Development of Aqueous and Lyophilized Formulations for LBURGINAZE™

LBURGINAZE™ is an engineered human arginase-1 fusion protein developed to exploit tumor metabolic vulnerabilities via systemic arginine depletion. Developing a stable formulation is necessary to preserve its enzymatic activity during long-term storage and clinical translation. Thus, this study aimed to systematically screen excipients and rationally develop aqueous and lyophilized formulations of LBURGINAZE.

A panel of pharmaceutically acceptable excipients, including sugars, polyols, and amino acids, was screened under thermal stress at 25℃ by measuring arginine hydrolytic activity. Formulations were evaluated in both liquid and freeze-dried states, with lead single excipients that preserved catalytic activity subsequently incorporated into multi-component formulations. Following stability assessments, the biological potency of reconstituted LBURGINAZE was evaluated against 3D HepG2 hepatocellular carcinoma spheroids alone and in combination with the multi-kinase inhibitor sorafenib.

In aqueous state, a multi-component formulation containing glycerol, L-alanine, NaCl, and polysorbate 80 provided optimal stability, preserving catalytic activity over 4 weeks. For solid-state storage, a single-excipient lyophilization containing sucrose provided superior lyoprotection, effectively maintaining 100% of arginine hydrolytic activity across 90 days at 25℃. Upon reconstitution, LBURGINAZE retained potent, concentration-dependent anti-proliferative efficacy against HepG2 3D tumor spheroids (IC₅₀ = 5.49 µg/mL) and displayed strong pharmacological synergism when combined with sorafenib.

These rational formulation approaches provide strong stability for LBURGINAZE™ in both aqueous and solid states at ambient conditions, providing a scalable platform to support its ongoing preclinical development.