Ischemia in tumors induces early and sustained phosphorylation changes in stress kinase pathways but does not affect global protein levels.
Author(s): Mertins P, Yang F, Liu T, Mani DR, Petyuk VA, Gillette MA, Clauser KR, Qiao JW, Gritsenko MA, Moore RJ, Levine DA, Townsend R, Erdmann-Gilmore P, Snider JE, Davies SR, Ruggles KV, Fenyo D, Kitchens RT, Li S, Olvera N, Dao F, Rodriguez H, Chan DW, Liebler D, White F, Rodland KD, Mills GB, Smith RD, Paulovich AG, Ellis M, Carr SA
Publication: Mol Cell Proteomics, 2014, Vol. 13, Page 1690-704
PubMed ID: 24719451 PubMed Review Paper? No
Purpose of Paper
The purpose of this paper was to determine the effects of cold ischemia time on the proteome and phosphoproteome in snap frozen ovarian cancer specimens.
Conclusion of Paper
While cold ischemia times of up to 60 min did not alter the overall abundance of any individual protein, changes in the phosphoproteome were noted. Specifically, 4.6% of phosphorylation sites were up-regulated, and 1.2% of sites were down-regulated (>25,000 were quantified) compared to baseline. The timing of the ischemia-induced phosphorylation changes was variable. A strong correlation was observed between results obtained with reverse phase protein analysis and LC-MS/MS. Reverse phase protein analysis also showed no proteomic changes due to cold ischemia. The majority of cold ischemia-affected phosphosites detected by reverse phase protein analysis but not by LC-MS/MS were not well suited for mass spectrometry for reasons such as peptide length.
Studies
-
Study Purpose
The purpose of this study was to determine the effects of cold ischemia time on the proteome and phosphoproteome in snap frozen ovarian cancer specimens. Tumors from four patients were used in this study and dissected into 4 pieces which were exposed to various room temperature cold ischemia times before being snap frozen in liquid nitrogen. The authors note that blood vessels were not ligated prior to tumor excision so no warm ischemia time occurred. The authors also present data obtained from breast cancer xenograft tumors which is not discussed here.
Summary of Findings:
While cold ischemia times of up to 60 min did not alter the overall abundance of any individual protein levels, changes in the phosphoproteome with cold ischemia were noted. Specifically, 4.6% of phosphorylation sites were up-regulated and 1.2% of sites were down-regulated (>25,000 were quantified) compared to baseline. Tyrosine and serine/threonine phosphorylation sites were equally affected by cold ischemia. The timing of the ischemia-induced phosphorylation changes was variable and included activation of the Mitogen-activated protein kinase (MAPK) cascade, regulation of the Rho, Ras, and GTPase signaling pathways, activation of transcriptional regulators, regulation of chromatin modification and assembly, and regulation of cytoskeleton organization. A strong correlation was observed between results obtained using reverse phase protein analysis and LC-MS/MS (r=0.79). Reverse phase protein analysis also showed no proteomic changes due to cold ischemia. The majority of cold ischemia-affected phosphosites detected by reverse phase protein analysis but not by LC-MS/MS were not well suited for mass spectrometry for reasons such as peptide length.
Biospecimens
Preservative Types
- Frozen
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform Protein LC-MS or LC-MS/MS Peptide LC-MS or LC-MS/MS Protein Reverse phase protein microarray Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) LC-MS or LC-MS/MS Specific Technology platform Reverse phase protein analysis
Biospecimen Acquisition Cold ischemia time <1 min
5 min
30 min
60 min