IMMUNOTOXIN AND ONCOTOXIN THERAPY OF CANCER CELLS

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Pseudomonas exotoxin (PE) or genetically modified forms of PE have been attached to monoclonal antibodies (mAbs) or growth factors to create cell-specific cytotoxic agents. Mutant PE molecules in which domain I has been replaced by growth factors, CD4 or single chain antibody combining regions have been created by gene fusion, the chimeric proteins produced in E. coli, and purified to near homogeneity. We have constructed and studied the activity of the following chimeric toxins: TGFalpha-PE40, IL2-PE40; IL4-PE40, IL6-PE40, IGF1-PE40, acidic FGF-PE40, CD4-PE40, antiTac(Fv)-PE40 and antitransferrin(Fv)-PE40. TGFalpha-PE40 which kills cells with EGF receptors has now been shown to have an antitumor effect in mice when injected I.P. against intraperitoneal tumors and against subcutaneous tumors. IL2-PE40 is very effective in killing mouse and rat cells with IL2 receptors but is less active against primate and human cells. To overcome this deficiency, a single chain immunotoxin anti-Tac(Fv)-PE40 was constructed which is extremely cytotoxic to human and primate cells containing IL2 receptors including cells directly isolated from patients with adult T cell leukemia. IL6-PE40 and a variant, IL6-PE664GIu, is cytotoxic to several myeloma cells lines and hepatoma cell lines; cells with as few as 400 receptors per cell can be killed. Because tumors are dependent on a new blood supply, we constructed acidic FGF-PE40 and PE664Glu which are cytotoxic to FGF receptor bearing cells. CD4-PE40 has been tested in combination with AZT and shown to act synergistically to arrest the spread of HIV infection in culture. A monoclonal antibody, B3, reactive with many colon, breast, lung and ovary tumors has been isolated, the genes encoding the variable regions cloned and a single chain immunotoxin made. B3(Fv)-PE40 has a strong antitumor effect against human tumors growing in nude mice and is currently undergoing preclinical development. Pseudomonas exotoxin mutants with increased activity have been created by changing the carboxy terminus from REDLK to KDEL. These molecules are two to ten-fold more cytotoxic to target cells. To pen-nit prolonged therapy with immunotoxins which are very immunogenic, the effect of an immunosuppressive agent, 15deoxyspergualin, has been studied and the drug shown to suppress primary antibody formation to PE in mice. Modified clones of PE have been constructed with foreign polypeptides inserted into the translocating domain of PE which introduces these inserts into the cytosol of target cells.