Inhibition of the non-lysosomal glucosylceramidase GBA2 accounts for the efficacy of sinbaglustat in a murine model of Sandhoff disease
Sinbaglustat is a small molecule under investigation as a treatment for lysosomal storage disorders. It inhibits two enzymes involved in glycosphingolipid (GSL) metabolism, the non-lysosomal glucosylceramidase (GBA2) and, with a lower affinity, glucosylceramide synthase (GCS). High  doses of structurally related dual GBA2/GCS inhibitors such as miglustat, and pure GCS inhibitors have shown efficacy in a transgenic mouse model (Hexb-/-) of Sandhoff disease, aGM2 gangliosidosis. Whether the inhibition of GBA2 alone confers protection is unknown. We first assessed the functional profile of sinbaglustat relating to GBA2 and GCS inhibition in cellular models and wild-type (WT) mice. Then we investigated sinbaglustat's potential efficacy in preventing disease manifestation in Hexb-/- mice upon chronic treatment with two dose levels (30 and 300 mg/kg/day), aimed at predominant inhibition of GBA2 or GBA2 and GCS. Non-treated WT mice served as controls. Sinbaglustat inhibited GBA2 more potently than GCS in GM2 gangliosidosis patient-derived fibroblasts and primary mouse neurons. The dual inhibition profile was also reflected in a dose-dependent, bell-shaped modulation of glucosylceramide, the substrate of GBA2 and product of GCS  , in wild-type mouse brain. At the two sinbaglustat dose levels that we tested in Hexb-/- mice, glucosylceramide levels in cerebral cortex were elevated, indicating GBA2 target engagement. Only the high 300 mg/kg/day dose reduced in addition the levels of various other GSLs compared to Hexb-/- untreated mice, demonstrating GCS target engagement in the central nervous system. In the liver and plasma, sinbaglustat dose-dependently decreased many GSLs including hexosaminidase A substrates. Sinbaglustat partially reversed the increase in mRNA expression of several neuroinflammation-related genes in the hippocampus of Hexb-/- mice. Sinbaglustat treatment at both dose levels delayed the age at which Hexb-/- mice started to develop motor impairments by about 2-4 weeks. The 30 mg/kg/day dose increased median survival of Hexb-/- mice by approximately 13.5%, the 300 mg/kg/day dose by approximately 18%.  The study indicates that sinbaglustat could represent a viable treatment option for GM2 gangliosidosis if data from the mouse model translate to human. Importantly, our data also reveal that GBA2 inhibition in the central nervous system is largely responsible for sinbaglustat's therapeutic effect in Hexb/- mice. Overall design: RNAseq of the hippocampus of Hexb/- mice (hexßtm1R1p) and WT littermates treated with singablustat from week 4 to week 15 at either 30 mg/kg/day, for primary inhibition of GBA2, or 300 mg/kg/day, for inhibition of both GBA2 and GCS. Untreated Hexb /- and untreated WT mice served as comparator and control.