Volume 24 Issue 1 - May 24, 2013 PDF
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Cell secretome analysis using hollow fiber culture system leads to the discovery of CLIC1 protein as a novel plasma marker for nasopharyngeal carcinoma
Ying-Hwa Changa, Chih-Ching Wub, Kai-Ping Changc, Jau-Song Yub,d, Yu-Chen Change, Pao-Chi Liaoa,e,f,*
a Institute of Biopharmaceutical Sciences, College of Medicine, National Cheng Kung University, Tainan, Taiwan
b Molecular Medicine Research Center, Chang Gung University, Tao-Yuan, Taiwan
c Department of Otolaryngology-Head Neck Surgery, Chang Gung Memorial Hospital, Lin-Kou, Taiwan
d Department of Biochemistry and Molecular Biology, Chang Gung University, Tao-Yuan, Taiwan
e Department of Environmental and Occupational Health, College of Medicine, National Cheng Kung University, Tainan, Taiwan
f Sustainable Environment Research Center, National Cheng Kung University, Tainan, Taiwan
 
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Nasopharyngeal carcinoma (NPC) is one of the most common malignant tumors in Southeast Asia. The incidence rate of NPC among the Chinese, particularly population of southern China, is approximately 100-fold higher than that of other countries1. Due to its deep location and vague symptoms, majority of NPC victims, however unfortunately, have had metastasis at the time when first diagnosed. Discovery of sensitive and specific biomarkers for improving detection of NPC remains a challenge. The proteins secreted from cells are referred as secretome. In cancer progression, some particularly interesting proteins released from the tumor cells can enter into blood circulation. These proteins could be detected in serum or plasma and used as cancer screening markers. A strategy for cell secretome analysis using a hollow fiber culture (HFC) system combined with liquid chromatography mass spectrometry2 was previously established earlier in house as shown in Fig. 1.

Figure 1. Outline of experimental work-flow for NPC-TW04 cell secretome analysis, and potential biomarker selection.


Using the platform NPC secretome was collected and analyzed for the discovery of relevant clinical biomarkers. A total of 66 identified proteins were identified, among which, chloride intracellular channel 1 (CLIC1) was sieved out as a potential candidate for NPC biomarker. Approximately 75% of NPC tissue specimens showed positive CLIC1 staining by IHC. Figure 2 shows the plasma levels of CLIC1 in NPC patients (N=70), as presented by sandwich ELISA, were significantly higher than those in the healthy controls (N=74) (mean ± SD, 16.38 ± 26.53 vs. 2.39 ± 5.32, µg/mL; p = 0.00005). This is the first report for the detection of novel CLIC1 as a plasma marker for the early detection of NPC supported by statistical analysis. Our results indicate that the analytical platform could provide a feasible strategy to benefit the tumor cell secretome analysis for the identification of cancer biomarkers.

Figure 2. Elevated CLIC1 levels in NPC plasma samples. (A) The plasma CLIC1 levels of healthy controls (N=74), NPC patients (N=70), lung cancer patients (N=43), and colon cancer patients (N=45) were measured by sandwich ELISA using 5 μL of plasma samples. (B) ROC curve analysis of the diagnostic efficacy of CLIC1. (C) The correlation of CLIC1 plasma levels between the early and late TNM stages of NPC. (D) The correlation of CLIC1 plasma levels between the healthy controls and the early TNM stages of NPC.


References
  1. Chan, A. T.; Teo, P. M.; Johnson, P. J., Nasopharyngeal carcinoma. Ann. Oncol. 2002, 13, (7), 1007-15.
  2. Wu, H. Y.; Chang, Y. H.; Chang, Y. C.; Liao, P. C., Proteomics analysis of nasopharyngeal carcinoma cell secretome using a hollow fiber culture system and mass spectrometry. J. Proteome Res. 2009, 8, (1), 380-9.
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