Monitoring of cellular resistance to cancer chemotherapy: Drug retention and efflux

Abstract

Publisher Summary Methods for identification of multiple drug resistance (MDR) include molecular procedures for gene expression (mRNA) in situ polymerase chain reaction (PCR) methods, use of monoclonal antibodies by immunocytochemistry (FCM), and flow cytometry. However, some of these methods cannot determine the heterogeneity of MDR marker expression or determine if the efflux protein is functional and active in reducing cellular drug retention. In several cases, phosphorylation of the P-glycoprotein, cell cycle stage, and proliferation can modulate the functionality of the efflux pump. This can result in discordance between the presence of the efflux proteins and the functional activity of the pump. Similarly, mutations could alter the specificity or activity of the efflux proteins, resulting in altered drug efflux. Analytical methods such as high-pressure liquid chromatography and spectrofluorometry can be used for monitoring of cellular drug retention and efflux, but of necessity these methods are slow, need large samples, and cannot measure drug retention in single cells. The flow cytometric functional assays that basically monitor the retention of a fluorescent drug and compare the total cellular drug content of the resistant versus sensitive cells, or of resistant cells with or without coincubation with an efflux blocker, overcome some of these obstacles.