An Analysis of Biomarkers for the Evaluation of Gene Therapy in Niemann–Pick Disease Type C1 Mice

Abstract

Niemann–Pick disease type C1 (NPC1) is an autosomal recessive lysosomal storage disorder caused by pathogenic variants of the NPC1 gene that encodes a protein essential for lysosomal cholesterol transport. A deficiency in NPC1 results in the accumulation of unesterified cholesterol and sphingolipids, leading to neurological, psychiatric, and hepatic manifestations from infancy to adulthood. The currently approved treatment is palliative. Although the efficacy of gene therapy has been demonstrated in murine models, reliable biomarkers for evaluating the treatment effects remain unknown. We evaluated adeno-associated virus (AAV) vector-mediated NPC1 gene therapy in Npc1 homo-knockout (Npc1^−/−) mice, focusing on blood-based biomarkers. An AAV vector carrying human NPC1 under a cytomegalovirus promoter (AAV-hNPC1) was administered intraperitoneally on days 6–8 after birth at varying vector doses and analyzed at multiple time points: 1.8 × 10¹¹ vector genomes/mouse analyzed at 7 weeks (Low/7w) and 1.0 × 10¹² vector genomes/mouse at 4 weeks (High/4w) and 9 weeks (High/9w). hNPC1 is expressed in the brain and liver, and a degree of neuronal cell survival is observed. High-dose AAV treatment improves body weight and rotarod performance. Plasma N-palmitoyl-O-phosphocholine-serine (PPCS) and lysosphingomyelin (lyso-SM) levels were significantly elevated in Npc1^−/− mice. PPCS increased with disease progression but was significantly decreased after later points of high-dose AAV treatment (saline-treated Npc1^−/− mice: 12.88 ± 3.53 ng/mL, AAV-treated Npc1^−/− mice: 7.87 ± 1.67 ng/mL, p = 0.0008). Lyso-SM and oxysterols showed limited changes after therapy. Vector genome analysis revealed higher and more sustained levels in the brain than in the liver, which is consistent with rapid hepatocyte proliferation-reducing vector persistence. These findings demonstrate that systemic AAV-hNPC1 therapy ameliorates motor and neurological deficits but has a limited impact on several cholesterol-related biomarkers. PPCS has been suggested as a sensitive biomarker of therapeutic response and warrants further evaluations in preclinical and clinical NPC1 gene therapy trials.