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High sensitivity isoelectric focusing to establish a signaling biomarker for the diagnosis of human colorectal cancer
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Stockholm Bioinformat Ctr, Sci Life Lab, Box 1031, S-17121 Solna, Sweden.;Natl Cheng Kung Univ, Dept Mech Engn, 1 Univ Rd, Tainan 70101, Taiwan..
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Colorectal Surgery.
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2016 (English)In: BMC Cancer, ISSN 1471-2407, E-ISSN 1471-2407, Vol. 16, 683Article in journal (Refereed) Published
Abstract [en]

Background: The progression of colorectal cancer (CRC) involves recurrent amplifications/mutations in the epidermal growth factor receptor (EGFR) and downstream signal transducers of the Ras pathway, KRAS and BRAF. Whether genetic events predicted to result in increased and constitutive signaling indeed lead to enhanced biological activity is often unclear and, due to technical challenges, unexplored. Here, we investigated proliferative signaling in CRC using a highly sensitive method for protein detection. The aim of the study was to determine whether multiple changes in proliferative signaling in CRC could be combined and exploited as a "complex biomarker" for diagnostic purposes. Methods: We used robotized capillary isoelectric focusing as well as conventional immunoblotting for the comprehensive analysis of epidermal growth factor receptor signaling pathways converging on extracellular regulated kinase 1/2 (ERK1/2), AKT, phospholipase C gamma 1 (PLC gamma 1) and c-SRC in normal mucosa compared with CRC stage II and IV. Computational analyses were used to test different activity patterns for the analyzed signal transducers. Results: Signaling pathways implicated in cell proliferation were differently dysregulated in CRC and, unexpectedly, several were downregulated in disease. Thus, levels of activated ERK1 (pERK1), but not pERK2, decreased in stage II and IV while total ERK1/2 expression remained unaffected. In addition, c-SRC expression was lower in CRC compared with normal tissues and phosphorylation on the activating residue Y418 was not detected. In contrast, PLC gamma 1 and AKT expression levels were elevated in disease. Immunoblotting of the different signal transducers, run in parallel to capillary isoelectric focusing, showed higher variability and lower sensitivity and resolution. Computational analyses showed that, while individual signaling changes lacked predictive power, using the combination of changes in three signaling components to create a "complex biomarker" allowed with very high accuracy, the correct diagnosis of tissues as either normal or cancerous. Conclusions: We present techniques that allow rapid and sensitive determination of cancer signaling that can be used to differentiate colorectal cancer from normal tissue.

Place, publisher, year, edition, pages
2016. Vol. 16, 683
Keyword [en]
Colorectal cancer, Isoelectric focusing, Signal transduction, Proliferation, ERK, c-SRC
National Category
Cancer and Oncology
Identifiers
URN: urn:nbn:se:uu:diva-305935DOI: 10.1186/s12885-016-2725-zISI: 000384194500001PubMedID: 27562229OAI: oai:DiVA.org:uu-305935DiVA: diva2:1046644
Funder
Swedish Cancer Society, CAN2013/661Knut and Alice Wallenberg Foundation, KAW 2015.0275eSSENCE - An eScience Collaboration
Available from: 2016-11-14 Created: 2016-10-24 Last updated: 2016-11-14Bibliographically approved

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Padhan, NarendraNordling, Torbjorn E. M.Sundström, MagnusÅkerud, PeterBirgisson, HelgiNygren, PeterNelander, SvenClaesson-Welsh, Lena
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