G(D3) VACCINES FOR MELANOMA - SUPERIOR IMMUNOGENICITY OF KEYHOLE LIMPET HEMOCYANIN CONJUGATE VACCINES

Cell surface gangliosides show altered patterns of expression as a con sequence of malignant transformation and have therefore been of intere st as potential targets for immunotherapy, including vaccine construct ion. One obstacle has been that some of the gangliosides that are over expressed in human cancers are poorly immunogenic in humans. A case in point is G(D3), a prominent ganglioside of human malignant melanoma. Using an approach that has been effective in the construction of bacte rial carbohydrate vaccines, we have succeeded in increasing the immuno genicity of G(D3) in the mouse by conjugating the ganglioside with imm unogenic carriers. Several conjugation methods were used. The optimal procedure involved ozone cleavage of the double bond of G(D3) in the c eramide backbone, introducing an aldehyde group, and coupling to amino lysyl groups of proteins by reductive amination. Conjugates were const ructed with a synthetic multiple antigenic peptide expressing repeats of a malarial T-cell epitope, outer membrane proteins of Neisseria men ingitidis, cationized bovine serum albumin, keyhole limpet hemocyanin, and polylysine. Mice immunized with these conjugates showed a stronge r antibody response to G(D3) than mice immunized with unconjugated G(D 3). The strongest response was observed in mice immunized with the key hole limpet hemocyanin conjugate of the G(D3) aldehyde derivative and the adjuvant QS-21. These mice showed not only a long-lasting high-tit er IgM response but also a consistent high-titer IgG response (predomi nantly IgG1), indicating recruitment of T-cell help, although the tite rs of IgM and IgG antibodies following booster immunizations were not as high as they are in the response to classical T-cell-dependent anti gens. This method is applicable to other gangliosides, and it may be u seful in the construction of immunogenic ganglioside vaccines for the immunotherapy of human cancers expressing gangliosides on their cell s urface.