Spontaneous, mutagen-induced and adenovirus-induced anchorage independent tumorigenic variants of mouse cells

Normal C57 Black mouse embryo cells did not form colonies in agarose, but rare variant (ar+) cells able to grow in agarose were detected. Fluctuation analysis showed that ar+ variants arose by spontaneous mutation in the cultured cells. The frequency of ar+ variants was increased by treating cells with N-methyl-N'nitro-N-nitrosoguanidine or ethyl methane sulphonate, or by abortive infection by human adenovirus type 5. Induced ar+ cells were fibroblastic; most grew slowly and had slightly reduced saturation density and increased serum requirement, but formed colonies in agarose. Fourteen of twenty ar+ clones induced by Ad5 were T antigen negative and two of these were also negative when tested for viral DNA. Six clones contained a few cells that were T antigen positive when first tested, but were negative when retested later. The ar+ variants were tumorigenic in athymic and in normal syngeneic mice. The results suggest that the ar+ phenotype can arise by spontaneous or chemically-induced mutation, and can be induced by adenovirus by a process different from classical transformation.