Cytotoxic CD4⁺ lymphocyte clones reactive with melanoma: The role of HLA and accessory molecules

Tumor-infiltrating lymphocytes (TILs) were isolated from a patient with disseminated melanoma during active specific immunotherapy. Purified CD8⁺ TILs lysed autologous melanoma cells and one of three allogeneic melanoma cell lines. CD4⁺ TILs also lysed melanoma cell lines, but cytotoxicity was considerably lower than with CD8⁺ cultures. Neither the CD4⁺ nor the CD8⁺ culture lysed autologous EBV-transformed B cells in a 6- or 14-hr assay. These data were confirmed using CD4⁺ or CD8⁺ TIL clones. Coculture of autologous melanoma cells with rIFN-γ (100 U/ml) for 24 hr significantly enhanced cytolysis by CD4⁺ clones from 1.5-fold to more than 10-fold. This enhancement was associated in part with an increased cell surface expression of HLA class II molecules, because lysis by seven of nine CD4⁺ clones was almost completely blocked by anti-HLA class II MAb L227. Interestingly, both anti-HLA class I and anti-HLA class II MAbs blocked cytolysis by four of the nine CD4⁺ clones. Lysis by two CD4⁺ clones was blocked by anti-HLA class I MAb w6/32, but not by L227. The CD4 molecule was involved in the binding to HLA class II molecules or in signal transduction, since anti-CD4 MAb consistently inhibited cytotoxicity. Blocking of the known accessory molecule-ligand interactions CD11a,CD18 (LFA-1)-CD54 (ICAM-1 or -2) and CD2-CD58 (LFA-3) demonstrated the involvement of both interactions in the cytotoxicity by CD4⁺ clones. CD4⁺ CTLs may be important effector cells in vivo against melanomas expressing HLA class I and/or class II molecules.