Influence of microenvironmental pH on adriamycin resistance

Resistance to Adriamycin (ADR) is frequently dependent upon enhanced efflux associated with the expression of the MDR1-encoded P membrane glycoprotein. Since enhanced expression of the MDR1 gene in ADR-resistant cells may be the result of spontaneous genetic mutation or amplification, it is presumed to be relatively stable and unalterable. Yet, reducing ADR efflux could increase sensitivity, and has been attempted using calcium channel blockers and other drugs. However, since the tumor cell microenvironment varies with respect to pH because of differences in vascularization, oxygenation, and metabolite clearance, the possibility exists that these factors could influence drug transport and the critical biochemical pathways which determine cytotoxicity, even in resistant cells. Using flow cytometric analysis of ADR fluorescence, the influx and efflux of 10 microM ADR dissolved in MES buffer (pH 6.5) and 4-(2-hydroxyethylene)-1-piperazineethanesulfonic acid buffer (pH 7.5 and 8.5) was measured in sensitive P388 and resistant P388/R84 cells in vitro. Substantially enhanced uptake of ADR was detected at alkaline pH in both cell populations, while the proportion of ADR-positive cells and the level of ADR uptake was decreased at lower pH. Acidification reduced ADR efflux, whereas alkalinization increased efflux when the uptake pH was 6.5 or 7.5. At uptake pH 8.5, the pH of the external buffer had little effect, even in resistant cells. In resistant cells in an alkaline microenvironment, ADR transport and retention were superior to that observed in sensitive cells in an acidic microenvironment. No differences were observed in ADR transport when the transmembrane pH gradient was equilibrated. These observations are especially relevant to the effect of ADR on tumor cell subpopulations that are acidic, and in which drug diffusion is inefficient. Efforts to alkalinize tumor cells prior to ADR therapy might reduce ADR resistance, even of genetic origin.