PubMed · 8763346
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L J Goldstein. 1996. MDR1 gene expression in solid tumours.. https://doi.org/10.1016/0959-8049(96)00100-1
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P-Glycoprotein (P-gp) causes a multidrug resistance (MDR) phenotype in tumour cells. In some cancers, the expression of P-gp has been correlated with low clinical response to chemotherapy and survival of patients. Previous studies have shown that certain lipophilic drugs bind to P-gp and reverse the MDR phenotype of tumour cells. In this study, we extend that list of compounds and present evidence for the capacity of a potent and clinically safe anthelmintic, ivermectin (IVM), as an MDR-reversing drug. Using a highly drug-resistant human cell line, we compared IVM with other MDR-reversing agents and showed that IVM is 4- and 9-fold more potent than cyclosporin A and verapamil, respectively. The capacity of IVM to inhibit iodoaryl-azidoprazosin photolabeling of P-gp is consistent with direct binding to P-gp. Studies showed that [3H]IVM binding to membranes from resistant cells is specific and saturable with KD and Bmax values of 10.6 nM and 19.8 pmol/mg, respectively. However, while cyclosporin A or vinblastine inhibited [3H]IVM binding to membranes from drug-resistant but not drug-sensitive cells, neither verapamil nor colchicine had any effect. Furthermore, both IVM and cyclosporin A and, to a lesser extent, verapamil also inhibited [3H]vinblastine binding to membranes from drug-resistant cells. Drug transport studies showed that [3H]IVM is a substrate for the P-gp drug efflux pump. However, it was transported less efficiently by P-gp than [3H]vinblastine. Moreover, only cyclosporin A was effective in potentiating the accumulation of [3H]IVM in drug-resistant cells. Taken together, the high efficiency of MDR reversal by IVM combined with its low toxicity are consistent with the properties of an ideal MDR-reversing agent.
Cholesterol esterification, catalyzed by acyl-CoA:cholesterol acyltransferase (ACAT), plays a central role in cellular cholesterol homeostasis and in physiologic processes that lead to coronary heart disease. Although ACAT resides in the endoplasmic reticulum (ER), the cholesterol substrate for esterification originates in the plasma membrane and must be transported to the ER for esterification. Progesterone inhibits esterification, possibly by blocking the transport of cholesterol to the ER. Recent studies suggest that progesterone acts by inhibiting the activity of one or more of the multidrug-resistant (MDR) P-glycoproteins. In the current manuscript, we demonstrate that progesterone's ability to inhibit esterification is not mediated through the progesterone receptor. We evaluate a series of steroid hormones and find a strong correlation between a steroid hormone's hydrophobicity and its ability to inhibit both cholesterol esterification and MDR-catalyzed drug efflux. We also find that cholesterol esterification is inhibited by nonsteroidal MDR inhibitors, and that this inhibition specifically affects the esterification of cholesterol derived from the plasma membrane. MDR inhibitors also inhibit cholesterol esterification in a wide range of cultured human cell lines. These observations suggest that MDR activity normally functions in a general process of intracellular cholesterol transport.
There is growing evidence that abnormal protein folding or trafficking (protein kinesis) leads to diseases. We have used P-glycoprotein as a model protein to develop strategies to overcome defects in protein kinesis. Misprocessed mutants of the human P-glycoprotein are retained in the endoplasmic reticulum as core-glycosylated biosynthetic intermediates and rapidly degraded. Synthesis of the mutant proteins in the presence of drug substrates or modulators such as capsaicin, cyclosporin, vinblastine, or verapamil, however, resulted in the appearance of a fully glycosylated and functional protein at the cell surface. These effects were dose-dependent and occurred within a few hours after the addition of substrate. The ability to facilitate processing of the misfolded mutants appeared to be independent of the cell lines used and location of the mutation. P-glycoproteins with mutations in transmembrane segments, extracellular or cytoplasmic loops, the nucleotide-binding domains, or the linker region were processed to the fully mature form in the presence of these substrates. These drug substrates or modulators acted as specific chemical chaperones for P-glycoprotein because they were ineffective on the deltaF508 mutant of cystic fibrosis transmembrane conductance regulator. Therefore, one possible strategy to prevent protein misfolding is to carry out synthesis in the presence of specific substrates or modulators of the protein.