The human prostatic epithelial cell line BPH-1 is normally nontumorigenic in nude mice. The present report demonstrates that this cell line can be permanently transformed by its microenvironment to become tumorigenic. The establishment of a series of tumorigenic sublines based on this parental cell line is described. BPH-1 cells were induced to form tumors either by recombination with human prostatic carcinoma-associated fibroblasts (CAFs) or by exposure to carcinogenic doses of testosterone and estradiol (T+E2) after recombination with rat urogenital sinus mesenchyme. Epithelial cells isolated from these tumors were established as cell strains in culture. When regrafted to nude mouse hosts epithelial cells isolated from CAF- or T+E2-induced tumors were found to be consistently tumorigenic even in the absence of CAF or T+E2. The T+E2-induced cell strains have been designated BPH1^TETD-A and -B and the CAF-induced strains are designated BPH1^CAFTD-01 through -08. In vitro, the cells had an epithelial morphology with a less well-defined cobblestone pattern than the parental line. They express SV40 large T antigen, confirming their derivation from the parental BPH-1 line. The BPH1^CAFTD strains formed colonies in soft agar, whereas the parental BPH-1 cells and the BPH1^TETD sublines did not. There was no immunocytochemically detectable expression of androgen (AR), α-estrogen (ERα), or progesterone (PR) receptors by the parental BPH-1 cell line or by any of the tumor-derived cell strains. The cells uniformly coexpressed both basal and luminal cell-type cytokeratins and the basal cell marker p63. When grafted beneath the renal capsule of athymic mouse hosts, all of the tumor-derived cell strains consistently formed tumors. These were predominantly poorly or moderately differentiated squamous or adenosquamous tumors, similar in organization to the primary tumors from which the cell strains were derived. The cell strains continued to express both basal- and luminal-type cytokeratins in vivo. Some of the cell strains also coexpressed vimentin. E-cadherin expression was absent from many of the cells, although patches of cells expressing this marker were seen. The cells continued to express SV40T antigen. These cell strains, which are all derived from a common nontumorigenic progenitor, represent a useful resource for examining genetic and phenotypic changes during carcinogenesis.