Characterization of human cells transformed by chemical and physical carcinogens in vitro

Summary Several different classes of chemical carcinogens induced the transformation of human fibroblasts grown in vitro. Characteristics of the events that occur from time of treatment through the expression of neoplastic transformation are presented. The S-phase appeared to be the portion of the cell cycle most vulnerable to insult. Staging of the cells by blocking them in G₁ before releasing them to proceed through scheduled DNA synthesis (S) was required to induce reproducible transformation. Compounds such as insulin were added to the cells upon release from the block to sensitize the cells to the carcinogen that was added during S. Growth of the transformed cells as distinct from nontransformed cells was promoted by growth in medium supplemented with 8X nonessential amino acids. Carcinogen-treated cells in the early stage of transformation exhibited abnormal colony morphology and were able to grow at 41°C, in air atmosphere, and in medium supplemented with only 1% serum. In addition, the transformed cells were insensitive to KB cell lysate and exhibited density independent, as well as anchorage independent, growth (i.e., growth in 0.33% agar). Cells that grew in soft agar also produced undifferentiated mesenchymal tumors in preirradiated nude mice. This work was supported in part by National Cancer Institute Grant ROI-CA-25907 and Air Force Office of Scientific Research Grant F49620-77-C0110 and EPA-R806638. The hydrazine compounds were furnished by Ms. Marilyn George and Dr. Kenneth Back, AFSOR Toxicology Division, Wright Patterson Air Force Base, Dayton, OH. The hydroxylate and phenyl napthylamines were furnished by Dr. Fred Kadlubar, Division of Chemical Carcinogenesis at the National Center for Toxicological Research, Jefferson, AR.