Generation and characterization of human iPSC-derived <i> NPC1 ^{I1061T/I10161T} </i> i ³ Neurons as a model for NPC1 disease

Niemann-Pick disease, type C is an autosomal recessive, fatal, neurodegenerative disorder caused by pathological variants in <i>NPC1</i> or <i>NPC2</i>. Dysfunction of either NPC1 or NPC2 results in impaired intracellular cholesterol transport and subsequent storage of unesterified cholesterol in endolysosomal compartments. Earlier cell-based studies utilized patient fibroblasts to study this disease; however, neuronal cells allow for investigation of the neurodegenerative aspect of NPC1. Expression of neurogenin in induced pluripotent stem cells leads to the generation of i³Neurons (integrated, isogenic, and inducible), allowing for rapid, synchronized growth of homogenous neurons. In this study, we report the development and characterization of a human iPSC-derived <i>NPC1</i> ^{<i>I1061T/I1061T</i>} i³Neuronal model system. <i>NPC1</i> ^{<i>I1061T</i>} is a missense variant resulting in a misfolded protein targeted for proteasomal degradation in the ER. <i>NPC1</i> ^{<i>I1061T/I1061T</i>} i³Neurons phenocopied the cellular pathological features of NPC1 disease including endolysosomal cholesterol accumulation, lysosomal morphological changes, and response to the proteostasis modulator, mo56HC. The NPC1 phenotype was alleviated by 2-hydroxypropyl-β-cyclodextrin treatment, a drug demonstrating efficacy both <i>in vitro</i> and <i>in vivo</i>. This <i>NPC1</i> ^{<i>I1061T/I1061T</i>} i³Neuronal cell line can facilitate future high-throughput drug and genomic screens, particularly those aimed at identifying proteostasis regulators that improve the expression/stability of the mutant NPC1 protein.