Impact of dipeptide on ADC physicochemical properties and efficacy identifies Ala–Ala as the optimal dipeptideElectronic supplementary information (ESI) available. See DOI: https://doi.org/10.1039/d3md00473b

Side chains of natural occurring amino acids vary greatly in terms of charge state, polarity, size and hydrophobicity. Using a linear synthetic route, two amino acids were sequentially coupled to a potent glucocorticoid receptor modulator (GRM) to afford a library of dipeptide-GRM linker payloads with a range of in silicoproperties. The linker payloads were conjugated to a mouse anti-TNF antibody through interchain disulfide Cys. Impact of various dipeptide linkers on ADC physical properties, including solubility, hydrophobicity, and aggregation were evaluated and the in silicoproperties pI, Log Pand tPSA of the linker drugs used to correlate with these properties. ADCs were screened in a GRE luciferase reporter assay to compare their in vitroefficacy. Data identified Ala–Ala as a superior dipeptide linker that allowed a maximum drug load of 10 while affording ADCs with low aggregation.