Antigenic differences among B16 melanoma variants selected for their differing abilities to metastasize: A possible mechanism for effective adjuvant immunotherapy

Most C57BL/6J mice will develop pulmonary metastases four to six weeks after excision of B16 melanoma isografts and begin to die within 40 days. When applied in an adjuvant setting after isograft excision, specific B16 immune RNA (I‐RNA) therapy prevents pulmonary metastases and prolongs survival in 50% of treated animals. Since increased in vitro cell‐mediated cytotoxicity (CMC) against B16 targets has been demonstrated in animals whose survival is improved by B16 I‐RNA therapy, we have proposed that the treatment functioned, at least in part, by specifically altering host CMC in vivo. In this study, C57BL/6J splenocytes were specifically sensitized in vitro by B16 I‐RNA exposure and examined for cytotoxic effect on variants of B16 melanoma selected for their differing metastatic potential in vivo. F10 tumor targets (explanted from a B16 melanoma vairant producing frequent pulmonary metastases in vivo) were consistently more sensitive to specific in vitro cytotoxic effect than F1 target cells (from a B16 melanoma variant with low metastatic potential in vivo). Lung metastases (F1 mets) were explanted after intravenous (IV) injection of F1 and used as target cells in the in vitro CMC assays. F1 mets demonstrated greater cytotoxic effect than F1 targets after exposure to B16 I‐RNA‐treated syngeneic splenocytes. C57BL/6J mice were sacrificed at weekly intervals following adjuvant in vivo B16 I‐RNA therapy (after B16 isograft excision), and their splenocytes were shown to be consistently more cytotoxic in vitro to B16 and F10 than to F1 target cells. Splenocytes harvested from mice treated with I‐RNA specific to antigenically distinct Lewis lung carcinoma (3LL) after 3LL isograft excision had no cytotoxic effect on any of the B16 variants. When nonsensitized C57BL/6J splenocytes were examined for cytotoxic effect in natural killer (NK) assays or in NK inhibition assays, differences in cytotoxic sensitivity of B16, F10, F1, and F1 mets could not be demonstrated. We conclude, therefore, that specifically sensitized effector cells could distinguish antigenic differences among B16 variants selected for differing in vivo metastatic potiential. These differences were tumor specific and could be demonstrated by cytotoxic splenocytes after either in vivo B16 I‐RNA treatment or in vitro B16 I‐RNA exposure. The relationship of these antigenic differences to in vivo metastatic potential and to the effectiveness of adjuvant B16 I‐RNA therapy is discussed.