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Inhibiting EGFR dimerization using triazolyl-bridged dimerization arm mimics.

Hanold LE, Oruganty K, Ton NT, Beedle AM, Kannan N, Kennedy EJ - PLoS ONE (2015)

Bottom Line: In this study, we demonstrate that these peptides have significantly improved proteolytic stability over the non-modified peptide sequence, and their inhibitory effects are dependent on the number of the methylene units and orientation of the introduced triazolyl bridge.We identified a peptide, EDA2, which downregulates receptor phosphorylation and dimerization and reduces cell viability.This is the first example of a biologically active triazolyl-bridged peptide targeting the EGFR dimerization interface that effectively downregulates EGFR activation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical and Biomedical Sciences, College of Pharmacy, University of Georgia, Athens, Georgia, United States of America.

ABSTRACT
The epidermal growth factor receptor (EGFR) is overexpressed in multiple carcinomas and is the focus of a variety of targeted therapies. Here we report the design of peptide-based compounds that mimic the EGFR dimerization arm and inhibit allosteric activation of EGFR. These peptides are modified to contain a triazolyl bridge between the peptide strands to constrain the EGFR dimerization arm β-loop. In this study, we demonstrate that these peptides have significantly improved proteolytic stability over the non-modified peptide sequence, and their inhibitory effects are dependent on the number of the methylene units and orientation of the introduced triazolyl bridge. We identified a peptide, EDA2, which downregulates receptor phosphorylation and dimerization and reduces cell viability. This is the first example of a biologically active triazolyl-bridged peptide targeting the EGFR dimerization interface that effectively downregulates EGFR activation.

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Design and synthesis of EDA peptides.(a) Peptide sequences were derived from the dimerization arm sequence of EGFR. The overall linker length and positioning of the azide and alkyne amino acids were varied. Non-natural amino acids are show in red and blue. (b) Dimerization arm mimics were synthesized by incorporating non-natural amino acids into the peptide sequence using solid phase peptide synthesis (SPPS). Peptides were cyclized on solid support via copper (I)-catalyzed azide-alkyne cycloaddition prior to resin cleavage. (c) Non-natural amino acids used for the triazole cross-link: N-Fmoc-L-propargylglycine (Pg), N-Fmoc-4-azido-L-homoalanine (Aha), N-Fmoc-5-azido-L-norvaline (Anv), N-Fmoc-6-azido-L-norleucine (Anl).
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pone.0118796.g002: Design and synthesis of EDA peptides.(a) Peptide sequences were derived from the dimerization arm sequence of EGFR. The overall linker length and positioning of the azide and alkyne amino acids were varied. Non-natural amino acids are show in red and blue. (b) Dimerization arm mimics were synthesized by incorporating non-natural amino acids into the peptide sequence using solid phase peptide synthesis (SPPS). Peptides were cyclized on solid support via copper (I)-catalyzed azide-alkyne cycloaddition prior to resin cleavage. (c) Non-natural amino acids used for the triazole cross-link: N-Fmoc-L-propargylglycine (Pg), N-Fmoc-4-azido-L-homoalanine (Aha), N-Fmoc-5-azido-L-norvaline (Anv), N-Fmoc-6-azido-L-norleucine (Anl).

Mentions: Since the dimerization arm plays a major role in the stabilization of the extracellular receptor dimer, multiple mimics were previously designed [17–21]. As an alternative strategy to covalently constrain the dimerization arm, we utilized cycloaddition chemistry to introduce a 1,4-disubstituted [1,2,3]-triazolyl-containing bridge between the terminal residues of the sequence. A panel of EGFR Dimerization Arm (EDA) peptides was designed using the native sequence of human EGFR (residues 269–278, Fig. 2a). The β-strand and turn residues were conserved from the original amino acid sequence since the majority of these residues make extensive contacts with the other receptor half-site.


Inhibiting EGFR dimerization using triazolyl-bridged dimerization arm mimics.

Hanold LE, Oruganty K, Ton NT, Beedle AM, Kannan N, Kennedy EJ - PLoS ONE (2015)

Design and synthesis of EDA peptides.(a) Peptide sequences were derived from the dimerization arm sequence of EGFR. The overall linker length and positioning of the azide and alkyne amino acids were varied. Non-natural amino acids are show in red and blue. (b) Dimerization arm mimics were synthesized by incorporating non-natural amino acids into the peptide sequence using solid phase peptide synthesis (SPPS). Peptides were cyclized on solid support via copper (I)-catalyzed azide-alkyne cycloaddition prior to resin cleavage. (c) Non-natural amino acids used for the triazole cross-link: N-Fmoc-L-propargylglycine (Pg), N-Fmoc-4-azido-L-homoalanine (Aha), N-Fmoc-5-azido-L-norvaline (Anv), N-Fmoc-6-azido-L-norleucine (Anl).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4366150&req=5

pone.0118796.g002: Design and synthesis of EDA peptides.(a) Peptide sequences were derived from the dimerization arm sequence of EGFR. The overall linker length and positioning of the azide and alkyne amino acids were varied. Non-natural amino acids are show in red and blue. (b) Dimerization arm mimics were synthesized by incorporating non-natural amino acids into the peptide sequence using solid phase peptide synthesis (SPPS). Peptides were cyclized on solid support via copper (I)-catalyzed azide-alkyne cycloaddition prior to resin cleavage. (c) Non-natural amino acids used for the triazole cross-link: N-Fmoc-L-propargylglycine (Pg), N-Fmoc-4-azido-L-homoalanine (Aha), N-Fmoc-5-azido-L-norvaline (Anv), N-Fmoc-6-azido-L-norleucine (Anl).
Mentions: Since the dimerization arm plays a major role in the stabilization of the extracellular receptor dimer, multiple mimics were previously designed [17–21]. As an alternative strategy to covalently constrain the dimerization arm, we utilized cycloaddition chemistry to introduce a 1,4-disubstituted [1,2,3]-triazolyl-containing bridge between the terminal residues of the sequence. A panel of EGFR Dimerization Arm (EDA) peptides was designed using the native sequence of human EGFR (residues 269–278, Fig. 2a). The β-strand and turn residues were conserved from the original amino acid sequence since the majority of these residues make extensive contacts with the other receptor half-site.

Bottom Line: In this study, we demonstrate that these peptides have significantly improved proteolytic stability over the non-modified peptide sequence, and their inhibitory effects are dependent on the number of the methylene units and orientation of the introduced triazolyl bridge.We identified a peptide, EDA2, which downregulates receptor phosphorylation and dimerization and reduces cell viability.This is the first example of a biologically active triazolyl-bridged peptide targeting the EGFR dimerization interface that effectively downregulates EGFR activation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical and Biomedical Sciences, College of Pharmacy, University of Georgia, Athens, Georgia, United States of America.

ABSTRACT
The epidermal growth factor receptor (EGFR) is overexpressed in multiple carcinomas and is the focus of a variety of targeted therapies. Here we report the design of peptide-based compounds that mimic the EGFR dimerization arm and inhibit allosteric activation of EGFR. These peptides are modified to contain a triazolyl bridge between the peptide strands to constrain the EGFR dimerization arm β-loop. In this study, we demonstrate that these peptides have significantly improved proteolytic stability over the non-modified peptide sequence, and their inhibitory effects are dependent on the number of the methylene units and orientation of the introduced triazolyl bridge. We identified a peptide, EDA2, which downregulates receptor phosphorylation and dimerization and reduces cell viability. This is the first example of a biologically active triazolyl-bridged peptide targeting the EGFR dimerization interface that effectively downregulates EGFR activation.

Show MeSH
Related in: MedlinePlus