Stromal cells isolated from normal human endometrium were cotransfected in primary culture with pSVc-m.vc, a plasmid containing a truncated c-myc gene regulated by simian virus 40 promoter, and pRSVneo, a plasmid containing a neomycin resistance gene regulated by Rous sarcoma virus (RSV) promoter. These cells demonstrated properties of transformed cells in vitro, including altered morphology, focus formation, anchorage-independent growth, chromosomal alterations, and tumor formation in athymic mice. When these cells were treated subsequently with a direct-acting carcinogen, N-methyl-N′-nitro-N-nitrosoguanidine, they demonstrated higher colony-forming efficiency in soft agar and reduced tumor latency. Cells transfected with pRSVneo alone exhibited some properties associated with neoplastic transformation, including altered morphology and formation of colonies in soft agar. It is presumed that in normal cells transfected with pRSVnpo, RSV long terminal repeats activated cellular genes that normally regulate growth of human endometrial stromal cells. © 1990 S. Karger AG, Basel.