Characterization of tumor cell heterogeneity of a murine leukemia cell line (L1210) in response to arabinosylcytosine: quantitation using a computerized image analysis system.

Cytosine arabinoside (Ara-C) is one of the most effective drugs in inducing remission in acute nonlymphocytic leukemia (ANLL) patients. However, the high recurrence rate indicates that a subpopulation of leukemic cells escapes drug effect. This cellular heterogeneity in drug response may play a major role in chemotherapeutic outcome. We have recently developed the individual colony-formation assay (ICFA) to study drug effects on the kinetics of proliferation of individual cells and their progeny. Thus parameters of proliferation are calculated for individual colonies. Three categories of drug responses were defined, including immediate growth cessation, delayed growth cessation (growth stops several days after drug exposure) and growth slowdown (logarithmic growth at a reduced rate compared to control). In the experiments included in this report, murine leukemia (L1210) cells were exposed to various concentrations of Ara-C for 1, 6 or 24 hours, and their responses quantified. Regardless of the Ara-C concentration or exposure time, subpopulations of cells were observed in each of the three response categories: immediate or delayed arrest or growth slowdown. As expected, the fraction of cells exhibiting immediate growth cessation generally increased with increasing drug dose and was markedly increased with longer exposure time. Delayed arrest was most prevalent at intermediate drug concentrations at all exposure times. If exposure was limited to 1 hour, at least 30% of cells continued to grow, although at a reduced rate (71% control rate after exposure to 1 mM Ara-C). This limited effect was paralleled by saturation of Ara-C triphosphate (Ara-CTP) formation. Six-hour exposure left at least 6.4% of cells growing, with an average rate of 45% of control. Under these conditions, no saturation in Ara-CTP formation was observed. Even 24-hour exposure to 5 microM Ara-C left 4.8% of colonies growing, at 42% of control rate. Thus a subpopulation of cells continued to grow even after 24-hour exposure to a relatively high concentration of Ara-C. Surviving, but slowly growing, cells may represent a previously unrecognized population that may contribute to therapeutic failure.