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In the present study, we induced melanomas in C57BL/6 mice by a single application of 7,12-dimethylbenz(a)anthracene to the scapular region of 4-day-old mice, followed by twice-weekly applications of croton oil. Of 20 mice treated, melanomas arose in two female littermates. The first melanoma (JB/MS) arose 16 weeks after initiation of treatment, and the second melanoma (JB/RH) arose 23 weeks later. The melanomas maintained their melanotic appearance after s.c. transplantation to normal C57BL/6 mice and metastasized spontaneously in the transplant recipients. To our knowledge, these are the first melanomas to have been induced in C57BL/6 mice since the B16 melanoma arose spontaneously in 1954. We feel that the JB/MS and JB/RH melanomas provide an excellent comparative system for studies done with the B16 melanoma. These melanomas of recent origin will also facilitate the investigation of biological, immunological, and biochemical parameters that influence the growth and metastasis of malignant melanomas.

This study was designed to evaluate the phenotypic stability of the carcinogen-induced JB/MS and JB/RH melanomas. The JB/MS melanoma maintained its original slow-growing melanotic phenotype during in vivo passage over a 2-yr period. However, both melanin production and tumorigenicity decreased rapidly after propagation of JB/MS cells in vitro. The JB/RH melanoma became essentially amelanotic after the second transplantation in vivo. Cultured JB/RH cells produced tumors identical to those obtained by transplantation of JB/RH tumor fragments. However, after propagation in vitro for 2 yr, the JB/RH cell line decreased in tumorigenicity, requiring 10 times as many cells to produce tumors in C57BL/6 mice as did the original cell line. The JB/RH melanoma was highly immunogenic in syngeneic C57BL/6 mice, and passage of JB/RH cells through immunized mice resulted in tumors that were significantly more tumorigenic in normal mice than were JB/RH cells that had been passed through either nude or sublethally irradiated mice. These results indicate that, in studies of primary mouse melanomas, it is essential to: (a) limit the number of tumor passages; (b) choose methods of propagation that will preserve the original phenotype; and (c) distinguish those properties produced by technical manipulation from those produced by true tumor progression.

Sinclair swine melanoma usually regresses in vivo. In the present study, swine melanoma cells were adapted to long-term growth in culture. The morphology of cultured melanoma cells ranged from dendritic to cuboidal, similar to that described for human melanoma cells. Doubling times of the swine melanoma cells were also similar to those of human melanoma cells in vitro. 3,4-Dihydroxy-L-phenylalanine oxidase-positive cells were detected by light microscopy, and melanin and premelanosomes were detected by electron microscopy. Cell cultures could be propagated from progressing, partially regressed, and primary cutaneous lesions, as well as from visceral metastases. Thus, it appears that, under these cell culture conditions, Sinclair swine melanoma cells can be adapted to prolonged growth in vitro.