Exhibition at the American Society of Cell & Gene Therapy 2025 Meeting

Bile acid homeostasis is crucial for liver function, and the Sodium Taurocholate Co-Transporting Polypeptide (NTCP, SLC10A1 gene) is integral in the hepatic re-uptake of bile acids. Dysregulation of this process, leading to toxic bile acid build up, is implicated in liver cholestatic diseases such as Primary Sclerosing Cholangitis (PSC) and Biliary Atresia (BA). These conditions currently lack effective therapies, leaving a significant unmet medical need. Innovative therapeutic strategies, grounded in human genetics, could selectively lower NTCP's function in bile acid re-uptake without affecting its other cellular roles. Sufficient NTCP modulation leading to a 2-fold bile acid increase in plasma, is expected to have a therapeutic effect. Axiomer, an RNA editing platform, uses chemically modified editing oligonucleotides (EONs) complementary to the SLC10A1 mRNA to recruit and guide endogenous Adenosine Deaminase Acting on RNA (ADAR) to perform targeted A-to-I editing on the double stranded structure. This results in a single amino acid change in the protein sequence (NTCP Q68R), enabling alteration of the bile acid re-uptake function in NTCP. Here we present preclinical proof of concept data of the approach and the efficiency of Axiomer EONs in in vitro and in vivo studies. 
In-silico 3D structural modelling of the NTCP Q68R variant demonstrated disruption of hydrogen bonds and contacts for Na+ binding which are essential for bile acid transport. In in vitro assessment, a plasmid expressed NTCP Q68R variant demonstrated a 7-fold reduction in bile acid uptake vs. wild-type with no impact on protein expression nor localization. EONs targeting SLC10A1 RNA demonstrated a 20% RNA editing efficiency in vitro, and a dose-dependent bile acids uptake inhibition (40% decrease with the 50nM dose vs. control to a near complete inhibition with the 100nM dose vs. control). 
In follow-up in vivo studies, ADAR mediated editing of NTCP resulted in a mean editing efficiency of 13.3% in humanized liver mice and 18.3% in NHP respectively (reaching up to 24% in NHPs). These EONs led to a mean change of 4.4-fold in plasma bile acids, above the 2-fold change therapeutic threshold. Correlation between plasma bile acids and EONs editing efficiency (linear regression R2 = 0.51) was also reported. The observed change in plasma bile acids levels was associated with a change in bile acids profile, from an unconjugated (73%) / conjugated (27%) ratio to an unconjugated (13%) / conjugated (87%) ratio post treatment in NHPs. In safety analysis, EONs confirmed class safety, with no hepatotoxicity or immunostimulatory score. 
ADAR-mediated editing of NTCP is a promising approach to address unmet medical needs in cholestatic diseases, such as PSC and BA. By decreasing bile acid re-uptake in the liver and increasing plasma bile acids leading to compensatory mechanism limiting bile acid production, the technology can potentially alleviate the bile acid toxicity in the liver. A clinical candidate, AX-0810, using GalNAc-mediated delivery and with enhanced potency (reaching up to 57% editing, representing a 5.5-fold increase over early generation) and stability profile, is being investigated in longer term in vivo studies and will be further investigated in clinical trials starting this year.

Regulatory RNAs (regRNAs) are long, short-lived, noncoding RNAs expressed from enhancers and promoters that act locally to regulate gene expression. This regulation is accomplished through the ability of regRNAs to bind transcription factors and increase their local concentration near a gene’s promoter and enhancer elements. Our RAP Platform enables the identification of regRNAs and antisense oligonucleotides (ASOs) targeting these regRNAs to modulate target gene expression. Using this platform, we successfully identified CMP-CPS-001, a clinical candidate targeting a regRNA of the gene encoding carbamoyl-phosphate synthase 1 (CPS1), the rate-limiting enzyme in the urea cycle. 
The urea cycle is responsible for the elimination of ammonia from the body by its conversion to urea and subsequent excretion in urine. Mutations in enzymes of the urea cycle cause urea cycle disorders (UCDs), a group of rare disorders characterized by the toxic accumulation of ammonia that results in neurological symptoms and even death. At present, UCDs are inadequately treated, and a significant unmet need remains. 
CPS1 is the first enzyme in the urea cycle that directly acts upon ammonia. We posited that increasing CPS1 activity could reduce ammonia levels by increasing metabolic flux through the urea cycle. We identified ASOs that target a human CPS1 regRNA, as well as surrogate ASOs that target a mouse Cps1 regRNA, and increase the expression of these genes. Increasing Cps1 expression using a mouse surrogate ASO reduced plasma ammonia levels and increased urea production following an acute ammonia challenge in a disease-relevant mouse model carrying a mutation in ornithine transcarbamylase gene (Otc), a urea cycle gene mutated in ~60% of patients with UCDs. The clinical candidate – CMP-CPS-001 – increased CPS1 gene expression in vitro in both wild-type and patient-derived primary hepatocytes, and reduced plasma ammonia levels in a humanized-liver mouse model. Finally, administration of CMP-CPS-001 resulted in increased ureagenesis in wild-type cynomolgus monkeys. 
These preclinical data demonstrate that targeting a CPS1 regRNA and increasing CPS1 expression may directly ameliorate hyperammonemia, which is the key driver of symptomology in UCD patients. Moreover, the mouse model data show that increasing Cps1 expression can lower ammonia and increase ureagenesis in the context of a mutation in Otc. CMP-CPS-001 is currently being evaluated in a Phase I clinical trial in adult healthy volunteers and is in development for the potential treatment of multiple urea cycle disorders.