Title

Protaglandin EP2 Receptor Antagonism: a Novel Approach to Attenuate/Prevent the Inflammation during Auto-Inflammatory Disorders

Date of Award

2016

Degree Type

Thesis

Degree Name

Master of Science (MS)

First Advisor

Avadhesh C Sharma, PharmD, PhD

Second Advisor

Bonnie Buxton, PhD

Third Advisor

Srujana Rayalam, BVSc, MVSc, PhD

Fourth Advisor

Yue-Qiao Huang, BSc, PhD

Fifth Advisor

Rangaiah Shashidhararnurthy, PhD

Abstract

Arachidonic acid metabolites play a very significant role in initiating inflammatory signaling pathways resulting in target tissue damage. Prostaglandin E2 (PGE2), a bioactive prostanoid family lipid derivative of arachidonic acid (AA) produced by cyclooxygenases (COX) and PG synthases as an essential homeostatic factor under normal physiological conditions. The cellular PGE2 levels are precisely balanced by the synthesis from COX enzymes and degradation by 15-hydroxyprostaglandin dehydrogenase (15-PGDH) enzymes. In addition, PGE2 has been implicated as a key mediator of many immunopathological conditions including chronic inflammatory disorders such as autoimmune arthritis. In the past, several steroid and non-steroid anti-inflammatory drugs have been developed to target the cyclooxygenase to inhibit PGE2 synthesis during various inflammatory disorders to control/prevent PGE2-mediated inflammation. However, recent studies showed the potential inhibition of COX leads to deleterious cardiovascular side effects that resulted in the withdrawal of some drugs. Therefore, it is important to identify a target downstream of COX, rather than blocking the entire COX signaling pathway. Prostaglandin-E2 receptor (subtype EP2) with which PGE2 binds with very high affinity, promotes the inflammation during several pathological conditions. Hence, we hypothesize that targeting the EP2 receptor may offer an innovative alternative strategy to prevent/control inflammation during various autoimmune inflammatory disorders such as Rheumatoid Arthritis (RA). To accomplish this study we used an in vitro cell culture model with mouse macrophage cell line (P388Dl and RAW 264.7) to block functional activity of EP2 receptor with small molecular weight PGE2 antagonists to blunt inflammation during the development of inflammatory disorders. Using EP receptor agonists butaprost or PGE2 for activation of EP2 receptor, we observed that out of seven EP2 antagonists studied, three (TGLI-3) could significantly attenuated the upregulation of many pro-inflammatory cytokines such as TNF-α (4-5 fold), IL-1β (2-3 fold), IL-6 (4 fold), and IL-12 (1-2 fold) at mRNA level. Interestingly, these antagonists also significantly inhibited the up regulation of IL-17 (> 10 fold) and IL-23 (> 10 fold), which are potent cytokines that play a major role during the pathogenesis of autoimmune RA. We also observed around 10-20 fold down regulation of MAPK and ERK1/2 signaling molecules in macrophage cell line treatments with EP2 antagonists. These results suggest that the tested EP2 antagonists inhibit the pro- inflammatory cytokines response by targeting the MAPK pathway signaling molecules. With further confirmatory studies, our data suggests that inhibiting EP2 receptor function using EP2 receptor antagonists can be a novel therapeutic approach to control/prevent the inflammation during chronic inflammatory disorders including rheumatoid arthritis.

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