Computational Insights into the pH-Dependent Behavior of Ipilimumab-CTLA-4

Abstract

Therapeutic antibodies face on-target/off-tumor toxicity, causing adverse effects such as cardiotoxicity, skin rashes, and organ inflammation. To mitigate these challenges, pH-dependent antibodies have been engineered to preferentially bind in the acidic tumor microenvironment while reducing interactions under physiological pH conditions. Building on experimental work that generated Ipilimumab variants (Ipi95, Ipi105, Ipi106) through charged amino acid substitutions in complementarity-determining regions, we employed molecular dynamics simulations to examine their interactions with CTLA-4 in neutral and acidic conditions. All variants exhibited enhanced binding affinity at acidic pH, with good agreement between computational and experimental binding free energies (R 2 = 0.7736). Statistical analysis revealed significant differences across conditions, most notably for Ipi95, which demonstrated the highest degree of pH sensitivity. Key structural determinants distinguishing pH-dependent from non-pH-dependent variants were also identified. These results provide mechanistic insights into how targeted mutations modulate pH-dependent recognition, offering a framework for the rational design of safer therapeutic antibodies.