Validating absolute lymphocyte count and plasma sphingosine-1-phosphate as disease biomarkers of sphingosine phosphate lyase insufficiency syndrome in anticipation of a pyridoxine clinical trial - Sphingosine phosphate lyase insufficiency syndrome (SPLIS) is an inherited metabolic childhood syndrome
caused by recessive mutations in SGPL1. Affected children exhibit steroid-resistant nephrotic syndrome with
rapid progression to end stage renal disease, adrenal insufficiency, neurological defects and lymphopenia. A
range of severity has been observed, with the most severely affected patients dying in utero, while others live
to adulthood, albeit requiring dialysis or kidney transplantation. Importantly, there is no specific cure for SPLIS,
and thus there is a dire need for the development of novel safe and effective treatments. SGPL1 encodes
sphingosine phosphate lyase (SPL), an essential intracellular enzyme responsible for the irreversible
breakdown of sphingosine-1-phosphate (S1P) in the final step of sphingolipid metabolism. S1P is a bioactive
lipid that signals through its five G protein coupled S1P receptors to regulate lymphocyte trafficking and other
physiological processes. SGPL1 mutations impair SPL activity and cause sphingolipid accumulation leading to
organ dysfunction and failure. In addition, aberrant S1P signaling in SPLIS children leads to lymphopenia due
to a defect in lymphocyte egress. SPL activity depends on vitamin B6 as a cofactor. Independent of its role as
a coenzyme in over 160 enzymatic reactions, vitamin B6 can also function as a chaperone, stabilizing vitamin
B6-dependent enzymes. Pharmacological doses of vitamin B6 in the form of the B6 vitamer pyridoxine have
proven effective in inborn errors of metabolism involving B6-dependent enzymes. Based on encouraging
preliminary findings in two SPLIS patients treated with vitamin B6 supplementation, we are preparing to
evaluate the safety and efficacy of vitamin B6 supplementation as the first specific treatment for SPLIS.
Challenges to developing a reliable assay for monitoring SPL activity in blood samples from SPLIS patients
make reliance on disease biomarkers a critical component of any clinical trial in SPLIS. We hypothesize that
absolute lymphocyte count (ALC) and plasma S1P levels will serve as reliable biomarkers that reflect SPLIS
disease status. Unfortunately, pediatric reference ranges for plasma S1P and other major sphingolipids have
not been reported to date. Further, no study comprehensively profiling circulating immune cell populations in
SPLIS has been reported. To confirm our hypothesis and overcome these obstacles to advancing treatments
for children with SPLIS and other sphingolipid metabolic disorders, we will characterize the major plasma
sphingolipids and blood markers of immune function in children with SPLIS and a healthy pediatric and young
adult control cohort. In accomplishing our Specific Aims, we will validate plasma S1P and ALC as robust SPLIS
disease biomarkers. We will establish reference ranges for a comprehensive set of plasma sphingolipids in a
healthy pediatric and young adult cohort that will be useful in the future for diagnosis, monitoring and
conducting clinical trials in a broad range of sphingolipid disorders. Our results will specifically facilitate clinical
trial readiness for testing the safety and efficacy of vitamin B6 cofactor supplementation to treat SPLIS.
