Il-12 immunotherapy for cancer

Abstract

Compositions and methods for delivering immune modulatory molecules to result in a therapeutic effect are disclosed. The compositions and methods use stably integrating lentiviral delivery systems. The methods are useful for therapeutically and prophylactically treating cancer such as leukemia.

1 .- 23 . (canceled) 
     
     
         24 . A method of expressing IL-12 in a cell that is secreted above a threshold level, optionally a cancer cell, comprising: contacting the cell with a composition comprising a vector construct or an isolated virus comprising a lentiviral delivery vector and an IL-12 expression cassette, wherein the lentiviral delivery vector comprises a central polypurine tract (cPPT) sequence, optionally SEQ ID NO: 2 and/or a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE), optionally SEQ ID NO: 3, under conditions that permit transduction of the cell, thereby providing a transduced cell, wherein the IL-12 is secreted and the IL-12 secreted per cell is above a threshold level. 
     
     
         25 . The method of  claim 24 , further comprising a step of isolating the transduced cell or isolating a population of cells comprising the transduced cell. 
     
     
         26 . The method of  claim 24 , further comprising:
 a. growth arresting the transduced cell or the population of cells; and   b. optionally introducing the transduced growth arrested cell, population of cells and/or a composition comprising said transduced growth arrested cell or population of cells in a subject.   
     
     
         27 .- 29 . (canceled) 
     
     
         30 . The method of  claim 31 , wherein the cancer is leukemia, optionally ALL, AML, CML or CLL. 
     
     
         31 . A method of inducing or enhancing an immune response, optionally a memory immune response in a subject, optionally with cancer or an increased risk of cancer and/or treating a subject with cancer or an increased risk of cancer comprising administering: i) an isolated virus-comprising a lentiviral delivery vector and an IL-12 expression cassette wherein the lentiviral vector comprises a central polypurine tract (cPPT) sequence, optionally SEQ ID NO: 2 and/or a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE), optionally SEQ ID NO: 3, ii) a transduced cell comprising a lentiviral delivery vector and an IL-12 expression cassette wherein the transduced cell expresses IL-12 above a threshold level iii) a population of cells comprising the transduced cell of ii), or iv) a composition comprising i), ii) and/or iii). 
     
     
         32 . (canceled) 
     
     
         33 . (canceled) 
     
     
         34 . The method of  claim 31  wherein the transduced cell is growth arrested prior to administering to the subject, optionally wherein the transduced cell is irradiated prior to administering to the subject. 
     
     
         35 . (canceled) 
     
     
         36 . A method of delivering IL-12 to a subject, optionally with cancer or an increased risk or cancer, optionally, for enhancing cancer treatment comprising:
 a. generating a recombinant IL-12 secreting cell wherein the IL-12 secreted per cell is above a threshold level; and   b. introducing an effective number of the generated recombinant IL-12 secreting cells to the subject, optionally in a population of cells comprising the recombinant IL-12 secreting cell.   
     
     
         37 . (canceled) 
     
     
         38 . (canceled) 
     
     
         39 . The method of  claim 36  wherein the threshold level of IL-12 secreted is at least 1.5 fg/ml cells/2 hrs of IL-12, at least 1.5 pg/ml cells/2 hrs of IL-12, or at least 1500 pg/mL/10 6  cells/2 hrs of IL-12, optionally at least 1500 pg/mL/10 6  cells/2 hrs, 1500-2500 pg/mL/10 6  cells/2 hrs, 2500-5000 pg/mL/10 6  cells/2 hrs, 5000-7500 pg/mL/10 6  cells/2 hrs, 7500-10000 pg/mL/10 6  cells/2 hrs, 10000-12500 pg/mL/10 6  cells/2 hrs, 12500-15000 pg/mL/10 6  cells/2 hrs, 15000-17500 pg/mL/10 6  cells/2 hrs, 17500-20000 pg/mL/10 6  cells/2 hrs or at least 20000 pg/mL/10 6  cells/2 hrs of IL-12. 
     
     
         40 . The method of  claim 36 , wherein the recombinant IL-12 secreting cell is generated by contacting the cell with a composition comprising a lentiviral delivery vector and an IL-12 expression cassette, wherein the lentiviral delivery vector comprises a central polypurine tract (cPPT) sequence, optionally SEQ ID NO: 2 and/or a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE), optionally SEQ ID NO: 3. 
     
     
         41 . The method of  claim 36 , wherein the recombinant cell is a cancer cell, optionally derived from the subject with cancer. 
     
     
         42 . The method of  claim 36 , wherein the recombinant cell or population of cells are introduced by IP injection, subcutaneously or intradermally. 
     
     
         43 . The method of  claim 36 , wherein the cancer is a leukemia, optionally ALL, AML, CML or CLL. 
     
     
         44 . The method of  claim 36 , wherein the IL-12 secreting cell is growth arrested prior to introducing to the subject. 
     
     
         45 . (canceled) 
     
     
         46 . (canceled) 
     
     
         47 . The method of  claim 31 , wherein the immune response leads to long-term immune memory and/or does not induce or enhance antagonistic cytokines, and/or wherein the response is initiated substantially free of inducing or enhancing of a CD8 +  T cell-dependent immune response. 
     
     
         48 . (canceled) 
     
     
         49 . The method of  claim 31  wherein the level of IL-12 secreted is above a threshold level that enhances dendritic cell maturation and/or antigen presentation. 
     
     
         50 .- 71 . (canceled) 
     
     
         72 . The method of  claim 36 , wherein the number of cells introduced ranges from 10 5  cells to 10 9  cells, optionally about 10 5 , about 10 6  cells, about 10 7  cells, about 10 8  cells, or about 10 9  cells. 
     
     
         73 . (canceled) 
     
     
         74 . The method of  claim 36  wherein the population of cells optionally comprises at least 0.1 to 1% IL-12 producing leukemic cells, optionally about 0.5%, about 1%, about 1-5%, 5-10%, 10-20% or more IL-12 producing leukemic cells, and wherein the population of cells secretes above the threshold level optionally the threshold level necessary to induce or enhance a CD4+ T cell dependent immune response, optionally at least 1500 pg/mL/10 6  cells/2 hrs, 1500-2500 pg/mL/10 6  cells/2 hrs, 2500-5000 pg/mL/10 6  cells/2 hrs, 5000-7500 pg/mL/10 6  cells/2 hrs, 7500-10000 pg/mL/10 6  cells/2 hrs, 10000-12500 pg/mL/10 6  cells/2 hrs, 12500-15000 pg/mL/10 6  cells/2 hrs, 15000-17500 pg/mL/10 6  cells/2 hrs, 17500-20000 pg/mL/10 6  cells/2 hrs or at least 20000 pg/mL/10 cells/2 hrs of IL-12. 
     
     
         75 . The method of  claim 24 , wherein the IL-12 expression cassette comprises a polynucleotide optionally encoding a p35 polypeptide and a polynucleotide encoding a p40 polypeptide; or a polynucleotide encoding an IL-12 fusion polypeptide. 
     
     
         76 . The method of  claim 75  wherein the IL-12 fusion polypeptide has at least 70% sequence identity to SEQ ID NO: 4 and binds an IL-12 receptor. 
     
     
         77 . The method of  claim 24  wherein the vector construct further comprises one or more of: an activator polynucleotide encoding a polypeptide that converts a prodrug to a drug, optionally a modified tmpk polynucleotide; a detection cassette, optionally a CD19, truncated CD19, CD20, human CD24, murine HSA, human CD25 (huCD25), a truncated form of low affinity nerve growth factor receptor (LNGFR), truncated CD34, eGFP, eYFP, or any other fluorescent protein or erythropoietin receptor (EpoR) polynucleotide; or a polynucleotide with at least 70% sequence identity to said polynucleotide; and an immune modulatory cassette, optionally a polynucleotide that encodes a polypeptide that modulates an immune cell, optionally a dendritic cell or a T cell, optionally a CD4+ T cell, optionally CD40L, IL-7, or IL-15.