Synthesis of Peptide-Oligonucleotide Conjugates via Condensation of a Lysine Side-Chain with a Thioester

Peptide-oligonucleotide conjugates (POCs) are promising drug delivery tools for oligonucleotide therapeutics. They are conventionally prepared by separately synthesizing peptides and oligonucleotides, introducing reactive functional groups into both molecules, and forming conjugates via click chemistry or maleimide-thiol coupling. However, these approaches generally require prefunctionalization of peptides or rely on the presence of cysteine residues, limiting their applicability to native peptides. Consequently, conjugation of peptides lacking thiol groups on cysteine residues to oligonucleotides remains challenging. Therefore, we aimed to develop a new conjugation method for POCs that does not require chemical modification of native peptides. To achieve this objective, we devised a method to prepare POCs via condensation between the ε-amino group of a lysine side-chain and a thioester-modified oligodeoxynucleotide. In establishing our POC synthesis method, the addition of 2-sulfanylmethyl-4-dimethylaminopyridine, which increases thioester reactivity, enabled rapid and efficient conjugation under mild conditions. Under the optimized conditions, conjugation proceeded efficiently with amino acids and simple trimer peptides having unprotected α-amino groups. Furthermore, conjugation with functional peptides demonstrated that this method is applicable to the synthesis of both linear and cyclic peptide-oligodeoxynucleotide conjugates.