Abstract Chronic kidney disease is one of the most prevalent diseases around the world, affecting over 35 million people in the United States alone. Of these, roughly 15% of cases are attributed to glomerular diseases such as Alport Syndrome (AS), which progresses to end stage renal disease in 90% of patients by the age of 40. Despite knowing the genetic underpinnings of AS, the downstream mechanisms are not fully understood, making it difficult to create targeted therapies for patients. Work from us and others have implicated the dysregulation of sphingolipid metabolism and signaling in the progression of glomerular disease via injury to the podocyte, a specialized glomerular cell which is crucial for renal filtration. The involvement of sphingosine-1-phosphate (S1P), a highly bioactive sphingolipid, is implicated in glomerular pathology. We have shown that the expression of Apolipoprotein M (ApoM), the physiologic carrier of S1P, is inversely correlated with albuminuria in CKD patients. Similarly, our preliminary studies show an association between ApoM deficiency and podocyte/renal injury in vitro and in vivo. Aim 1 will investigate if ApoM deficiency exacerbates S1P-mediated podocyte injury. Our preliminary data show that Col4a3-/- mice, a model of AS, have significantly decreased APOM in the renal cortex, mirroring the phenotype in human patients. Furthermore, they also show S1P accumulation in kidney cortex in association with glomerular apoptosis, while treatment with recombinant APOM restores these phenotypes and improves renal function. This aim leverages lipidomic analyses, advanced microscopy modalities, and renal phenotype evaluation to determine the role of S1P in mediating podocyte injury. This aim will use various in vitro and in vivo analyses in lentiviral siAPOM cells and renal APOM deficient kAPOMASO-/- mice to establish the role of kidney- derived APOM in modulating S1P content and downstream podocyte injury. Aim 2 will Determine if renal ApoM replenishment is sufficient to rescue from S1P-mediated renal injury associated with AS. Our preliminary data show that treatment of Col4a3-/- mice with rhApoM in vivo prevents the progression of kidney injury. Importantly, Col4a3-/- mice have decreased ApoM expression in the kidney cortex, while treatment with rhApoM restores renal ApoM expression in vivo. We hypothesize that kidney derived ApoM plays a role in preventing S1P accumulation and consequent glomerular injury in AS. This aim seeks to show that restoring the expression of ApoM in Col4a3-/- podocytes will be sufficient to protect from S1P-mediated injury and restore S1P levels in vitro. We also expect that restoration of renal ApoM expression with AAV gene delivery will prevent progression of proteinuria in Col4a3-/- mice.