Interaction of maytansinoids with MCF7 cells.

A. Binding of the maytansinoid ³[H]-S-methyl DM1 to the cells, a representative experiment. Cells were incubated in culture medium with various concentrations of ³[H]-S-methyl DM1 at 37° C for 4 h (equilibrium was reached within 4 h) with (squares) or without (circles) an excess (5 μM) of non-labeled S-methyl DM1, washed two times with medium, and cell-associated radioactivity was measured on a scintillation counter. The specific binding (triangles) was calculated as the difference between the two values. Cell-associated ³[H]-S-methyl DM1 was fully retained by cells after multiple washes, and thus the equilibrium was not distorted during the washes. B. Scatchard plot of A. C. The relationship between the cytotoxicity (IC₅₀) of a maytansinoid and its apparent affinity (ED₅₀) to cells. The IC₅₀ values were determined in clonogenic assays after a 4-h exposure of cells to a maytansinoid. Each point (the mean ± standard error) is the result of three independent experiments. The ED₅₀ is a concentration of a maytansinoid that decreased the binding of a trace amount of ³[H]-S-methyl DM1 by 50% in a competition binding assay. D. Structures of the maytansinoids. E. Efflux of ³[H]-S-methyl DM1 from MCF7 cells at 37° C in medium (filled symbols), or in the presence of an excess of non-labeled S-methyl DM1 (open symbols). Triangles and squares represent two independent experiments.