MHC CLASS II-INDEPENDENT, V-BETA-SPECIFIC ACTIVATION OF T-CELLS BY SUPERANTIGEN MUTANTS FUSED TO ANTITUMOR F-AB FRAGMENTS - IMPLICATIONS FOR USE IN TREATMENT OF HUMAN COLON-CARCINOMA

Genetically engineered fusion proteins of the superantigen staphylococ cal enterotoxin A (SEA) and tumor-reactive monoclonal antibodies, C215 F(ab)-SEA and C242F(ab)-SEA, have been generated and shown to be effec tive in mediating superantigen-antibody directed cellular cytotoxicity against human carcinoma cells expressing the CA215 or CA242 antigens in an MHC class II-independent manner. In an attempt to reduce the in vivo toxicity of superantigen administration, alanine substitution mut ations in SEA at residues F47 and D227 that affect SEA binding to clas s II molecules have been created and genetically linked to C215F(ab) o r C242F(ab). The purpose of this study was to determine whether these F-ab-SEA mutant fusion proteins, that have low MHC class II binding af finities, were still able to stimulate human T cells in a V beta-speci fic manner in the presence or absence of MHC class II molecules. The S EA wt- and SEA-D227A-based fusion proteins shared the ability to activ ate V beta 5.2-, V beta 6-, V beta 7-, V beta 9- and V beta 18-bearing T cells, whereas F-ab-SEA-F47A protein activated only V beta 6- and V beta 7-bearing T cells. The fusion of F-ab fragments onto SEA wt, SEA -F47A or SEA-D227A had no effect on the V beta specificity of these su perantigens. F-ab fusion proteins containing either SEA wt or SEA muta nts were presented, in the absence of class II molecules, by CHO cells transfected with CA215 and CD80 and all induced the expansion of only V beta 6-, V beta 7- and V beta 18-bearing T cells. F-ab-SEA mutant f usion proteins may provide attenuated therapeutic agents that, while s till able to specifically target high affinity T cells for MHC class I I-independent local tumor killing, will not induce excessive systemic toxicity.