CY can potentiate cellular and antibody-mediated immune responses, including responses to malignant tumors, in experimental animals. Under the auspices of this grant, we have shown that, in patients with advanced cancer, the administration of CY 300.mg/M2 IV results in: 1) augmentation of the development of delayed-type hypersensitivity (DTH) and antibody to the primary antigen, KLH; 2) selective impairment of non-specific suppressor T cell function of peripheral blood lymphocytes (PBL); and 3) augmentation of DTH to an autologous melanoma cell vaccine. The combination of vaccine with CY pretreatment (3 days before) resulted in complete regression of metastases in two patients with advanced melanoma. In this proposal for renewal, we have outlined an experimental plan to elucidate the mechanism of CY immunopotentiation and to further study its therapeutic implications. We will determine whether CY selectively depletes suppressor or suppressor-inducer T lymphocytes by studying PBL composition with the newly-developed monoclonal antibodies, anti-Leu 8 and anti-Leu 15. We will use PBL from KLH-injected melanoma patients to determine whether CY inhibits KLH-specific suppressor T cells. The approach will be to use the KLH lymphoproliferative response to assay: 1) suppressor cells present in peripheral blood and 2) suppressors that can be generated in vitro. We will also test a novel technique for demonstrating KLH-specific suppressor cells - the local adoptive transfer of DTH. We will further study CY-potentiated immunity to human tumor-restricted antigens by administering CY followed by an autologous melanoma cell vaccine. We will test DTH, autologous mixed lymphocyte-tumor cell (AMLTC) (proliferative) response, lymphocyte cytotoxicity, and antibody response. Specificity will be addressed by testing with certain defined melanoma-associated antigens, e.g., the melanoma-associated proteoglycan. We will further define the therapeutic efficacy of CY + vaccine by conducting a phase II trial in melanoma patients with clinically-apparent, but limited metastatic disease. Finally, we will determine whether CY directly inhibits tumor-directed suppressor cells by using the same strategy that was outlined for KLH-specific suppressors. Future directions of this work include: 1) substitution of defined melanoma-restricted antigens for intact cells in the treatment of grossly metastatic and sub-clinically metastatic melanoma; 2) treatment of patients with colorectal cancer by immunization with carcinoembryonic antigen preceded by CY; and 3) demonstration of melanoma-directed suppressor cells in patients with stage II disease.
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