Date of Award

1-5-2023

Document Type

Doctoral Dissertation - Restricted (NYMC/Touro only) Access

Degree Name

Doctor of Philosophy

Department

Pharmacology

First Advisor

Victor G. Garcia, Ph.D

Abstract

The orphan G protein coupled receptor (GPCR), GPR75, is a Gq coupled GPCR and represents one of many potential targets for the development of therapeutic interventions to remedy disease. The ligand of GPR75, 20-hydroxyeicosatetraenoic acid (20-HETE), is the ω-hydroxylation product of arachidonic acid which is catalyzed by CYP4A/4F enzymes and is a vasoactive lipid with implications in a wide array of cardiometabolic diseases. 20-HETE has been shown to modulate blood pressure, to be elevated in the blood and urine of individuals with obesity where it is proposed to act as a pro-adipogenic agent, and it has even been found to impair insulin receptor signaling. All told, 20-HETE has implications ranging from hypertension to chronic kidney disease, ischemic stroke, and myocardial infarct driven heart failure. With so many implications in conditions detrimental to human health, 20-HETE has attracted a lot of attention not only historically but in the present day, where its roles in diabetes and implications in worsening COVID-19 outcomes have come to light. In Gpr75-transfected HTLA cells, 20-HETE stimulates intracellular Ca2+ levels, IP-1 accumulation and β-arrestin recruitment; all of which were negated by known 20-HETE functional antagonists. Mutation of Thr212 within the putative 20-HETE binding site abolished 20-HETE’s ability to stimulate GPR75 activation. Knockdown of Gpr75 in human endothelial cells nullified 20-HETE-stimulated intracellular Ca2+. C-C Motif Chemokine Ligand 5 (CCL5), a suggested GPR75 ligand, failed to activate GPR75; however, it inhibited 20-HETE’s ability to activate GPR75 signaling. With these results, we provide strong evidence for the deorphanization of GPR75 as the 20-HETE receptor, and lay the framework for the development and assessment of more potent GPR75 agonists, partial agonists, and antagonists that may be leveraged against 20-HETE:GPR75 driven illness.

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