Combinatorial Methods For Inducing Cancer Cell Death

Abstract

The present invention relates to methods and compositions for inhibiting proliferation and inducing cell death in a population of cancer cells by (i) increasing the amount of the differentiation associated protein MDA-7, and (ii) decreasing RAS activity within the population. It is based, at least in part, on the discovery that decreasing expression of a mutated, activated K-ras gene, together with introducing an expressible mda-7 gene, in pancreatic cells had a synergistic growth-inhibitory and anti-survival effect, and abolished tumorigenicity of the cells in athymic nude mice. The methods of the invention may be directed to the therapy of pancreatic cancer and other malignancies.

1 . A method for inhibiting proliferation and inducing cell death in a population of cancer cells by (i) increasing the amount of the differentiation associated protein, MDA-7 and (ii) decreasing RAS activity within the population.  
     
     
         2 . The method of  claim 1  wherein the amount of MDA-7 is increased by introducing, into one or more cell of the population, a nucleic acid encoding MDA-7 protein in expressible form.  
     
     
         3 . The method of  claim 2  wherein the nucleic acid encoding MDA-7 protein is comprised in a viral vector.  
     
     
         4 . The method of  claim 3  wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         5 . The method of  claim 1  wherein RAS activity is decreased by administering, to the cancer cell population, an effective amount of an anti-RAS agent, which may be, for example, an antisense molecule, a ribozyme, a precursor of a triple helix, or a farnesyl transferase inhibitor.  
     
     
         6 . The method of  claim 5 , wherein RAS activity is decreased by administering an effective amount of a viral vector encoding an antisense molecule.  
     
     
         7 . The method of  claim 6 , wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         8 . The method of  claim 5 , wherein RAS activity is decreased by administering an effective amount of an antisense molecule which is an oligonucleotide.  
     
     
         9 . The method of  claim 6 , wherein the viral vector further comprises a nucleic acid encoding MDA-7 in expressible form.  
     
     
         10 . The method of  claim 1  wherein RAS activity is decreased by administering, to the cancer cell population, an effective amount of an agent which inhibits a molecule selected from the group consisting of the epidermal growth factor receptor, RAF, MAPK kinase, MAP kinase, and PI3 kinase.  
     
     
         11 . A method for inhibiting proliferation and/or inducing cell death of a cancer cell by (i) increasing the amount of the differentiation associated protein, MDA-7 and (ii) decreasing RAS activity in the cancer cell.  
     
     
         12 . The method of  claim 11  wherein the amount of MDA-7 is increased by introducing, into the cancer cell, a nucleic acid encoding MDA-7 protein in expressible form.  
     
     
         13 . The method of  claim 12  wherein the nucleic acid encoding MDA-7 protein is comprised in a viral vector.  
     
     
         14 . The method of  claim 13  wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         15 . The method of  claim 11  wherein RAS activity is decreased by administering, to the cancer cell, an effective amount of an anti-RAS agent, which may be, for example, an antisense molecule, a ribozyme, a precursor of a triple helix, or a farnesyl transferase inhibitor.  
     
     
         16 . The method of  claim 15 , wherein RAS activity is decreased by administering an effective amount of a viral vector encoding an antisense molecule.  
     
     
         17 . The method of  claim 16 , wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         18 . The method of  claim 15 , wherein RAS activity is decreased by administering an effective amount of an antisense molecule which is an oligonucleotide.  
     
     
         19 . The method of  claim 16 , wherein the viral vector further comprises a nucleic acid encoding MDA-7 in expressible form.  
     
     
         20 . The method of  claim 11  wherein RAS activity is decreased by administering, to the cancer cell, an effective amount of an agent which inhibits a molecule selected from the group consisting of the epidermal growth factor receptor, RAF, MAPK kinase, and PI3 kinase.  
     
     
         21 . A method for inhibiting proliferation and inducing cell death in a population of pancreatic cancer cells having a mutated K-ras gene by (i) increasing the amount of the differentiation associated protein, MDA-7 and (ii) decreasing RAS activity within the population.  
     
     
         22 . The method of  claim 21  wherein the amount of MDA-7 is increased by introducing, into one or more cell of the population, a nucleic acid encoding MDA-7 protein in expressible form,  
     
     
         23 . The method of  claim 22  wherein the nucleic acid encoding MDA-7 protein is comprised in a viral vector.  
     
     
         24 . The method of  claim 23  wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         25 . The method of  claim 21  wherein RAS activity is decreased by administering, to the pancreatic cancer cell population, an effective amount of an anti-RAS agent, which may be, for example, an antisense molecule, a ribozyme, a precursor of a triple helix, or a farnesyl transferase inhibitor.  
     
     
         26 . The method of  claim 25 , wherein RAS activity is decreased by administering an effective amount of a viral vector encoding an antisense molecule.  
     
     
         27 . The method of  claim 26 , wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         28 . The method of  claim 25 , wherein RAS activity is decreased by administering an effective amount of an antisense molecule which is an oligonucleotide.  
     
     
         29 . The method of  claim 26 , wherein the viral vector further comprises a nucleic acid encoding MDA-7 in expressible form.  
     
     
         30 . The method of  claim 21  wherein RAS activity is decreased by-administering, to the pancreatic cancer cell population, an effective amount of an agent which inhibits a molecule selected from the group consisting of the epidermal growth factor receptor, RAF, MAPK kinase, MAP kinase and PI3 kinase.  
     
     
         31 . A method for inhibiting proliferation and/or inducing cell death of a pancreatic cancer cell having a mutated K-ras gene by (i) increasing the amount of the differentiation associated protein, MDA-7 and (ii) decreasing RAS activity in the pancreatic cancer cell.  
     
     
         32 . The method of  claim 31  wherein the amount of MDA-7 is increased by introducing, into the pancreatic cancer cell, a nucleic acid encoding MDA-7 protein in expressible form.  
     
     
         33 . The method of  claim 32  wherein the nucleic acid encoding MDA-7 protein is comprised in a viral vector.  
     
     
         34 . The method of  claim 33  wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         35 . The method of  claim 31  wherein RAS activity is decreased by administering, to the cancer cell, an effective amount of an anti-RAS agent, which may be, for example, an antisense molecule, a ribozyme, a precursor of a triple helix, or a farnesyl transferase inhibitor.  
     
     
         36 . The method of  claim 35 , wherein RAS activity is decreased by administering an effective amount of a viral vector encoding an antisense molecule.  
     
     
         37 . The method of  claim 36 , wherein the viral vector is selected from the group consisting of an adenovirus vector, an adeno-associated virus vector, a retrovirus vector, and a vaccinia virus vector.  
     
     
         38 . The method of  claim 35 , wherein RAS activity is decreased by administering an effective amount of an antisense molecule which is an oligonucleotide,  
     
     
         39 . The method of  claim 36 , wherein the viral vector further comprises a nucleic acid encoding MDA-7 in expressible form.  
     
     
         40 . The method of  claim 31  wherein RAS activity is decreased by administering, to the pancreatic cancer cell, an effective amount of an agent which inhibits a molecule selected from the group consisting of the epidermal growth factor receptor, RAF, MAPK kinase, MAP kinase and PI3 kinase.  
     
     
         41 . A method for treating a subject having pancreatic cancer, comprising, administering, to the subject, mounts of agents which are effective, in combination, in (i) increasing the amount of the differentiation associated protein, MDA-7 and (ii) decreasing RAS activity in cells of the pancreatic cancer.  
     
     
         42 . A method for treating a subject having pancreatic cancer, comprising, administering, to the subject, (a) a viral vector comprising an mda-7 gene in expressible form; and (b) an antisense ras oligonucleotide, in amounts which are effective, in combination, in (i) increasing the amount of the differentiation associated protein, MDA-7 and (ii) decreasing RAS activity in cells of the pancreatic cancer.  
     
     
         43 . A method for identifying a suitable cancer cell for treatment with mda-7/anti-RAS combination therapy, comprising (i) administering, to a test cancer cell, a first agent which increases the amount of MDA-7 protein in combination with a second agent that decreases RAS activity in the cancer cell; (ii) determining whether the cancer cell exhibits at least one characteristic of apoptosis; wherein the presence of a characteristic of apoptosis has a positive correlation with the suitability of the cancer cell for treatment with mda-7/anti-RAS combination therapy.  
     
     
         44 . A method for identifying a suitable cancer cell for treatment with mda-7/anti-RAS combination therapy, comprising (i) administering, to a culture of test cancer cells, a first agent which increases the amount of MDA-7 protein in combination with a second agent that decreases RAS activity; (ii) measuring the proliferation of cancer cells in the culture; and (iii) comparing the proliferation of cells measured in step (ii) with the proliferation of control cultures of the cancer cells in the presence of the same concentration of first agent or second agent, used alone; wherein if the combination of first and second agent results in a decrease in cell proliferation which is greater than the additive effect of the first agent and the second agent used alone, the cancer cell is suitable for treatment with mda-7 anti-RAS combination therapy.  
     
     
         45 . (canceled)  
     
     
         46 . A method of inhibiting the proliferation of a cancer cell, comprising exposing the cancer cell to an effective concentration of extracellular MDA-7 protein.  
     
     
         47 . The method of  claim 46 , wherein the extracellular MDA-7 protein is provided by introducing an isolated MDA-7 protein into a fluid which contacts the cancer cell.  
     
     
         48 . The method of  claim 46 , wherein the extracellular MDA-7 protein is produced by secretion from a cell into which an mda-7 gene, in expressible form, has been introduced,  
     
     
         49 . The method of  claim 48 , wherein the cell into which the mda-7 gene is introduced is a non-malignant cell.  
     
     
         50 . The method of  claim 49 , wherein the non-malignant cell is a hepatocyte.