Unprotected peptide macrocyclization and stapling via a fluorine-thiol displacement reaction

Md Shafiqul Islam,Robert Maloney,Vincent A Voelz,Weidong Yang,Parmila Kafley,Zakey Yusuf Buuh,Si Zhang,Samuel L Junod,Rongsheng E Wang,Carson Cohen,Mi Zhao,Kishan H Kaneria,Zhigang Lyu,Yifu Guan

doi:10.1038/s41467-022-27995-5

Md Shafiqul Islam, Robert Maloney + Show 12 more

Open Access

https://doi.org/10.1038/s41467-022-27995-5

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Journal: Nature Communications	Publication Date: Jan 17, 2022
Citations: 13	License type: open-access

Affiliation: Temple University

Abstract

We report the discovery of a facile peptide macrocyclization and stapling strategy based on a fluorine thiol displacement reaction (FTDR), which renders a class of peptide analogues with enhanced stability, affinity, cellular uptake, and inhibition of cancer cells. This approach enabled selective modification of the orthogonal fluoroacetamide side chains in unprotected peptides in the presence of intrinsic cysteines. The identified benzenedimethanethiol linker greatly promoted the alpha helicity of a variety of peptide substrates, as corroborated by molecular dynamics simulations. The cellular uptake of benzenedimethanethiol stapled peptides appeared to be universally enhanced compared to the classic ring-closing metathesis (RCM) stapled peptides. Pilot mechanism studies suggested that the uptake of FTDR-stapled peptides may involve multiple endocytosis pathways in a distinct pattern in comparison to peptides stapled by RCM. Consistent with the improved cell permeability, the FTDR-stapled lead Axin and p53 peptide analogues demonstrated enhanced inhibition of cancer cells over the RCM-stapled analogues and the unstapled peptides.

Highlights

We report the discovery of a facile peptide macrocyclization and stapling strategy based on a fluorine thiol displacement reaction (FTDR), which renders a class of peptide analogues with enhanced stability, affinity, cellular uptake, and inhibition of cancer cells
We have demonstrated a new, mild, and clean synthetic strategy to cyclize and/or staple unprotected peptides
The developed fluorine–thiol displacement reaction (FTDR) approach operates at mild temperature in aqueous solutions and offers excellent chemoselectivity and functional group tolerance, e.g., sparing intrinsic cysteines

Summary

Introduction

We report the discovery of a facile peptide macrocyclization and stapling strategy based on a fluorine thiol displacement reaction (FTDR), which renders a class of peptide analogues with enhanced stability, affinity, cellular uptake, and inhibition of cancer cells. This approach enabled selective modification of the orthogonal fluoroacetamide side chains in unprotected peptides in the presence of intrinsic cysteines. Targeting PPIs has since emerged as a promising therapeutic strategy that inhibits specific molecular pathways without compromising other functions of the involved proteins[5] This avenue is challenging due to the generally flat, shallow, and extended nature of PPI interfaces[1,6,7]. The development of stapled peptides with designable cell permeability remains challenging and requires a delicate balance between positive charges, hydrophobicity, alpha-helicity, and staple position[35,36,37]

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Conclusion