P-glycoprotein and Vacuolar ATPase Synergistically Confer Anthracycline Resistance to Fission Yeast and Human Cells

Abstract

Drug resistance is a major hurdle to the success of chemotherapy. The permeability glycoprotein (P-gp) is an important factor dictating drug access to the cells, as it controls the efflux of chemotherapeutic agents against the concentration gradient. Pmd1, a P-gp-like protein, was recently isolated as a doxorubicin resistance gene in fission yeast. Although the null mutant of pmd1 (Δpmd1) exhibited sensitivity to doxorubicin, it showed an unexpectedly high resistance to the drug at relatively high concentrations. The data presented here suggest that this is due to the presence of cooperative processes that can complement and counteract drug cytotoxicity in the absence of Pmd1. One such factor, Rav1, is an essential factor in controlling the assembly of the pH-regulating transporter vacuolar-ATPase (V-ATPase) in fission yeast. The simultaneous disruption of Pmd1 and Rav1 resulted in a prominent accumulation of doxorubicin in the cytoplasm of cells, accompanied by a decline in cell viability. With concurrent treatment of pharmacological inhibitors in human cervical cancer cells, P-gp and V-ATPase were further shown to act synergistically to sensitize cells to doxorubicin also in the human cells. Furthermore, a novel Cornichon-like protein SPAC2C4.05 (herein named as Cor1) was demonstrated for the first time to be involved in the interaction with P-gp and V-ATPase to counteract doxorubicin-dependent cytotoxicity. Therefore this study identified a molecular cooperation between multiple membrane transporter proteins that confers chemoresistance to cells against the chemical insult of doxorubicin. Interestingly, this network exhibited differential effects to doxorubicin as compared with its close epimeric analog epirubicin, suggestive of the intricacy of the drug response regulated by this synergistic interaction. A model is discussed on how the versatility of this network can differentiate closely related chemical drug structures yet allow for the robustness to counteract a vast range of drugs.

Calcium modulation of doxorubicin cytotoxicity in yeast and human cells

“…Low levels of calcium and doxorubicin were specifically chosen to show the synergistic effects (Tay et al . ). Instead of a synergistic effect, unexpectedly, we observed a suppression of the doxorubicin‐dependent growth defect upon the incorporation of calcium into the doxorubicin‐containing media.…”

“…; Tay et al . , ) (Figs , S1 in Supporting Information). Two replicates of the calcium sensitivity test were carried out for each strain (Fig.…”

“…The concentrations of doxorubicin used were 15, 30 and 50 μg/mL (Tay et al . ). Tetrandrine (Sigma‐Aldrich) was used at 0 and 1 m m ; verapamil (Sigma‐Aldrich) and nifedipine (Sigma‐Aldrich) at 0, 0.1 and 1 m m , as determined via empirical testing of drug‐dependent cell viability.…”

“…Exponentially growing cultures of the null mutant strains were obtained, and three steps of a 10-fold serial dilution were spotted onto agar media incorporated with decreasing concentrations of CaCl 2 . Wild-type (WT) and Drav1 strains were employed as negative and positive controls, respectively (Dawson et al 2008;Tay et al 2013Tay et al , 2014 (Figs 1, S1 in Supporting Information). Two replicates of the calcium sensitivity test were carried out for each strain ( Fig.…”

Influence of Ribosomal Protein L39-L in the Drug Resistance Mechanisms of Lacrimal Gland Adenoid Cystic Carcinoma Cells

“…Within this field, the studies were concentrated on two aspects: the multiple drug resistance(MDR) (Koshkin et al, 2012) mechanism gene, which encoded 170 P protein, and non-multiple drug resistance gene (Tay et al, 2014). The synthesis and overexpression of multidrug resistance gene is inseparable from the effect of a series of ribosomal protein.…”