Stimulation of cis-Diamminedichloroplatinum(II) Accumulation by Modulation of Passive Permeability with Genistein: An Altered Response in Accumulation-defective Resistant Cells1

The effect of the tyrosine kinase inhibitor genistein on the accumulation of cisplatin (DDP) was investigated in DDP-sensitive and -resistant human 2008 ovarian cardnoma cell lines. DDP accumulation after a 1-h exposure was maximally increased by concurrent 40 piM genistein. The maximal stimulation of accumulation was observed after 2 h of total genistein exposure and was 83 ± 13% (n = 5) higher than controls. With resistant C13* cells, however, the stimulation of accumulation was delayed until 4 h and was increased only 46 ± 18% compared to controls. Revertant RH4 cells that retained the accumulation defect behaved like the C13 cells. Genistein stimulated [3H]mannitol accumulation (a marker of passive permeability) by 43 ± 9% (n = 3) in 2008 cells, and the effect was maximal after 2 h of total genistein exposure. Changes in [3Hjmannitol accumulation in 2008 parent cells were highly correlated with DDP accumulation (r = 0.9010). These experiments also revealed that [3H]mannitol accumulation after 2 h in C13* cells was reduced 38% compared to 2008 cells, a decrease that reflected the DDP accumulation defect. Fluid-phase pinocytosis determined with lucifer yellow CH as a marker showed no difference between 2008 and C13* cells and no effect of genistein. Genistein was demonstrated to clearly inhibit protein-tyrosine phosphorylation initiated by the epidermal growth factor receptor kinase. Differences were noted in the phosphotyrosine pattern between the 2008 and C13* cells. Under the conditions that had the maximal effect on DDP accumulation in 2008 cells, genistein decreased the IC50 of DDP 8.2-fold in 2008 cells and 4.7-fold in C13* cells. We conclude that: (a) genistein stimulates DDP accumulation by modulating the passive permeability of the plasma membrane; (b) C13* cells are less permeable to passively diffusing small molecules, which offers a mechanism for the DDP accumulation defect without invoking carrier proteins; (C) the effect of tyrosine kinase inhibition on passive permeability is altered in C13 cells; and (d) pinocytosis contributes Received 2/14/95; revised 1/24/96; accepted 3/4/96. I Supported by Grant BE-175 from the American Cancer Society. 2 Present address: Dipartimento di Scienze Biomediche, Sezione di Chimica Biologica, Universit#{225} di Modena, via Campi 287. 41 100 Modena, Italy. 3 To whom requests for reprints should be addressed, at Food and Drug Administration. 5600 Fishers Lane HFD-lSO, Rockville, MD 20857. insignificantly to DDP accumulation. Genistein, a dietary isoflavone, thus seems to be a promising clinical candidate for combination with DDP.