Browsing by Author "Chan, Chung-Chuan"

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An Informatics-assisted Label-free Approach for Personalized Tissue Membrane Proteomics: Case Study on Colorectal Cancer
(2011-04-01)
Han, Chia-Li
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Chen, Jinn-Shiun
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Chan, Err-Cheng
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Wu, Chien-Peng
;
Yu, Kun-Hsing
We developed a multiplexed label-free quantification strategy, which integrates an efficient gel-assisted digestion protocol, high-performance liquid chromatography tandem MS analysis, and a bioinformatics alignment method to determine personalized proteomic profiles for membrane proteins in human tissues. This strategy provided accurate (6% error) and reproducible (34% relative S. D.) quantification of three independently purified membrane fractions from the same human colorectal cancer (CRC) tissue. Using CRC as a model, we constructed the personalized membrane protein atlas of paired tumor and adjacent normal tissues from 28 patients with different stages of CRC. Without fractionation, this strategy confidently quantified 856 proteins (>= 2 unique peptides) across different patients, including the first and robust detection (Mascot score: 22,074) of the well-documented CRC marker, carcinoembryonic antigen 5 by a discovery-type proteomics approach. Further validation of a panel of proteins, annexin A4, neutrophils defensin A1, and claudin 3, confirmed differential expression levels and high occurrences (48-70%) in 60 CRC patients. The most significant discovery is the overexpression of stomatin-like 2 (STOML2) for early diagnostic and prognostic potential. Increased expression of STOML2 was associated with decreased CRC-related survival; the mean survival period was 34.77 +/- 2.03 months in patients with high STOML2 expression, whereas 53.67 +/- 3.46 months was obtained for patients with low STOML2 expression. Further analysis by ELISA verified that plasma concentrations of STOML2 in early-stage CRC patients were elevated as compared with those of healthy individuals (p < 0.001), suggesting that STOML2 may be a noninvasive serological biomarker for early CRC diagnosis. The overall sensitivity of STOML2 for CRC detection was 71%, which increased to 87% when combined with CEA measurements. This study demonstrated a sensitive, label-free strategy for differential analysis of tissue membrane proteome, which may provide a roadmap for the subsequent identification of molecular target candidates of multiple cancer types. Molecular & Cellular Proteomics 10: 10.1074/mcp.M110.003087, 1-15, 2011.
Detection of autoantibodies against Rabphilin-3A-like protein as a potential biomarker in patient's sera of colorectal cancer
(2011-07-15)
Chen, Jinn-Shiun
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Kuo, Yung-Bin
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Chou, Yeh-Pin
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Chan, Chung-Chuan
;
Fan, Chung-Wei
Background: Rabphilin-3A-like (RPH3AL) protein functions in the regulation of hormone exocytosis, and mutations in the RPHA3L gene have been associated with tumorigenesis in colorectal cancer (CRC). We evaluated the potential use of anti-RPH3AL autoantibodies as a marker for CRC detection. Methods: Sera from 84 patients with CRC and 63 healthy controls were analysed for the presence of RPH3AL autoantibodies with a Western blotting assay. Results: The frequencies of RPH3AL autoantibodies in the early stage, advanced stage and all CRC patients were 64.7%, 78.0% and 72.6%, respectively. These values are significantly higher than the frequency of RPH3AL autoantibodies in healthy controls (15.9%, P<0.001). Although the presence of RPH3AL autoantibodies did not correlate with clinical parameters, RPH3AL autoantibodies were found in 69.4% (34/49) of CRC patients who were negative for carcinoembryonic antigen. The value of the area under the receiver operating characteristic curve of RPH3AL autoantibody was 0.84, which suggests that screening for these autoantibodies could potentially be used for CRC diagnosis. Conclusion: Circulating RPH3AL autoantibodies are prevalent in patients with CRC, and detection of these autoantibodies might provide a novel non-invasive approach for CRC diagnosis. (C) 2011 Elsevier B.V. All rights reserved.
Identification of Phospholipid Scramblase 1 as a Biomarker and Determination of Its Prognostic Value for Colorectal Cancer
(2011-01-01)
Kuo, Yung-Bin
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Chan, Chung-Chuan
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Chang, C. Allen
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Fan, Chung-Wei
;
Hung, Ray-Ping
The purpose of this study was to examine the expression of phospholipid scramblase 1 (PLSCR1) in tumor tissues and plasma specimens of patients with colorectal cancer (CRC), as well as analyze its association with clinical parameters. The expression levels of PLSCR1 protein in 104 matched CRC and adjacent normal tissue sections and 50 pairs of CRC tissue blocks were determined by use of immunohistochemical and Western blot analyses, respectively. To evaluate the diagnostic potential of PLSCR1, the plasma levels of PLSCR1 were investigated in 111 additional subjects (59 CRC patients and 52 healthy controls) by Western blot. PLSCR1 was overexpressed in malignant adenocarcinoma tissues compared with normal colorectal mucosa (P < 0.001). In addition, the plasma level of PLSCR1 was not only significantly elevated in CRC patients compared with healthy individuals (P < 0.001), but it was also substantially increased in early stage CRC (P < 0.001). Importantly, the overall sensitivity and specificity of PLSCR1 for CRC detection were 80% and 59.6%, respectively. The area under the ROC curve of PLSCR1 for CRC diagnosis is 0.75, which increases to 0.8 if combined with the measurement of carcinoembryonic antigen. Univariate analysis with the Cox regression model revealed that elevated PLSCR1 expression indicated a poor prognosis for CRC, This study showed that PLSCR1 protein levels were significantly elevated in both the cancer tissue and plasma of CRC patients. Moreover, the plasma levels of PLSCR1 were significantly elevated in patients with early stage CRC compared with healthy individuals, suggesting that PLSCR1 might be used as a noninvasive serological diagnostic and prognostic biomarker for CRC. (c) 2011 The Feinstein Institute for Medical Research, www.feinsteininstitute.org Online address: hffp://www.molmed.org doi: 10.2119/molmed.2010.00115
Identification of SEC61 beta and its autoantibody as biomarkers for colorectal cancer
(2011-05-12)
Fan, Chung-Wei
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Chan, Chung-Chuan
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Chen, Kuei-Tien
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Twu, Jen
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Huang, Ya-Shu
Background: To identify novel serological biomarkers for human colorectal cancer (CRC). we analyzed CRC tissues using gel-assisted digestion and isobaric tags with related and absolute quantitation (iTRAQ) labeling mass spectrometry (MS). By comparing pairs of tumor tissues and matched normal tissues. we discovered the SEC61 beta with expression changes 3.3-fold and a marginal statistical significance (p=0.052) previously. Methods: SEC61 beta expression in CRC tissues was further analyzed by western blotting and immunohistochemistry. We next assessed the putative diagnostic value of the SEC61 beta autoantibody as a serum marker. Results: Using western blotting analysis. SEC61 beta expression was increased 1.9-fold in tumor tissues. Immunohistochemical analysis of 64 CRC specimens showed that SEC61 beta was positively detected in 64% of the tumors, but weakly or not detected in > 80% of the adjacent nontumor epithelial cells. Western blot analysis with plasma samples showed that the sensitivity and specificity of the SEC61 beta autoantibody from patients with CRC were 79% and 75%. respectively. Importantly, the results of the SEC61 beta autoantibody for early detection of colorectal cancer revealed a higher sensitivity of 77% than the carcinoembryonic antigen (CEA) assay. Conclusions: Measurement of SEC61 beta autoantibody levels may provide an alternative detection indicator for CRC, particularly among early-stage patients. (C) 2011 Elsevier B.V. All rights reserved.
Multiple serological biomarkers for colorectal cancer detection
(2010-04-01)
Chan, Chung-Chuan
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Fan, Chung-Wei
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Kuo, Yung-Bin
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Chen, Yu-Han
;
Chang, Pi-Yueh
The aim of this study was to initiate a survey of human autoantibody responses to a panel of select colorectal tumor-associated antigens identified by previous serological analysis of a cDNA expression library and to subsequently identify multiple serological biomarkers for the detection of colorectal cancer. For screening of autoantibodies against colorectal tumor-associated antigens, sera from 94 colorectal cancer patients and 54 normal controls were analyzed by enzyme-linked immunosorbent assay using recombinant rCCCAP, rHDAC5, rP53, rNMDAR and rNY-CO-16 proteins as coating antigens. Seropositivity among colorectal cancer patients to the 5 individual coating antigens varied from 18.1% to 35.1%. Seropositivity to any of the 5 coating antigens was 58.5% and combining this analysis with evaluation of serum carcinoembryonic antigen (>= 5 ng/ml) significantly increased the seropositivity to 77.6%. Seropositivity of early-stage (Dukes' Stages A and B) colorectal cancer patients to CEA was 21.9%, and seropositivity to any of the 5 colorectal cancer-associated antigens was 53.7%, and the combination of these 2 measurements resulted in a higher diagnostic capacity (65.9%) than either marker alone. In conclusion, these results collectively indicated that combined detection of serum autoantibody profiles against our panel of colorectal tumor-associated antigens and the analysis of carcinoembryonic antigen provides a promising diagnostic biomarker for colorectal cancer, particularly among early-stage patients.