MULTIDRUG-RESISTANCE PROTEIN-MEDIATED TRANSPORT OF CHLORAMBUCIL AND MELPHALAN CONJUGATED TO GLUTATHIONE

The human multidrug resistance protein (MRP1) confers resistance of ce lls to a number of different cytostatic drugs and functions as an expo rt pump for glutathione S-conjugates, glucuronides and other amphiphil ic anions. The present study details for the first time MRP1-mediated ATP-dependent transport of various glutathione S-conjugates of the bif unctional alkylating agents chlorambucil and melphalan. In membrane ve sicles prepared from cells expressing recombinant MRP1, the conjugates were transported at rates in the following order: monoglutathionyl ch lorambucil > bisglutathionyl chlorambucil > monohydroxy monoglutathion yl chlorambucil and monoglutathionyl melphalan > monohydroxy monogluta thionyl melphalan. In addition, we show that membranes from chlorambuc il-resistant GST-alpha-overexpressing CHO cells as well as from their parental cells express the hamster homologue of MRP1. With both CHO ce ll membrane preparations, we observed ATP-dependent transport of monog lutathionyl chlorambucil and of leukotriene C-4, a glutathione S-conju gate and high-affinity substrate of MRP1. The transport rates measured in the resistant cells were only two- to three-fold higher than those measured in the control cells. These results together with cytotoxici ty assays comparing MRP1-overexpressing cell pairs with the CHO cell p air indicate that, although MRP1-mediated transport is active, it may not be the rate-limiting step in chlorambucil resistance in these cell lines.