Antibodies raised against human targets often fail to recognize their animal orthologs, limiting preclinical evaluation in relevant models. We developed a Deep Mutational Scanning (DMS)-coupled deep learning strategy to engineer potent cross-reactive antibodies with minimal sequence divergence. Starting from C4, a fully human anti-PD-L1 antibody with weak recognition of murine PD-L1, DMS identified substitutions that improved binding to both human and mouse antigens. Conventional recombination of beneficial mutations generated highly cross-reactive antibodies but required 13 to 15 substitutions. To reduce this mutational burden, a deep learning model trained on DMS-derived sequence-binding data was used to identify minimal mutation combinations predicted to retain high affinity. This approach yielded variants carrying only 4 to 5 substitutions, with in vitro and cellular binding properties comparable to highly mutated antibodies. Epitope mapping, structural modeling and in vivo assessment further confirmed that these engineered antibodies retained PD-1/PD-L1 blockade and demonstrated therapeutic activity in a mouse tumor model.
H. Dorison, Anne-Laure Grindel, François Thenier et al.· bioRxiv· 0 citations
Mimicking complex protein-protein interfaces with small, well-defined molecular scaffolds remains a major challenge in chemical biology. Here, we report a foldamer-based downsizing strategy that compresses the quaternary architecture of the histone H3-H4 dimer into compact peptide-oligourea hybrids acting as high-affinity ligands of the histone chaperone Anti-Silencing Function 1 (ASF1). Guided by multiple high-resolution co-crystal structures, we designed a series of foldamer mimetics that accurately reproduce both the H3 α-helix and the H4 β-strand epitopes. Systematic optimization of linker geometry, β-strand mimicry, formal charge, and selective backbone N-methylation yielded highly stable ligands with nanomolar affinities, enhanced proteolytic resistance, and robust cytosolic penetration. Notably, the optimized constructs and their N-methylated analogues recapitulate the binding mode of the native H3-H4 dimer on ASF1 with high fidelity and engage endogenous ASF1 in cell extracts, demonstrating effective intracellular target recognition. Together, these results show that peptide-oligourea foldamers can reproduce the structural features of a protein quaternary structure surface, combining high affinity, high stability and cell permeability.
Bo Li, Marie E. Perrin, Emma Maillard et al.· Angewandte Chemie· 0 citations