Overcoming multidrug resistance in cancer cells: Biochemical evaluation of new lead drug candidates

Abstract

P‐glycoprotein (P‐gp) overexpression is often correlated with multidrug‐resistance (MDR) of cancer cells due to its role in the transport of chemotherapeutic drugs out of the cells in an ATP‐dependent process. To search for P‐gp inhibitors as co‐therapeutics to combat MDR, we previously used high‐throughput in silico ligand docking studies to identify drug‐like compounds. To assess functionality of the hit compounds we measured their effects on P‐gp ATPase hydrolysis activity. Some of the compounds inhibited ATP hydrolysis of purified P‐gp and successfully reversed the MDR in prostate and ovarian cancer cell lines. We subsequently used computational approaches and structure‐based design to produce variants of one of the hit compounds with predicted higher affinity to P‐gp. The efficacy of these variants in reversing MDR in cell culture as well as the biochemical evaluation of inhibition was assessed. High expression of human P‐gp in the yeast Pichia pastoris in a biologically active form was previously reported and the purified P‐gp was used on our recent assays. Currently, reconstitution of P‐gp into nanodiscs allowed for human P‐gp to be solubilized in its active form with ATPase activity enhanced by 2–3 fold as well as increased stability. Here we report the evaluation of the potential of our previously discovered drug‐like compounds to serve as inhibitors of human P‐gp ATP hydrolysis activity.Support or Funding InformationThis work was supported by NIH NIGMS [R15GM094771‐02] to PDV and JGW, SMU University Research Council, SMU Hamilton Undergraduate Research Scholars and Undergraduate Research Assistantship Programs, the SMU Center for Drug Discovery, Design and Delivery, the Communities Foundation of Texas, and private gifts from Ms. Suzy Ruff of Dallas, Texas, and Ms. Myra Williams, Ph.D. of Naples Florida. The authors would like to thank Dr. Ina L. Urbatsch, Texas Tech University Health Science Center, for providing the P. pastoris cells expressing the human wild‐type MDR1 used in this work.