Self-assembling polymer-based short chain fatty acid prodrugs ameliorate non-alcoholic steatohepatitis and liver fibrosis

Abstract

With the preponderance of a high-calorie diet and sedentary lifestyle, the prevalence of non-alcoholic steatohepatitis (NASH), a state of abnormally elevated lipid accumulation in the liver with chronic inflammation, is increasing at an alarming rate worldwide. Hence, cost-effective therapeutic interventions are required to manage this disease at an early stage. Numerous reports have suggested a link between gut microbial dysbiosis, particularly a decrease in the abundance of short-chain fatty acids (SCFA)-producing microbiota and NASH pathogenesis. Considering these low molecular weight (LMW) SCFAs such as acetic, propionic, and butyric acids have been used to inhibit hepatic steatosis in mouse models. However, the poor pharmacokinetic (PK) profile of SCFAs, caused due to their LMW, renders them therapeutically ineffective. Thus, to improve the PK characteristic-based therapeutic efficacy of LMW SCFAs, we designed SCFA-based prodrugs that possess self-assembling characteristics in aqueous media. The designed SCFA prodrugs consist of enzyme-metabolizable amphiphilic block copolymers, [poly(ethylene glycol)-b-poly(vinyl ester)s] conjugated to propionic acid (PA) or butyric acid (BA) by an ester linkage, which self-assemble into stable nanosized micelles several tens of nanometers in diameter (Nano^PA and Nano^BA). Via pharmacological analysis, we confirmed that, after oral administration, LMW BA decreased to a physiological level within 24 h in the liver, whereas BA liberated from Nano^BA was observed until 72 h post-administration, implying a sustained release profile. Here, we evaluated the therapeutic efficacy of Nano^SCFA in a choline-deficient, L-amino acid-defined high-fat diet (CDAHFD)-induced NASH and liver fibrosis mouse model by ad libitum drinking. Nano^SCFA, particularly Nano^BA, exhibited the remarkable potential to ameliorate the phenotypic features of fatty liver disease by reducing hepatic lipogenesis and fibrosis, with negligible adverse effects. In contrast, conventional LMW SCFAs failed to prevent the pathogenesis of fatty liver disease, which plausibly can be explained by their rapid clearance and discernible adverse effects. Mechanistic studies revealed that Nano^BA restored the nuclear expression of PPARα, a transcriptional factor regulating mitochondrial fatty acid oxidation, in the periportal hepatocytes and decreased the CPT1A expression level in the hepatic tissues, reflecting the therapeutic effects of Nano^BA. Taken together, we confirmed that our Nano^SCFA potentially improved the PK properties of SCFAs, and it consequently alleviated NASH symptoms and fibrotic liver compared to LMW SCFAs. Our study establishes Nano^SCFA as a suitable nano-assembled prodrug for NASH treatment.

Keywords: Butyric acid; Liver fibrosis; NASH; Polymer micelles; SCFA-based self-assembling prodrugs; Short-chain fatty acids.