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Stable transformants of Saos-2 cells that contain the luciferase reporter gene under the control of the human p16INK4a transcriptional regulatory region were established, and were used to identify growth-inhibiting substances from culture broths of actinomycetes and extracts of plants. Among the active substances so far identified were an aclacinomycin-derivative, cenerubin B, and a cardiac glycoside, periplocin. These substances inhibited growth of normal human fibroblasts, and induced senescent phenotypes including a flattened morphology and increased acidic beta-galactosidase activity, although the activities of their derivatives to induce p16CDKN2 and beta-galactosidase did not coincided with each other. These results suggest that the reporter system using the p16CDKN2 transcriptional regulatory region is a useful means for screening growth inhibiting substances that are potential anti-tumor agents.

Induction of p16INK4a Transcription and of Cellular Senescence by Aclacinomycin-Derivatives and Cardiac Glycosides.