Development and preclinical evaluation of an asialoglycoprotein receptor 1-targeted antibody-drug conjugate in hepatic inflammation.

Hepatic inflammation leads to progressive conditions such as metabolic dysfunction-associated steatohepatitis, cirrhosis, liver cancer, or failure. Due to the high prevalence rates of hepatitis-induced serious liver pathologies, effective interventions are needed. In this study, we developed an antibody-drug conjugate (ADC) from an in-house generated and well characterized monoclonal antibody targeting asialoglycoprotein receptor 1 (ASGPR1) in hepatocytes. The mAb showed strong affinity for ASGPR1 protein with a KD value of 4.2 ± 0.55 nM and high specificity for ASGPR1 in primary hepatocytes and mice liver tissues. After confirming the biodistribution of mAb ASGPR1 in the liver, we employed AAV8-TBG vector enabled green fluorescent protein (GFP)-labeled hepatocytes to validate the specific localization of mAb ASGPR1 to liver hepatocytes. Immunofluorescence analysis showed >80% hepatocytes expressing the GFP reporter and low uptake in other cells. Further, to establish mAb ASGPR1-mediated drug delivery, an anti-inflammatory drug dexamethasone (DEX) was conjugated to mAb ASGPR1 through an ester linker generating mAb ASGPR1-DEX ADC. mAb ASGPR1-DEX conjugate was stable and did not undergo nonspecific cleavage in plasma over 6 days at 37 °C. Besides, mAb ASGPR1-DEX ADC retained binding specificity and affinity toward ASGPR1 comparable to those of the unconjugated mAb ASGPR1 antibody. The mAb ASGPR1-DEX conjugate exhibited significant efficacy against lipopolysaccharide-induced hepatic inflammation by attenuating levels of interleukin-6, interleukin-1beta, and tumor necrosis factor in mouse precision cut liver slices and hepatocyte organoids. Collectively, these findings hold significant implications for mAb ASGPR1-based ADC as a promising therapeutic modality that warrants further investigation in inflammation-induced liver pathologies.