Tissue phosphoproteomics with PolyMAC identifies potential therapeutic targets in a transgenic mouse model of HER2 positive breast cancer

Adam C. Searleman, Anton B. Iliuk, Timothy S. Collier, Lewis A. Chodosh, W. Andy Tao, Ron Bose

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

Abstract

Altered protein phosphorylation is a feature of many human cancers that can be targeted therapeutically. Phosphopeptide enrichment is a critical step for maximizing the depth of phosphoproteome coverage by MS, but remains challenging for tissue specimens because of their high complexity. We describe the first analysis of a tissue phosphoproteome using polymer-based metal ion affinity capture (PolyMAC), a nanopolymer that has excellent yield and specificity for phosphopeptide enrichment, on a transgenic mouse model of HER2-driven breast cancer. By combining phosphotyrosine immunoprecipitation with PolyMAC, 411 unique peptides with 139 phosphotyrosine, 45 phosphoserine, and 29 phosphothreonine sites were identified from five LC-MS/MS runs. Combining reverse phase liquid chromatography fractionation at pH 8.0 with PolyMAC identified 1571 unique peptides with 1279 phosphoserine, 213 phosphothreonine, and 21 phosphotyrosine sites from eight LC-MS/MS runs. Linear motif analysis indicated that many of the phosphosites correspond to well-known phosphorylation motifs. Analysis of the tyrosine phosphoproteome with the Drug Gene Interaction database uncovered a network of potential therapeutic targets centered on Src family kinases with inhibitors that are either FDA-approved or in clinical development. These results demonstrate that PolyMAC is well suited for phosphoproteomic analysis of tissue specimens.

Original languageEnglish
Pages (from-to)3463-3469
Number of pages7
JournalElectrophoresis
Volume35
Issue number24
DOIs
StatePublished - Dec 1 2014

Keywords

  • Breast Cancer
  • Drug identification
  • HER2
  • Phosphoproteomics
  • PolyMAC

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