Comparison of in vitro methods to determine drug-induced cell lethality

In proliferating cell populations, the inability to reproduce indefinitely is the only relevant criterion to assess cell lethality. The in vitro colony formation technique (CF) used to determine reproductive death is, however, too slow and has several technical limitations. For finding suitable, more rapid techniques that assessed drug-induced cell killing, a human lymphoma cell line was exposed in vitro to increasing concentrations of adriamycin, bleomycin, and 1,3-bis(2-chloroethyl)-1-nitrosourea for 1 hr. Survival was assayed immediately after treatment and at regular intervals thereafter. Data from CF were compared to those resulting from the following tests: doubling time, labeling index, dye exclusion, 51Cr release, and rate of [3H]thymidine uptake (scintillation index). Dye exclusion and 51Cr release failed to demonstrate any killing effect for the 3 drugs. The percentage of killing calculated from doubling time determinations, although dose dependent, failed to correlate with CF. Scintillation and labeling index values displayed similar temporal fluctuations but were not clearly dose dependent and did not correlate with CF. Thus, CF appears as the most reliable, dose-dependent index of cell lethality. Tests that measure metabolic death grossly overestimate or underestimate killing activity induced by 3 of the most effective antitumor drugs.