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 ind ucing remission in acute nonlymphocytic leukemia (ANLL) patients. Howe ver, the high recurrence rate indicates that a subpopulation of leukem ic cells escapes drug effect. This cellular heterogeneity in drug resp onse may play a major role in chemotherapeutic outcome. We have recent ly developed the individual colony-formation assay (ICFA) to study dru g effects on the kinetics of proliferation of individual cells and the ir progeny. Thus parameters of proliferation are calculated for indivi dual colonies. Three categories of drug responses were defined, includ ing immediate growth cessation, delayed growth cessation (growth stops several days after drug exposure) and growth slowdown (logarithmic gr owth at a reduced rate compared to control). In the experiments includ ed in this report, murine leukemia (L1210) cells were exposed to vario us concentrations of Ara-C for 1, 6 or 24 hours, and their responses q uantified. Regardless of the Ara-C concentration or exposure time, sub populations of cells were observed in each of the three response categ ories: immediate or delayed arrest or growth slowdown. As expected, th e fraction of cells exhibiting immediate growth cessation generally in creased with increasing drug dose and was markedly increased with long er exposure time. Delayed arrest was most prevalent at intermediate dr ug concentrations at all exposure times. If exposure was limited to 1 hour, at least 30% of cells continued to grow, although at a reduced r ate (71% control rate after exposure to 1 mM AraC). This limited effec t was paralleled by saturation of Ara-C triphosphate (Ara-CTP) formati on. Six-hour exposure left at least 6.4% of cells growing, with an ave rage rate of 45% of control. Under these conditions, no saturation in Ara-CTP formation was observed. Even 24-hour exposure to 5 mu M AraC l eft 4.8% of colonies growing, at 42% of control rate. Thus a subpopula tion of cells continued to grow even after 24-hour exposure to a relat ively high concentration of Ara-C. Surviving, but slowly growing, cell s may represent a previously unrecognized population that may contribu te to therapeutic failure.