Pharmacokinetics, Biodistribution, Immunogenicity, and Model-Informed-Based PK/PD Model of a Next-Generation Advanced Novel Gene Therapy for Hemophilia

Hemophilia B is an X-linked hereditary coagulation condition resulting from a defect in the factor IX gene. Gene-based delivery offers a promising alternative to protein-based medicines. The efficacy and safety may be influenced by several parameters, including the dosage of gene therapy, biological distribution, transduction efficiency, immunogenicity risk, or the molecular causes of inhibitor formation. The mechanism for determining the clinical first-in-human (FIH) dose of AAV-based gene therapy continues to pose challenges. This study aims to develop and validate a population pharmacokinetic and pharmacodynamics model (PK/PD) of VGB-R04 gene therapy for prediction of clinical dose. The pharmacodynamics, pharmacokinetic, and immunogenicity studies of VGB-R04 via intravenous injection in mice and cynomolgus monkeys were conducted to support an investigational new drug (IND) application. The end points included pharmacodynamic biomarkers, biodistribution, viral shedding, clinical pathology and histopathology, anti-AAV8 neutralizing antibodies, and anti-hFIX Padua protein antibody test. The peak concentration was noted 1 h after injection, subsequently exhibiting a distinct decline over time. The elimination rate of target genes in mice blood exceeded that in cynomolgus monkeys. The concentration in liver tissues indicated distinct liver tissue tropism. The elimination rate of target genes in mice livers exceeded that in cynomolgus monkeys. The plasma concentration of hFIX Padua protein exhibited a dose-dependent elevation in mice at doses of 8 × 10¹¹, 2.4 × 10¹², and 8 × 10¹² vg/kg. Cynomolgus monkeys exhibited significant elevation in plasma concentrations of hFIX Padua protein at 4 × 10¹³ vg/kg. A significant reduction in FIX activity and hFIX protein was observed in most of the animals starting about 4 weeks after dosing. In most animals, anti-hFIX Padua neutralizing antibody titers were detected at about week 4 of the monkeys and correlated with the preceding reductions in hFIX expression. Anti-AAV8 neutralizing antibodies can be detected in both species, but no antibodies against anti-hFIX Padua were found in mice. The research revealed the potential pharmacological and immunogenicity benefits, pharmacokinetic characteristics with target distribution, and possible safety of VGB-R04 in mice and cynomolgus monkeys following a single dosage. The model’s adequacy and robustness were assessed by VPC and bootstrap methods. Utilizing these data, we formulated a semimechanistic pharmacokinetic/pharmacodynamic quantitative model at a dosage of 4 × 10¹² vg/kg to enhance clinical translation, optimize clinical decision-making, and inform personalized therapy, despite the absence of suitable quantitative published data for developing pharmacokinetic models in gene therapy.