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Novel Aqueous Chemical Stabilization Method for Proteins Based on the Diels-alder Reaction

Published:
Lead Inventor: Raymond Moellering
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SUMMARY

  • Stabilization of therapeutic proteins and peptides is often necessary to improve pharmacokinetic properties and reduce proteolysis in vivo. However, pre-existing stabilization strategies require expensive catalysts or harsh reagents that prevent preservation of certain sterically challenging protein structures.
  • The inventors found that the Diels-Alder reaction, when performed aqueously or in solid phase, can stabilize turn and loop peptides by macrocyclization.
  • The invention is a method of stabilizing peptides through carbon-carbon bonds that works on sterically challenging structures not amenable to traditional stabilization chemistry. The reaction can be performed aqueously or in solid phase, without harsh or expensive reagents.
  • In proof-of-concept studies with a model loop peptide (i, i+7), NMR showed the Diels-Alder reaction added structural rigidity in the peptide backbone. Moreover, in vitro experiments showed reduced protein lysis in the stabilized peptide as compared to the control.

FIGURE

 

In vitro stabilization assay for Diels-Alder stabilized peptides (red) as compared to linear peptides (blue). Both different peptides and different degradation enzymes were tested. As shown, stabilized peptides retain original shape for longer before degradation.

 

 

ADVANTAGES

ADVANTAGES

  • Performed in aqueous or solid phase
  • No expensive catalysts or toxic chemicals
  • Preserves sterically challenging structures
  • Stereo selective

 

APPLICATIONS

  • Protein engineering
  • Antibody pharmacokinetics
  • Stapled peptides

 

PUBLICATIONS

  • Montgomery, JE; Donnelly, JA; Speltz, TE; Coulos, JS; Moellering, RE. Versatile peptide macrocyclization with Diels-Alder cycloadditions. In preparation.