Quantitative Analysis of Gene Expression in Fixed Colorectal Carcinoma Samples As a Method for Biomarker Validation

Overview

Journal Mol Med Rep

Specialty Molecular Biology

Date 2016 Apr 29

PMID 27121919

Citations 10

Authors

Beata Ostasiewicz

Pawel Ostasiewicz

Kamila Dus-Szachniewicz

Katarzyna Ostasiewicz

Piotr Ziolkowski

Affiliations

Soon will be listed here.

Abstract

Biomarkers have been described as the future of oncology. Modern proteomics provide an invaluable tool for the near‑whole proteome screening for proteins expressed differently in neoplastic vs. healthy tissues. However, in order to select the most promising biomarkers, an independent method of validation is required. The aim of the current study was to propose a methodology for the validation of biomarkers. Due to material availability the majority of large scale biomarker studies are performed using formalin‑fixed paraffin‑embedded (FFPE) tissues, therefore these were selected for use in the current study. A total of 10 genes were selected from what have been previously described as the most promising candidate biomarkers, and the expression levels were analyzed with reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) using calibrator normalized relative quantification with the efficiency correction. For 6/10 analyzed genes, the results were consistent with the proteomic data; for the remaining four genes, the results were inconclusive. The upregulation of karyopherin α 2 (KPNA2) and chromosome segregation 1‑like (CSE1L) in colorectal carcinoma, in addition to downregulation of chloride channel accessory 1 (CLCA1), fatty acid binding protein 1 (FABP1), sodium channel, voltage gated, type VII α subunit (SCN7A) and solute carrier family 26 (anion exchanger), member 3 (SLC26A3) was confirmed. With the combined use of proteomic and genetic tools, it was reported, for the first time to the best of our knowledge, that SCN7A was downregulated in colorectal carcinoma at mRNA and protein levels. It had been previously suggested that the remaining five genes served an important role in colorectal carcinogenesis, however the current study provided strong evidence to support their use as biomarkers. Thus, it was concluded that combination of RT‑qPCR with proteomics offers a powerful methodology for biomarker identification, which can be used to analyze FFPE samples.

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References

Yang B, Cao L, Liu B, McCaig C, Pu J . The transition from proliferation to differentiation in colorectal cancer is regulated by the calcium activated chloride channel A1. PLoS One. 2013; 8(4):e60861. PMC: 3625186. DOI: 10.1371/journal.pone.0060861. View

Wisniewski J, Ostasiewicz P, Dus K, Zielinska D, Gnad F, Mann M . Extensive quantitative remodeling of the proteome between normal colon tissue and adenocarcinoma. Mol Syst Biol. 2012; 8:611. PMC: 3472694. DOI: 10.1038/msb.2012.44. View

Lawrie L, Dundas S, Curran S, Murray G . Liver fatty acid binding protein expression in colorectal neoplasia. Br J Cancer. 2004; 90(10):1955-60. PMC: 2409459. DOI: 10.1038/sj.bjc.6601828. View

Tung M, Stella Tsai C, Tung J, Tsao T, Chen H, Yeh K . Higher prevalence of secretory CSE1L/CAS in sera of patients with metastatic cancer. Cancer Epidemiol Biomarkers Prev. 2009; 18(5):1570-7. DOI: 10.1158/1055-9965.EPI-08-0948. View

Alshareeda A, Negm O, Green A, Nolan C, Tighe P, Albarakati N . KPNA2 is a nuclear export protein that contributes to aberrant localisation of key proteins and poor prognosis of breast cancer. Br J Cancer. 2015; 112(12):1929-37. PMC: 4580386. DOI: 10.1038/bjc.2015.165. View

Satoh Y, Mori K, Kitano K, Kitayama J, Yokota H, Sasaki H . Analysis for the combination expression of CK20, FABP1 and MUC2 is sensitive for the prediction of peritoneal recurrence in gastric cancer. Jpn J Clin Oncol. 2011; 42(2):148-52. DOI: 10.1093/jjco/hyr179. View

Hubbard K, Catalano J, Puri R, Gnatt A . Knockdown of TFIIS by RNA silencing inhibits cancer cell proliferation and induces apoptosis. BMC Cancer. 2008; 8:133. PMC: 2390572. DOI: 10.1186/1471-2407-8-133. View

Ma S, Zhao X . KPNA2 is a promising biomarker candidate for esophageal squamous cell carcinoma and correlates with cell proliferation. Oncol Rep. 2014; 32(4):1631-7. DOI: 10.3892/or.2014.3381. View

Liao C, Luo S, Li L, Lin C, Chen Y, Jiang M . CSE1L/CAS, the cellular apoptosis susceptibility protein, enhances invasion and metastasis but not proliferation of cancer cells. J Exp Clin Cancer Res. 2008; 27:15. PMC: 2474842. DOI: 10.1186/1756-9966-27-15. View

10.

Yamazaki T, Kanda T, Sakai Y, Hatakeyama K . Liver fatty acid-binding protein is a new prognostic factor for hepatic resection of colorectal cancer metastases. J Surg Oncol. 1999; 72(2):83-7. DOI: 10.1002/(sici)1096-9098(199910)72:2<83::aid-jso8>3.0.co;2-c. View

11.

Ibusuki M, Fu P, Yamamoto S, Fujiwara S, Yamamoto Y, Honda Y . Establishment of a standardized gene-expression analysis system using formalin-fixed, paraffin-embedded, breast cancer specimens. Breast Cancer. 2011; 20(2):159-66. DOI: 10.1007/s12282-011-0318-x. View

12.

Torre L, Bray F, Siegel R, Ferlay J, Lortet-Tieulent J, Jemal A . Global cancer statistics, 2012. CA Cancer J Clin. 2015; 65(2):87-108. DOI: 10.3322/caac.21262. View

13.

Sharaf R, Butte A, Montgomery K, Pai R, Dudley J, Pasricha P . Computational prediction and experimental validation associating FABP-1 and pancreatic adenocarcinoma with diabetes. BMC Gastroenterol. 2011; 11:5. PMC: 3037918. DOI: 10.1186/1471-230X-11-5. View

14.

Kheirelseid E, Chang K, Newell J, Kerin M, Miller N . Identification of endogenous control genes for normalisation of real-time quantitative PCR data in colorectal cancer. BMC Mol Biol. 2010; 11:12. PMC: 2825202. DOI: 10.1186/1471-2199-11-12. View

15.

Kong H, Zhu M, Cui F, Wang S, Gao X, Lu S . Quantitative assessment of short amplicons in FFPE-derived long-chain RNA. Sci Rep. 2014; 4:7246. PMC: 5384205. DOI: 10.1038/srep07246. View

16.

Hashimoto T, Kusakabe T, Watanabe K, Sugino T, Fukuda T, Nashimoto A . Liver-type fatty acid-binding protein is highly expressed in intestinal metaplasia and in a subset of carcinomas of the stomach without association with the fatty acid synthase status in the carcinoma. Pathobiology. 2004; 71(3):115-22. DOI: 10.1159/000076465. View

17.

Antalis T, Reeder J, Gotley D, Byeon M, Walsh M, Henderson K . Down-regulation of the down-regulated in adenoma (DRA) gene correlates with colon tumor progression. Clin Cancer Res. 1998; 4(8):1857-63. View

18.

Kondo T . Inconvenient truth: cancer biomarker development by using proteomics. Biochim Biophys Acta. 2013; 1844(5):861-5. DOI: 10.1016/j.bbapap.2013.07.009. View

19.

Di Meo A, Diamandis E, Rodriguez H, Hoofnagle A, Ioannidis J, Lopez M . What is wrong with clinical proteomics?. Clin Chem. 2014; 60(10):1258-66. DOI: 10.1373/clinchem.2014.225185. View

20.

Wang B, Tao X, Huang C, Liu J, Ye Y, Huang A . Decreased expression of liver-type fatty acid-binding protein is associated with poor prognosis in hepatocellular carcinoma. Hepatogastroenterology. 2014; 61(133):1321-6. View