Closed-Loop Multi-Objective Optimization for Receptor-Selective Cell-Penetrating Peptide Design

Abstract Cell-penetrating peptides (CPPs) can deliver diverse cargoes into cells. However, designing CPPs with receptor-selective interaction profiles remains difficult because interactions with individual cell-surface components cannot be tuned independently. Here, we developed a closed-loop in silico framework for receptor-selective CPP design, in which receptor interactions are formulated as explicit objectives in a multi-objective optimization problem. We first constructed a CPP-like candidate library by using a sequence generative model fine-tuned on known CPPs. The framework then evaluated candidate peptides by receptor-wise docking, molecular dynamics simulations, and MM/GBSA to compute receptor-wise binding scores as computational proxies for receptor-associated interactions. These scores were used iteratively to propose subsequent candidates by multi-objective Bayesian optimization. Applied to a CXCR4/NRP1 design setting, the framework identified candidates with more favorable predicted interaction profiles characterized by higher CXCR4 binding scores and lower NRP1 binding scores. We selected 10 peptides from the computationally identified candidates for cell-based imaging and found that 4 showed higher enrichment in CXCR4-positive regions than in NRP1-positive regions under the tested conditions. These results show that the proposed framework provides a practical in silico approach for designing CPPs with receptor-selective interaction profiles.