P-glycoprotein (Pgp) is a highly dynamic ATP-binding cassette (ABC) membrane transporter that effluxes a diversity of molecules out of cells. It is a major determinant of drug absorption, distribution, and excretion in intestines, liver, kidney and brain, and also an impediment to successful chemotherapy in some cancers, HIV and central nervous system (CNS) diseases. About half of marketed drugs are estimated to be transport substrates or inhibitors of Pgp, and the evaluation of Pgp susceptibility of drug candidates has become an important step in the development of new therapeutics in the pharmaceutical industry. However, Pgp binding molecules exhibit complex activities, acting as ATPase stimulators, inhibitors, transport substrates, or non- transportable ligands. Despite extensive studies, we still have very limited understanding of the mechanisms for the complex and polyspecific Pgp-drug interactions pertaining to Pgp transport, inhibition and evasion. This study aims to (1) define how structural and chemical properties of a ligand affect its interactions with Pgp, (2) characterize how Pgp reacts upon the binding of different classes of ligands, and (3) rationalize chemical synthesis to modify existing drugs to evade Pgp transport. We will use an integrative chemistry, structural and functional approach to tackle these aims, which is based on several major advances in the literature and that we have most recently made on Pgp structural determination and ligand interactions. First, we propose to conduct a thorough structure-activity relationship study within a focused library of ligands bearing common scaffolds. We will use a battery of functional assays, including ATPase activity, detailed drug binding and competition, cell-based transport, and drug resistance assays, as well as structural characterizations to evaluate these compounds. Second, we will use several complementary biophysical techniques, including X- ray crystallography, single particle electron microscopy (EM), and luminance resonance energy transfer (LRET), for the determination of Pgp/ligand complex structures, conformational distributions, and the kinetics of conformational changes. The characterization of Pgp conformations by EM and LRET, together with the screen of novel ligands, detergents and lipids, as well as new constructs to stabilize Pgp (and certain conformations), will facilitate higher resolution (< 3.0 ) structural studies, a critical barrier that has eluded Pgp thus far. Third, we will modify several Pgp drug substrates on positions that have been identified without diminution of drug potency, thus with a focus on resulting changes in Pgp interactions. By the related three aim studies we will achieve a detailed and fundamental understanding of polyspecific Pgp-drug interactions, which will have a far- reaching impact on drug discovery given the pharmacological and clinical significance of Pgp and that many current and investigational drugs are susceptible to Pgp efflux.

Public Health Relevance

P-glycoprotein (Pgp) gives rise to multidrug resistance that causes numerous patient deaths in several diseases, most notably in cancers. It also affects the bioavailability and tissue distribution of many drugs in clinical use. Understanding how Pgp interacts with drugs is critical to the treatment of human diseases given its pharmacological and clinical significance and that many drugs lose efficacy because of Pgp-mediated resistance.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM118594-01
Application #
9080833
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Okita, Richard T
Project Start
2016-03-01
Project End
2019-12-31
Budget Start
2016-03-01
Budget End
2016-12-31
Support Year
1
Fiscal Year
2016
Total Cost
Indirect Cost
Name
Scripps Research Institute
Department
Type
DUNS #
781613492
City
La Jolla
State
CA
Country
United States
Zip Code
92037
Waghray, Deepali; Zhang, Qinghai (2018) Inhibit or Evade Multidrug Resistance P-Glycoprotein in Cancer Treatment. J Med Chem 61:5108-5121
Zoghbi, Maria E; Mok, Leo; Swartz, Douglas J et al. (2017) Substrate-induced conformational changes in the nucleotide-binding domains of lipid bilayer-associated P-glycoprotein during ATP hydrolysis. J Biol Chem 292:20412-20424