Vincristine (VCR) is an active agent in the treatment of acute lymphoblastic leukemia (ALL). We evaluated the relationship between the cytotoxic activity of VCR and the degree of VCR saturation of cellular drug binding sites, using the MOLT-3 ALL cell line. When MOLT-3 cells at a density of 1 x 10(6) or 1 x 10(8) cells/mL of pH-controlled medium were exposed to VCR for 1 hr, its cytotoxic activity on cells at high density was 10-fold less than on cells at low density (inoculum effect). The number of VCR binding sites measured by Scatchard analysis was 9.25 x 10(6)/cell. At high cell density, the saturation of VCR binding sites was one log order less than that at low density. Irrespective of cell density, curves of cell-kill versus the degree of VCR saturation of the cellular binding sites overlapped each other. Minimal cytotoxic activity was observed at 0.3% VCR saturation, and nearly maximal cytotoxic activity occurred at about 25% saturation, with the Ic50 at about 4% saturation. These data show that the VCR-induced cell-kill effect is dependent on the degree of saturation of VCR binding sites rather than on the extracellular VCR concentration. The lesser cell-kill on cells at high density can be explained by the lack of drug molecules to sufficiently saturate cellular binding sites. This phenomenon may be responsible, at least in part, for the poor chemotherapeutic outcome of ALL patients with high leukocyte counts at presentation.