GENERATION OF TUMOR SPECIFIC T CELLS THAT RECOGNIZE A SHARED ACUTE MYELOGENOUS LEUKEMIA ANTIGEN.

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GENERATION OF TUMOR SPECIFIC T CELLS THAT RECOGNIZE A SHARED ACUTE MYELOGENOUS LEUKEMIA ANTIGEN.
Myra Coppage MS , Todd Belanger MS , Maurice Zauderer PhD and Deepak Sahasrabudhe MD . Rochester NY, University of Rochester Medical Center, 14642, Department of Pathology ; Rochester NY, University of Rochester Medical Center, 14642, James P. Wilmot Cancer Center and Rochester NY, Vaccinex, Inc, 14642 .

Identification of immunologically relevant tumor antigens has been hampered by the difficulty of generating tumor-specific cytotoxic T cells (CTL). We present data demonstrating in vitro induction of autologous acute myelogenous leukemia (AML)-specific CTL using tumor pulsed dendritic cells (DCs). Method: Peripheral blood leukemic blasts from an AML patient were cryopreserved. When the patient entered complete remission, PBL were cryopreserved, a B cell line (B-LCL) established, and monocyte precursors used to generate immature DCs. The PBL underwent three cycles of stimulation with DCs pulsed with UV-irradiated autologous leukemic blasts. T cell growth cytokines (IL-2 and IL-7) were added at the second stimulation. Prior to the third stimulation, the cultures were enriched for CD8+ cells by magnetic bead selection. Following the third stimulation the T cells were tested for tumor specific cytotoxicity by standard chromium release assay. Results: Low levels of AML blast specific lysis was observed in the early bulk cultures. T cells were cloned by limiting dilution; 22/1000 wells grew. Five of the 22 exhibited AML-specific production of IFNg by ELISA. One clone (5) was expanded, and activity assayed by cytokine ELISA against 8 different AML tumors, K562, and autologous B-LCL. The T cells secreted GM-CSF, TNFa, and IFNg when stimulated with AML blasts from 3 of 8 patients or cell lines, but not with K562 or autologous B-LCL. The three AML samples recognized by the CTL clone share a single HLA Class I antigen, HLA-A24. Conclusion: This strategy successfully produced an HLA-A24 restricted, CD8+ T cell clone that recognizes a shared AML antigen. Lethality-based selection method is being used to characterize this antigen.