IDENTIFICATION OF A K-B-RESTRICTED CTL EPITOPE OF BETA-GALACTOSIDASE - POTENTIAL USE IN DEVELOPMENT OF IMMUNIZATION PROTOCOLS FOR SELF-ANTIGENS

The use of recombinant and synthetic vaccines in the treatment of canc er has recently been explored using model tumor associated antigens (T AA), many of which do not model the immunological state of affairs in which the TAA is expressed by normal tissues. One potentially useful m odel Ag is beta-galactosidase (beta-gal). Because the activity of this enzyme is so easily detectable, this gene has been inserted into a la rge number of recombinant viruses and tumors useful to the cancer vacc inologist. In addition, numerous transgenic mouse colonies that have t issue-specific expression of beta-gal have been developed, enabling th e modeling of tolerance to ''self'' Ags. Since most of these mice have an H-2(b) background, we generated cytotoxic T lymphocytes (CTL) capa ble of recognizing beta-gal-expressing tumor cells of C57BL\6 origin a nd have determined that their restriction element is the K-b molecule. Using an allele-specific epitope forecast to generate a panel of cand idate peptides, we have determined that the K-b-restricted sequence is DAPIYTNV and corresponds to amino acids 96-103 of the intact beta-gal molecule. A recombinant vaccinia virus (rVV- pes beta-gal(96-103)) wa s constructed that encoded the peptide epitope preceded by an endoplas mic reticulum insertion signal sequence. Tumor cells infected with thi s rVV were recognized by the original CTL that had been used to identi fy the epitope. Furthermore, splenocytes of mice immunized with a rVV encoding the full-length beta-gal molecule and restimulated with the D APIYTNV peptide specifically recognized tumor cells expressing beta-ga l. The identification of this immunogenic beta-gal sequence enables th e modeling of immunization strategies in animal models of malignant di sease in which the target antigen is a ''self'' protein.