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Phosphoproteome and protein kinase activity in fresh-frozen colorectal cancer tissue obtained from patients

Xiao Wang

Kinase activity profiling in tumour samples by peptide microarrays and mass spectrometry-based phosphoproteomics may influence the selection of protein kinase inhibitors in cancer patients due to their ability to detect aberrant cellular signaling in connection to biological function. Variable tissue handling processes in clinical practise can affect protein phosphorylation state and kinase activity, making biomarker finding more difficult. Using peptide microarrays and mass spectrometry-based phosphoproteomics, the influence of cold ischemia time (CIT) on the stability of kinase activity and protein phosphorylation status in fresh-frozen clinical tissue samples was investigated. Five patients' biopsies of colorectal cancer resection specimens were taken and snap frozen immediately after surgery and at intervals between CIT. A peptide microarray was used to profile kinase activity in all samples. At several time points, MS-based global phosphoproteomics was done in malignancies from patients. Changes in kinase activity and phosphoproteome as a result of CIT were studied using statistical and cluster analysis. The bulk of the phosphoproteome and protein kinase activity in colorectal cancer resection tissue is stable up to minutes after CIT and reflects tumour features. However, with increased CIT, specific alterations in kinase activity were detected. To limit changes in kinase activity during CIT, strict tissue collection protocols are recommended.