PROJECT SUMMARY SPLIS is an ultra-rare, often lethal inborn error of metabolism recognized just five years ago. Most affected children exhibit a rapidly progressive form of nephrotic syndrome which leads to failure-to-thrive and kidney failure, the main cause of death. Affected children may exhibit adrenal insufficiency, hypothyroidism, skin and neurological defects, and immunodeficiency. Although kidney transplantation can be lifesaving, there is no cure for SPLIS. SPLIS is caused by recessive mutations in SGPL1, which encodes sphingosine-1-phosphate (S1P) lyase, a vitamin B6-dependent enzyme responsible for catabolism of the bioactive lipid S1P in the final step of sphingolipid metabolism. SPLIS-associated SGPL1 mutations impair S1P lyase activity, enzyme localization and/or stability, and cause sphingolipid accumulation leading to multi-organ dysfunction and failure. Unlike classical sphingolipid metabolic disorders involving lysosomal enzymes (such as Tay-Sachs disease), SPLIS is an atypical (non-lysosomal) sphingolipid disorder. The study of SPLIS biology is revealing shared features of atypical sphingolipid disorders and the potential for common diagnostic, monitoring and treatment strategies. As pioneers in the field —having identified the first S1P lyase gene from budding yeast and studied its function for decades—we are now directing our expertise to develop targeted therapies for SPLIS. To our knowledge, we are the only group working toward that goal. Some children with less severe forms of SPLIS may respond to supplementation with vitamin B6, the enzyme’s cofactor. Studies in a mouse model of SPLIS have provided proof-of-concept for the use of adeno-associated virus-mediated SGPL1 gene therapy as potentially curative treatment for SPLIS. Clinical trials testing these two therapeutic strategies are in the planning stages. Further, we have developed disease biomarkers and are preparing to launch the first prospective SPLIS natural history study. However, there are major challenges to existing interventions. Pyridoxine is only useful for a limited number of patients harboring B6-responsive variants. Kidney transplantation requires adequate weight gain, is costly, and has high morbidity. Gene therapy is irreversible, can have severe toxicities, and cannot be used prenatally. We hypothesize that enzyme replacement therapy (ERT) represents a lifesaving treatment for SPLIS that acts by preserving kidney function and offers many advantages over other treatment strategies. In this project we will use two powerful and complementary mouse models of SPLIS, a well-established knock-out mouse and a novel knock-in mouse, to test our hypothesis with two specific aims: 1) Establish proof-of-concept for ERT as a lifesaving treatment for SPLIS; 2) Establish proof-of-concept for the ability of ERT to prevent or reverse SPLIS nephrosis. Our results will provide the rationale for further developing ERT as a treatment for SPLIS. Our team, comprised of an exp...