Melanoma-specific lysis by cloned CD4⁺ and CD8⁺ T cells from actively immunized melanoma patients

We have previously shown that T cells from actively immunized melanoma patients, stimulated in 9-day mixed lymphocyte-tumor cell (MLTC) bulk cultures with allogeneic melanoma cells, lysed various melanomas but not lung carcinoma or lymphoid tumor cells. To further characterize these T cells, we cloned MLTC-stimulated T cells from nine melanoma patients. One hundred seventeen clones were generated from peripheral blood lymphocytes (PBL), Of which 64 were CD4⁺ and 53 were CD8⁺ From two of the patients we also grew 18 CD4⁺ clones from tumor infiltrating lymphocytes (TIL), which were propagated from one patient (WB) with only autologous tumor cells. One hundred clones were cytotoxic to one or more melanoma targets expressing HLA-A2 (or -A28) and/or HLA-B12 (or -B44) molecules, matching the T cell phenotype in at least that one allele. Melanoma cells completely lacking detectable surface major histocompatibility complex (MHC) antigens were not Iysed by any of 25 clones tested. However, we found a considerable number of clones that lysed melanoma cells lacking the 'appropriate' class of MHC. Thus, 11 of 58 CD4⁺ clones lysed the HLA class II-negative M-2 line, while 10 of 30 CD8⁺ clones Iysed the HLA class I-negative M-8 melanoma. Reactivity against the allogeneic melanoma M-7 and its lymphoid counterpart LCL-7 was analyzed with PBL and TIL clones from patient WB. Four of 5 CD4⁺ and 4 of 5 CD8⁺ clones lysed the melanoma cell lines but not the HLA-matched B cells, confirming their specificity for melanoma. Since melanoma M-7 lacked HLA class II antigens, both the CD4⁺ and the CD8⁺ clones apparently recognized melanoma-associated antigens presented in an MHC class I context. No significant difference was observed between the reactivity patterns of clones derived from PBL or TIL, perhaps because of the prior in vivo immunization. Cloned T cells propagated in the presence of either allogeneic or autologous melanomas thus lysed both autologous and allogeneic melanomas sharing at least one MHC allele with the T cells. In 8 of 9 patients the MHC molecules in common were HLA-A2/A28, or HLA-B12/B44, or both.