Adaptational origin of some purine-analogue resistant phenotypes in cultured mammalian cells

Abstract • In an attempt to elucidate further the reasons for phenotypic diversity amongst purine-analogue resistant clones, Chinese hamster and mouse lymphoma cells were exposed to single and multistep selection procedures. The phenotypes of cells during exposure to, and on removal from, continuous low dose selection were determined and compared with those of cells surviving short-term exposure to high selective doses. • On exposure to low doses of 8-azaguanine (8AZ), of 6-thioguanine (6TG), HAT + first step resistance developed within 2–3 days. First-step resistant lines showed no reduction in either HGPRT activity or [ 14 C]hypoxanthine utilisation, consistent with their retained ability to grow in HAT. They were unstable when grown in the absence of selective pressure and gradually lost purine-analogue resistance over a period of 6–9 weeks. • Continued growth in the presence of low doses 8AZ resulted in progression to a higher level of resistance accompanied by a loss of ability to grow in HAT, a reduction in HGPRT activity, and in [ 14 C]hypoxanthine utilisation. • Second step HAT − resistant lines gradually regained their ability to grow in HAT when cultured in the absence of selective pressure over 15–20 weeks. Recovery of ability to grow in HAT was accompanied by a rise in [ 14 C]hypoxanthine incorporation and a reduction in level of resistance. The instability of these resistant lines, in contrast to those isolated after single high doses, suggests that they are non-mutants and are the result of drug-induced modifications in cellular biochemistry. The similarity of the phenotypes of these lines with those of many resistant lines described by others suggests that many other purine-analogue resistant lines may be of non-mutant origin. The implicatiosn of these observations are discussed.