Podocyte-specific Rap1 agonism for treatment of glomerular disease

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Project Summary/Abstract: The incidence of proteinuric kidney diseases are increasing with well over 500 million cases worldwide. Over the last two decades, it has become clear that specialized kidney cells, called podocytes, regulate kidney filtration and are injured in all forms of proteinuric diseases regardless of etiology. Despite this, targeted effective therapies that protect podocytes and slow chronic kidney disease progression are completely lacking. Previously, we reported that the small GTPase Rap1 regulates fundamental biological processes in podocytes by cycling between inactive GDP-bound and active GTP-bound forms. We demonstrate that the podocyte Rap1 activation state is controlled by many upstream factors, both positive and negative, that converge to control the ratio of GTP- and GDP-bound forms. The essential role of these upstream regulators is emphasized by the presence of human familial nephrotic syndrome caused by mutations to several of these genes, including newly identified gain of function mutations in the negative Rap1 regulator, Rap1GAP. Each of these mutations has in common that they cause relative depletion of podocyte Rap1-GTP. In the current proposal, we show that levels of podocyte Rap1-GTP are also diminished in human glomerular diseases including in diabetic kidney disease (DKD). Augmenting levels of podocyte Rap1 activation genetically or pharmacologically protects podocytes in short term injury models. By targeting podocyte-specific upstream Rap1 regulatory pathways, we are developing novel Rap1 agonist compounds that activate Rap1 in podocytes, but not in other cell types. Such an approach allows for podocyte-specific pharmacological Rap1 activation and avoids potential systemic toxicities. The goals of the current grant are to explore the therapeutic potential of Rap1 agonists in podocyte diseases including in DKD. We accomplish this via three specific aims: i) Test whether enhanced podocyte Rap1 activation will mitigate chronic glomerular injury utilizing novel inducible podocyte-specific constitutively active Rap1 transgenic mice. ii) Characterize the glomerular phenotype of a novel Rap1GAP knock-in mouse model that expresses a double missense Rap1GAP human disease-associated variant and then utilize both novel mouse models to elucidate specific Rap1 downstream effectors that establish podocyte injury susceptibility versus protection. iii) Synthesize podocyte-targeted Rap1 agonist compounds and screen their ability to rescue podocyte injury in transgenic Rap1-GTP deficient zebrafish. The efficacy of lead compounds will be tested in mouse models of kidney disease including in DKD. Our current lead compound, BT-529, a Rap1-Rap1GAP interaction antagonist, induces Rap1 activation in podocytes but not in other kidney cells and dramatically protects cultured podocytes from injury. Overall, this work sets the stage for urgently needed new podocyte-targeted therapies for kidney diseases.