Chemical and biological stability of anticancer drugs used in a human tumor clonogenic assay.

Abstract

Human tumor clonogenic assays (HTCA) are being used to evaluate the chemosensitivity of human cancers to both standard and experimental anticancer drugs as well as to predict clinical tumor response and resistance to these agents. To enable us to design and accurately interpret drug assay data we quantitated the chemical and biological stability characteristics of various cell cycle-specific and cell cycle-nonspecific drugs commonly used in our laboratory for chemosensitivity testing in the HTCA. Aliquots of the culture media were obtained immediately after drug addition and after 6 and 24 h and 2, 4, and 10 days of incubation, and then frozen in liquid nitrogen for later chemical (HPLC or RIA) and biological (HTCA) assay. Adriamycin, actinomycin D, bisantrene, bleomycin, and vinblastine retained 80-100% of their biological activity against human tumor colony-forming units even after 10 days of incubation in culture media. In contrast, etoposide lost 60% of its in vitro antitumor activity over the same period. There was no loss in the chemical concentration of actinomycin D. bisantrene, bleomycin, or vinblastine in the culture media, but etoposide concentrations as measured by HPLC decreased to zero over 10 days. We conclude that the HTCA can be used as a simple test for biological stability of new investigational agents prior to the development of adequate chemical assay methodology, and that it can help clarify whether a drug's in vitro antitumor activity is due to the parent compound or to a degradation product.