Immunization of syngeneic mice against malignant melanoma with subcellular membrane fractions of a bromodeoxyuridine-grown variant

Abstract

Syngeneic C57BL/6 mice immunized with non-tumorigenic B16 melanoma cells and crude membrane fractions are able to reject challenge with the tumorigenic B559 parent clone. The immunogenic C3471 variant was derived from the malignant B559 clone by continuous growth in the presence of 1 μg 5-bromodeoxyuridine (BrdUrd)/ml. Fifty-one percent (35 of 69) of mice immunized by three inoculations of 106 cell equivalents of C3471 crude membranes (CM) isolated by nitrogen cavitation remained tumor-free for at least 50 days after challenge with a tumorigenic dose of B559 cells. This compares favorably with the virtually 100% protection evoked by 106 viable C3471 cells. For those CM-immunized mice failing to reject B559 challenge, the mean latent period for tumor formation was significantly increased (P≥0.001) over controls. In addition, mice immunized with cultured C3471 cells were able to reject, with equal efficiency, challenge with either cultured or tumor-derived B559 cells, indicating that the immunogen(s) present on C3471 cells and CMs was (were) not a tissue culture artifact. Freshly prepared syngeneic fascia cells and membranes as well as CM prepared from cultured malignant B559 cells had no tumor rejection activity. In vivo tumor rejection activity in plasma membrane vesicles prepared from C3471 cells by formaldehyde treatment also demonstrated tumor rejection activity. The host response to CMs, as to C3471 cells, could be transferred by lymphoid cells from mice immunized with C3471 CMs or cells. Co-injection of leucocytes from CM-immunized mice together with a tumorigenic dose of B559 cells into immunocompetent syngeneic mice resulted in abrogation of B559 tumorigenicity. The tumor rejection antigen(s) induced or increased by growth in BrdUrd has not yet been characterized biochemically, but is likely to involve a cell surface component common to both cell types and is retained by crude membrane fractions. The use of subcellular fractions from a spontaneous melanoma grown with BrdUrd, which elicits immunity against the malignant tumor, represents a model with immunoprophylactic potential for human neoplasia.