Investigating p.Ala1035Val in NPC1: New Cellular Models for Niemann–Pick Type C Disease

Niemann-Pick type C (NPC) is a lysosomal storage disorder (LSD) caused by pathogenic variants in either the <i>NPC1</i> or <i>NPC2</i> genes, which encode proteins involved in the lysosomal export of unesterified cholesterol. In patients of Western European descent, the p.Ile1061Thr variant in <i>NPC1</i> is especially prevalent. However, mounting evidence has positioned p.Ala1035Val as the most common variant in Portugal and the second most prevalent variant worldwide. By analyzing 10 Portuguese NPC patients homozygous for p.Ala1035Val, we found an SNP in <i>cis</i> on position 858 (p.Ile858Val), which we hypothesize could have a disease-modifying effect. To address this query, we created variant-specific in vitro models of NPC by stably transducing <i>NPC1</i>^-/- ARPE-19 cells with constructs encoding different fluorescently-tagged variants of NPC1, which we used, alongside patient-derived skin fibroblasts, to investigate lysosomal positioning and the trafficking routes elicited by p.Ile1061Thr and p.Ala1035Val (with and without the p.Ile858Val SNP in <i>cis</i>). Our results corroborate the previously described decrease in p.Ile1061Thr-NPC1 trafficking to the lysosome and suggest a similar, if not worse, scenario for the p.Ala1035Val variant, especially when in <i>cis</i> with p.Ile858Val. This is the first reported functional study addressing the impact of the p.Ala1035Val variant at the cellular level, paving the way for novel therapeutic options.