Lethal and kinetic response of cultured human lymphoid cells to melphalan

Melphalan was analyzed for its lethal and kinetic effects in a human lymphoma cell line. Cell kill was assessed by the reduction in colony formation. One hour treatment of asynchronous exponentially growing cell populations with increasing concentrations of melphalan resulted in a threshold exponential survival curve typical of the effects of alkylating agents and ionizing radiation on this cell line (D(q) = 0.5 μg/ml, 1 hr; and D(o) = 1.5 μg/ml, 1 hr). Prolonged exposure of asynchronous cells to 2.5 μg/ml of melphalan resulted in an initial (15 hr) logarithmic decrease in survival (3 log decades) followed by a plateau region with no increased killing in spite of continuous exposure to the drug. Synchronized cells showed moderate cycle stage dependent sensitivity to melphalan with cells in early G₁ phase being most sensitive (tenfold) to the lethal effect of the drug. The effects on cell cycle progression were analyzed employing sequential DNA histography, provided by pulse cytophotometry. Melphalan induced a concentration and exposure time dependent block in G₂ phase. Both degree and manifestation of the G₂ block varied with the particular stage of the cell cycle in which short term treatment had been conducted. Thus, G₁ cells were most effectively blocked in the immediate life cycle. Cells treated in S and G₂ phase underwent cell division without delay; however, the G₂ traverse of the immediate progeny was prolonged. Treatment with a concentration > 0.5 μg/ml for > 24 hr caused retardation of S phase transit, accounting for a delay in the onset of G₂ accumulation. Progression through G₁ into S phase was not affected by treatment with a concentration up to 2.5 μg/ml. There was no correlation between the perturbation of cycle progression with cell survival assessed by colony formation. However, manifestation of G₂ block in the immediate lifespan resulted from treatment in G₁ phase which was also most sensitive to the lethal effect of melphalan.