Abstract
2-Hydroxy-propyl-β-cyclodextrin (HPβCD), a cholesterol scavenger, is currently undergoing Phase 2b/3 clinical trial for treatment of Niemann Pick Type C-1 (NPC1), a fatal neurodegenerative disorder that stems from abnormal cholesterol accumulation in the endo/lysosomes. Unfortunately, the extremely high doses of HPβCD required to prevent progressive neurodegeneration exacerbates ototoxicity, pulmonary toxicity and autophagy-based cellular defects. We present unexpected evidence that a poly (ethylene glycol) (PEG)-lipid conjugate enables cholesterol clearance from endo/lysosomes of Npc1 mutant (Npc1−/−) cells. Herein, we show that distearyl-phosphatidylethanolamine-PEG (DSPE-PEG), which forms 12-nm micelles above the critical micelle concentration, accumulates heavily inside cholesterol-rich late endosomes in Npc1−/− cells. This potentially results in cholesterol solubilization and leakage from lysosomes. High-throughput screening revealed that DSPE-PEG, in combination with HPβCD, acts synergistically to efflux cholesterol without significantly aggravating autophagy defects. These well-known excipients can be used as admixtures to treat NPC1 disorder. Increasing PEG chain lengths from 350 Da-30 kDa in DSPE-PEG micelles, or increasing DSPE-PEG content in an array of liposomes packaged with HPβCD, improved cholesterol egress, while Pluronic block copolymers capable of micelle formation showed slight effects at high concentrations. We postulate that PEG-lipid based nanocarriers can serve as bioactive drug delivery systems for effective treatment of lysosomal storage disorders.
Highlights
2-Hydroxy-propyl-β-cyclodextrin (HPβCD), a cholesterol scavenger, is currently undergoing Phase 2b/3 clinical trial for treatment of Niemann Pick Type C-1 (NPC1), a fatal neurodegenerative disorder that stems from abnormal cholesterol accumulation in the endo/lysosomes
There are no current treatments available for children suffering from this devastating rare genetic disease. 95% of NPC cases result from defects in the NPC1 protein, a large glycoprotein with 13-membrane-spanning domains that resides on the surface of late endosomes/lysosomes, with a sterol-sensing domain in the lumen that binds to cholesterol and enables egress to a cytoplasmic location[3]
We evaluated autophagic activity using Npc1−/− and Npc1+/+ mouse embryonic fibroblasts (MEFs) that stably express a doxycycline-inducible firefly luciferase-p62 construct (p62-fluc). p62 acts as a marker for cellular cargo that is degraded by autophagy
Summary
2-Hydroxy-propyl-β-cyclodextrin (HPβCD), a cholesterol scavenger, is currently undergoing Phase 2b/3 clinical trial for treatment of Niemann Pick Type C-1 (NPC1), a fatal neurodegenerative disorder that stems from abnormal cholesterol accumulation in the endo/lysosomes. An intrathecal cyclodextrin injection in a single NPC1 patient led to an increase in cholesterol redistribution in the central nervous system and improved vertical gaze palsy, a clinical indicator of NPC-linked neurodegeneration[13] Based on these results, HPβCD has entered NIH initiated Phase 2b/3 clinical trial, and the FDA has granted orphan drug status to further accelerate use of HPβCD for treating NPC disease. We found that DSPE-PEG used in combination with HPβCD showed a substantial decline in the lysosomal cholesterol content in Npc1-deficient cells These studies strongly suggest that a simple mixture of two well-characterized biocompatible excipients can have a significant impact in the treatment of NPC lysosomal storage disorder
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