Poly-ion complex micelle effectively delivers CoA- conjugated CPT1A inhibitors to modulate lipid metabolism in brain cells

Abstract

Carnitine palmitoyltransferase 1 (CPT1) is a central player in lipid metabolism, catalyzing the first committed step to fatty acid oxidation (FAO), and is an appealing target for several diseases. However, CoA-conjugated CPT1 inhibitors are negatively-charged and have low cell membrane permeability. Herein, we report the use of a poly-ion complex (PIC) micelles to deliver the specific CPT1 inhibitors, (±)-, (+)-, and (-)-C75-CoA into U87MG glioma cells and GT1-7 neurons. PIC micelles were formed through charge-neutralization of the cargo with the cationic side chain of PEG-poly{N-[N'-(2-aminoethyl)-2-aminoethyl]aspartamide} (PEG-PAsp(DET)), forming 50-60-nm particles with a neutral surface charge. Upon short-term incubation with cells, the micelle-encapsulated CPT1 inhibitors resulted in 1.5 to 5-fold reduction of ATP synthesis, compared to the free drug, without an apparent decline in cell viability. Micelle treatment showed a discernible decrease in oxidation of 14C-palmitate into CO2 and acid-soluble FAO metabolites, confirming that the substantial lowering of ATP production was related to FAO inhibition. Micelle treatment also diminished IC50 by 2 to 4-fold over the free-drug-treated U87MG after long-term incubation. The resulting IC50 is comparable with etomoxir. We synthesized a fluorescent CoA derivative and encapsulated it in the PIC micelle to measure cellular uptake. The fluorescent micelle showed more efficient internalization in both cell types, especially in neurons where uptake reached up to 3-fold over the free dye. The results starkly demonstrate that the PIC micelle is a promising delivery system for anionic inhibitors of CPT1 in glioma cells and neurons, laying the groundwork for future research or clinical applications.