Chemotherapy induced peripheral neuropathy is a common dose-limiting toxicity that can reduce therapeutic effectiveness and impact quality of life for cancer patients. The overarching goal of this research is to determine the molecular basis of chemotherapy-induced peripheral neuropathy to support the development of targeted therapies to prevent and treat this toxicity. The proposed studies are based on a reverse translational pharmacogenetic approach that uses genetic association findings to implicate critical pathways in peripheral neuropathy. Recent genetic association and functional validation findings support a role for sphingosine-1phosphate (S1P) signaling in chemotherapy-induced neurotoxicity, which are consistent with previous studies in rodent models. The studies proposed in this application will extend these findings and address a significant gap in our knowledge of S1P signaling in target cells for toxicity, peripheral sensory neurons. The central hypothesis that will be tested is that modulation of S1P signaling in peripheral sensory neurons by microtubule targeting agents plays a critical role in their neurotoxicity. A human induced pluripotent stem cell derived sensory neuron model of chemotherapy neurotoxicity (iPS-SNs) will be employed for all studies. Pharmacological and genetic approaches will be used to modulate S1P signaling and interrogate chemotherapy toxicity linked to this signaling pathway. The three aims are complementary and address discrete functions of S1P. The first aim will investigate whether microtubule targeting agents alter sphingolipid metabolism in sensory neurons and will link specific S1P receptors to cytoskeletal changes. The studies proposed in the second aim will focus on Rho GTPase signaling downstream of S1P receptors and will establish the S1P signaling axis that is critical for chemotherapy-induced changes in neurite structure and the development of retraction bulbs. The third aim will use scRNA-seq and sc-ATACseq to elucidate whether paclitaxel-induced changes in gene expression in iPS-SNs involve S1P effects on chromatin accessibility. The ability of fingolimod, a multiple sclerosis therapy that targets S1P receptor signaling and is currently being tested for prevention and treatment of paclitaxel-induced peripheral neuropathy, to protect against chemotherapy-induced neurotoxicity will be examined. Collectively, these studies will reveal molecular mechanisms underlying the axon degeneration that occurs in sensory neurons in response to microtubule targeting agents and elucidate novel mechanisms for neuroprotection with fingolimod.