedure was carried out for 3 patients treated with the combined cell-based/anti-CTLA-4 prostate cancer immunotherapy. These patients were prioritized for SEREX analysis based upon serum PSA decline and radiographic changes in metastatic tumor sites (bone and lymph nodes. From the SEREX analysis of these 3 patients, multiple LNCaP/PC-3/Pituitary derived cell protein clones reactive to the patient sera post- immunotherapy were identified. Positive hits for the SEREX screen were then screened against pre-immunotherapy serum to determine if the antibody response to these proteins was augmented or induced following the immunotherapy .
  • antigens are de novo antigens, i.e., antibody responses that are not detectable pre-therapy in any patient screened so far, but are present in at least 2 (of the total patients screened) following immunotherapy. Response is absolute (on/off). These results indicate that these genes can serve as an important marker of clinical benefit in patients.
  • This example describes the results of experiments designed to assess humoral immune responses against prostate cancer antigens.
  • a selection of 20 genes that are associated with prostate cancer and have previously demonstrated an interaction with the immune response were selected for evaluation.
  • the genes are set forth in Table 3, below. This list includes:
  • PSA Prostate specific antigen
  • PSMA Prostate-specific membrane antigen
  • PAP Prostatic acid phosphatase
  • PSCA Prostate stem cell antigen
  • CEA Carcinoembryonic antigen
  • AMACR Her2/neu ⁇ -methylacyl-CoA racemase
  • Glucose-regulated protein-78 kDa GFP78
  • TARP T-cell receptor gamma alternate reading frame protein
  • antigen targets are further characterized for cellular immune response (T- cells) using peripheral blood mononuclear cells (PBMCs) harvested from patients administered the combination cell-based/anti-CTLA-4 prostate cancer immunotherapy.
  • PBMCs peripheral blood mononuclear cells
  • This example provides an exemplary method for detecting activation of cytotoxic T lymphocytes (CTLs) by monitoring IFN- ⁇ expression by the CTLs in response to exposure to an appropriate antigen, e.g., a filamin-B peptide presented on an MHC I receptor.
  • CTLs cytotoxic T lymphocytes
  • PBMCs peripheral blood monocytic cells
  • FACS fluorescence activated cell sorting
  • IFN- ⁇ release by the CTLs is measured using an IFN- ⁇ ELISA kit (PBL- Biomedical Laboratory, Piscataway, NJ). Briefly, purified IFN- ⁇ as standards or culture supernates from the CTL-T2 co-culture are transferred into wells of a 96-well plate pre-coated with a monoclonal anti-human IFN- ⁇ capture antibody and incubated for 1 h in a closed chamber at 24 °C. After washing the plate with PBS/0.05% Tween 20, biotin anti-human IFN- ⁇ antibody is added to the wells and incubated for 1 h at 24 0 C.
  • IFN- ⁇ ELISA kit PBL- Biomedical Laboratory, Piscataway, NJ.
  • the wells are washed and then developed by incubation with streptavidin horseradish peroxidase conjugate and TMB substrate solution. Stop solution is added to each well and the absorbance is determined at 450 run with a SpectraMAX Plus plate reader (Stratagene, La Jolla, CA). The amount of cytokine present in the CTL culture supernatants is calculated based on the IFN- ⁇ standard curve.
  • This example provides an exemplary method for detecting activation of cytotoxic T lymphocytes (CTLs) by CTL proliferation in response to exposure to an appropriate antigen, e.g., a filamin-B peptide presented on an MHC I receptor.
  • CTLs cytotoxic T lymphocytes
  • an appropriate antigen e.g., a filamin-B peptide presented on an MHC I receptor.
  • PBMCs peripheral blood monocytic cells
  • CD8+ cells are isolated by fluorescence activated cell sorting (FACS).
  • FACS fluorescence activated cell sorting
  • the CD8+ cells are then incubated with, e.g., T2 cells loaded with the filamin-B peptide to be assessed, produced as described above.
  • the samples are incubated for 12 hours, then 20 ⁇ l of 3H-thymidine is added to each well and the sample incubated for an additional 12 hours.
  • Cells are harvested and the plate is read in a beta counter to determine the amount of ⁇ unincorporated 3H-thymidine.
  • This example provides an exemplary method for detecting activation of cytotoxic T lymphocytes (CTLs) by monitoring lysis of cells displaying an appropriate antigen, e.g., a filamin-B peptide presented on an MHC I receptor.
  • CTLs cytotoxic T lymphocytes
  • CTLs The cytotoxic activity of the CTLs is measured in a standard ⁇ 1 Cr-release assay. Effector cells (CTLs) are seeded with ⁇ Cr-labeled target cells (5 x 10 ⁇ cells/well) at various effecto ⁇ target cell ratios in 96-well U-bottom microtiter plates.
  • CTLs Effector cells
  • Antibody immunity to prostate cancer associated antigens can be detected in the serum of patients with prostate cancer. J Urol. 2000 Nov; 164(5): 1825-9.
  • Chinnaiyan AM Integrative genomic and proteomic analysis of prostate cancer reveals signatures of metastatic progression. Cancer Cell. 2005 Nov;8(5):393-406.

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Abstract

La présente invention concerne des procédés et des compositions pour identifier le cancer de la prostate ou une réponse immunitaire à médiation humorale contre le cancer de la prostate. L'invention concerne également des procédés pour déterminer si un sujet répond ou est susceptible de répondre à une thérapie contre le cancer de la prostate.
PCT/US2008/004016 2007-03-28 2008-03-27 Procédés et compositions pour identifier le cancer de la prostate ou une réponse immunitaire à médiation humorale contre le cancer de la prostate WO2008121307A2 (fr)

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CN104583422A (zh) * 2012-06-27 2015-04-29 博格有限责任公司 标志物在诊断和治疗前列腺癌中的用途
CN108500301A (zh) * 2018-04-03 2018-09-07 台州屹捷数控机床有限公司 一种可延长的双向主轴
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Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US8119129B2 (en) 2008-08-01 2012-02-21 Bristol-Myers Squibb Company Combination of anti-CTLA4 antibody with dasatinib for the treatment of proliferative diseases
US8685394B2 (en) 2008-08-01 2014-04-01 Bristol-Myers Squibb Company Combination of anti-CTLA4 antibody with diverse therapeutic regimens for the synergistic treatment of proliferative diseases
US9320811B2 (en) 2008-08-01 2016-04-26 Bristol-Myers Squibb Company Combination of anti-CTLA4 antibody with diverse therapeutic regimens for the synergistic treatment of proliferative diseases
CN104583422A (zh) * 2012-06-27 2015-04-29 博格有限责任公司 标志物在诊断和治疗前列腺癌中的用途
EP2867375A4 (fr) * 2012-06-27 2016-06-01 Berg Llc Utilisation de marqueurs dans le diagnostic et le traitement du cancer de la prostate
US9797905B2 (en) 2012-06-27 2017-10-24 Berg Llc Use of markers in the diagnosis and treatment of prostate cancer
US10539566B2 (en) 2014-12-08 2020-01-21 Berg Llc Use of markers including filamin A in the diagnosis and treatment of prostate cancer
CN108500301A (zh) * 2018-04-03 2018-09-07 台州屹捷数控机床有限公司 一种可延长的双向主轴
CN108500301B (zh) * 2018-04-03 2019-09-20 台州屹捷数控机床股份有限公司 一种可延长的双向主轴

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