Isostearyl Mixed Anhydrides for the Preparation ofN-Methylated Peptides UsingC-Terminally UnprotectedN-Methylamino Acids

Abstract

Sustainable and efficient manufacturing methods for N-methylated peptides remain underexplored despite growing interest in therapeutic N-methylated peptides within the pharmaceutical industry. A methodology for the coupling of C-terminally unprotected N-methylamino acids mediated by an isostearic acid halide (ISTAX) and silylating reagent has been developed. This approach allows for the coupling of a wide variety of amino acids and peptides in high yields under mild conditions without the need for a C-terminal deprotection step in the process of C-terminal elongation. These advantages make this a useful synthetic method for the production of peptide therapeutics and diagnostics containing N-methylamino acids.

Extended Solution-phase Peptide Synthesis Strategy Using Isostearyl-Mixed Anhydride Coupling and a New C-Terminal Silyl Ester-Protecting Group for N-Methylated Cyclic Peptide Production

“…The condensation was completed quickly within 2 h. The Oxyma-based condensation agent has higher thermal stability than triazole-based reagents; however, concerns remain about HCN gas generation . Unlike Oxyma and chloroformate, ISTA-Cl is a less-hazardous coupling agent with high thermal stability that does not generate HCN or CO 2 gas . The introduction of the chloroacetyl group was performed by adding chloroacetic anhydride, which was activated by N , N ′-diisopropylcarbodiimide (DIC) in situ.…”

“…Amino acids were purchased from Watanabe Chemical Industries, Tokyo Chemical Industry, GL Biochem, and Merck. ISTA-Cl was synthesized as previously described . Di- tert -butylsilane and Di- tert -butylisobutyl trifluoromethanesulfonate were purchased from Gelest.…”

“…Reverse elongation permits the rapid and environmentally friendly synthesis of peptide fragments for convergent LPPS in fewer steps due to elimination of the deprotection step . In a previous study, we showed that tetrapeptides containing N -methyl amino acids could be efficiently synthesized without epimerization using this method . One of the limitations of peptide fragment synthesis by convergent LPPS is C-terminal carboxy protection during peptide elongation.…”

“…LPPS also has a better green chemistry profile than SPPS as the reagent and solvent requirements can be minimized. 25 As such, we have developed a new coupling method using isostearic acid halide (ISTA-X) and silylation agents, 26 by which N-and C-terminal unprotected amino acids can be introduced without double insertion by a temporary protecting effect of the carboxyl functionality by the silyl group (Scheme 1).…”

“…25 In a previous study, we showed that tetrapeptides containing N-methyl amino acids could be efficiently synthesized without epimerization using this method. 26 One of the limitations of peptide fragment synthesis by convergent LPPS is C-terminal carboxy protection during peptide elongation. The protective groups must be selectively deprotected over N α -and side chain-protecting groups.…”