Novel roles for RIP kinases in islet inflammation and beta-cell cytotoxicity in type 2 diabetes

Rlr Va Medical Center, Indianapolis IN

Investigators

Abstract

Background and Innovation: β-cell failure is a hallmark of type 2 diabetes (T2D) that arises with islet amyloid formation, islet inflammation, β-cell loss and hyperglycemia. However, the cellular and molecular mechanisms that underlie this pathogenic process are not well understood. Receptor interacting protein kinase 3 (RIPK3) is a pro-death signaling molecule that has been implicated in amyloid-associated brain pathology as well as β- cell cytotoxicity. Recent work from the Templin lab revealed that loss of RIPK3 protects from T2D stress- induced islet inflammation and β-cell cytotoxicity in vitro as well as β-cell loss and glucose intolerance in vivo. In the current project, the role of RIPK3 in T2D-associated β-cell dysfunction and cytotoxicity will be evaluated in greater detail using a humanized mouse model of T2D with β-cell specific (Ripk3βKO) or macrophage specific (Ripk3MϕKO) loss of RIPK3 expression. The translational potential of targeting RIPK3 will also be evaluated using a small molecule RIPK3 inhibitor identified as a lead compound. The proposed in vitro studies (Aim 1) will evaluate actions of RIPK3 in inflammatory gene expression via RNA sequencing, examine its role in kinase signal transduction using a novel chemical-proteomics approach called kinome profiling and characterize its role in islet cell death using a high-content live cell imaging and analysis system. The role of RIPK3 in β-cell insulin secretory function will also be examined. The proposed in vivo studies (Aim 2) will decipher the role of RIPK3 in T2D disease pathogenesis using β-cell specific (Ripk3βKO) or macrophage specific (Ripk3MϕKO) loss of RIPK3 in a T2D-relevant humanized mouse model of islet amyloid formation and β-cell cytotoxicity. These studies will determine how loss of RIPK3 alters body composition, glucose homeostasis, islet function and islet morphology following high fat diet feeding and islet amyloid formation in vivo. These studies will expand the current understanding of T2D pathogenesis by evaluating a novel mediator of islet inflammation and β-cell fate and may establish RIPK3 as a novel therapeutic target with potential to treat or prevent T2D. Significance and Impact to Veterans Healthcare: T2D disproportionally affects the veteran population, with recent studies reporting that 25% of VA patients have diabetes compared to 10% of the general population. Since T2D is a major contributor to morbidities including heart disease, kidney disease and stroke, new therapies to treat or prevent T2D will provide significant health benefits to Veterans. The research proposed here aims to develop novel oral therapeutics that improve β-cell function and/or survival and can be used to prevent or treat T2D and improve Veteran health. This research will establish the role of RIPK3 in T2D pathogenesis using a humanized mouse model of diabetes and evaluate a small molecule RIPK3 inhibitor identified as a translationally-relevant lead compound. Path to translation/implementation: Previous studies identified RIPK3 as a therapeutic target with relevance to T2D disease pathogenesis. The studies proposed here will extend those findings by deciphering the specific mechanisms by which RIPK3 mediates impaired glucose homeostasis in T2D. They will also assess the effects of a lead compound small molecule RIPK3 inhibitor on human islets. Successful completion of this project will result in additional translational studies to understand pharmacokinetic and pharmacodynamic properties of candidate RIPK3 inhibitors and to evaluate their effects on β-cell health and glucose homeostasis in animal models of T2D in vivo. The long-term goal of these studies is to develop small molecule RIPK3 inhibitors that can be used clinically to reduce islet inflammation and β-cell cytotoxicity in Veterans with prediabetes or T2D.