10.1021/pr900468k.s001 Hiroshi Kawase Hiroshi Kawase Kiyonaga Fujii Kiyonaga Fujii Masaki Miyamoto Masaki Miyamoto Kanako C. Kubota Kanako C. Kubota Satoshi Hirano Satoshi Hirano Satoshi Kondo Satoshi Kondo Fuyuhiko Inagaki Fuyuhiko Inagaki Differential LC−MS-Based Proteomics of Surgical Human Cholangiocarcinoma Tissues American Chemical Society 2009 DJ biomarker discovery phase protein LC candidate biomarkers test case samples SILAC bile duct tissue samples bile duct cells cholangiocarcinoma case samples Surgical Human Cholangiocarcinoma TissuesCholangiocarcinoma 2009-08-07 00:00:00 Journal contribution https://acs.figshare.com/articles/journal_contribution/Differential_LC_MS_Based_Proteomics_of_Surgical_Human_Cholangiocarcinoma_Tissues/2837800 Cholangiocarcinoma is an intractable cancer for which there is no effective therapy other than surgical resection, and many patients are not candidates for this treatment. Even for patients who undergo surgical resection, the 5-year survival rate is low. One reason for this is that the disease is often detected in late stages. Thus, there is a clear need for better biomarkers to facilitate early diagnosis and prognostication. During the biomarker discovery phase of our study, we used LC−MS-based proteomics with spectral counting, a semiquantitative approach to differential expression profiling, in paired cancerous and normal bile duct tissue samples from two cases. In total, 38 proteins up-regulated in the cancer samples were identified. These were verified using a SILAC method for MS-based validation. The results led to the identification of well-characterized proteins and proteins of unknown function that are up-regulated in cholangiocarcinoma. We used immunoblot analysis to validate four candidate biomarkers, actinin-1, actinin-4, protein DJ-1 and cathepsin B, with the test case samples and four additional cholangiocarcinoma case samples. Each of the four candidate proteins was overexpressed in a subset of five of the six cases tested. By immunohistochemistry, we further confirmed that expression of these proteins was elevated in cancer cells as compared with normal bile duct cells. Thus, we successfully identified several proteins up-regulated in cholangiocarcinoma. These proteins are candidate biomarkers and may also help to provide new insights into our understanding of the disease.