Routine preparation of thiol oligonucleotides: application to the synthesis of oligonucleotide-peptide hybrids.

Oligonucleotide-peptide hybrids have potential for use as antisense inhibitors of gene expression, with the peptide helping to increase the intracellular concentration of the active oligonucleotide. The preparation of such hybrids can be achieved by the coupling of thiol-derivatized oligonucleotides with maleimido-peptides. We have developed reliable methods for preparing 5'-thiol oligonucleotides in good yields using phosphoramidite chemistry and coupling 6-(tritylthio)hexyl phosphoramidite as the 5'-terminal residue. The use of highly pure thiol phosphoramidite as well as a manual iodine treatment after this coupling were found to be important. Oligonucleotide-peptide hybrids were prepared in high yield (85%) by reacting freshly purified 5'-thiol oligonucleotides with peptides derivatized at their N-terminus with a maleimido functionality.