A Precision Medicine for Alcohol-Associated Liver Disease

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ABSTRACT The goal of this study is to develop an RNAi-based genetic precision medicine to treat alcohol associated liver disease (AALD). AALD is a devastating health problem worldwide, accounts for the majority of alcohol-related mortality globally and is the second most indication for liver transplantation in the US. The current medical management for AALD remains limited, and no proven pathobiology-driven pharmacotherapy is available. There is thus an urgent need to identify novel targets and develop promising therapies for both prevention and treatment for AALD. Recent genome-wide association studies (GWAS) has identified that a single nucleotide polymorphism (SNP rs738409 C>G, or amino acid alteration at position 148 I>M) in the patatin-like phospholipase domain containing 3 (PNPLA3) gene is the single largest genetic risk factor contributing to the progression of AALD to advanced liver perturbations including liver cancer. The PNPLA3 148 I>M polymorphism is the molecular reason why this subset of AALD patients develop severe liver injuries. Mechanistic studies have revealed that overexpression of the mutant PNPLA3 (the 148M isoform) is underlying both liver steatosis and fibrosis and inflammation by interrupting hepatic triglyceride metabolism. Our creation of a humanized PNPLA3148M mouse model successfully recapitulates the AALD phenotypes and has corroborated the aforementioned mechanism. We hypothesize that transcriptional downregulation of the PNPLA3148M isoform is a cause-targeting strategy for the treatment of AALD with the PNPLA3148M allele. To this end, we pioneered the development and have issued patents for a precision allele-specific small interfering RNA (siRNA) that possesses excellent specificity for the PNPLA3148M isoform compared to its wild-type counterpart. We further discovered two more modified oligonucleotides which significantly improved the medicinal chemistry specifications of our therapeutic lead. Administration of our lead siRNA to the liver of the humanized PNPLA3148M model fed an ethanol-containing diet significantly reduced the PNPLA3148M expression as well as hepatic steatosis, inflammation and fibrosis. We also demonstrated that delivering our siRNA into the humanized PNPLA3148M model with the N-Acetylgalactosamine (GalNAc)-conjugation, a well-established liver-specific drug delivery strategy, effectively knocked down hepatic hPNPLA3148M mRNA and ameliorated the liver histology. With this success, we propose to advance our drug candidate development by focusing on an Investigational New Drug (IND) application to the FDA. Specifically, we aim: 1) to define the best drug candidate of our hPNPLA3148M-targeting, GalNAc-conjugated oligonucleotides (Phase I); 2) to collect IND-enabling quality assurance and safety data in rodents and non-human primates (Phase II). We expect that via this study, we will develop the first genetic medicine for AALD treatment, which could also serve as a new prototype for drug development for other common human diseases. The collected data in this study will enable us to file the IND application to support our first clinical trial in humans.