Comparative Proteomics of Ovarian Cancer Aggregate Formation Reveals an Increased Expression of Calcium-activated Chloride Channel Regulator 1 (CLCA1)

• Published in: The Journal of biological chemistry 2015-07, Vol.290 (28), p.17218-17227
• Main Authors: Musrap, Natasha, Tuccitto, Alessandra, Karagiannis, George S., Saraon, Punit, Batruch, Ihor, Diamandis, Eleftherios P.
• Format: Article
• Language: English
• Subjects: aggregation; Cell Adhesion; Cell Aggregation - genetics; Cell Aggregation - physiology; Cell Line, Tumor; Chloride Channels - antagonists & inhibitors; Chloride Channels - genetics; Chloride Channels - metabolism; CLCA1; Disease Progression; Female; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Genomics and Proteomics; Humans; mass spectrometry (MS); metastasis; Models, Biological; multicellular aggregates; Niflumic Acid - pharmacology; ovarian cancer; Ovarian Neoplasms - genetics; Ovarian Neoplasms - metabolism; Ovarian Neoplasms - pathology; Proteomics; RNA, Messenger - genetics; RNA, Messenger - metabolism; RNA, Neoplasm - genetics; RNA, Neoplasm - metabolism; RNA, Small Interfering - genetics; spheroids; Spheroids, Cellular - metabolism; Spheroids, Cellular - pathology; Tandem Mass Spectrometry
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M115.639773

Ovarian cancer is a lethal gynecological disease that is characterized by peritoneal metastasis and increased resistance to conventional chemotherapies. This increased resistance and the ability to spread is often attributed to the formation of multicellular aggregates or spheroids in the peritoneal cavity, which seed abdominal surfaces and organs. Given that the presence of metastatic implants is a predictor of poor survival, a better understanding of how spheroids form is critical to improving patient outcome, and may result in the identification of novel therapeutic targets. Thus, we attempted to gain insight into the proteomic changes that occur during anchorage-independent cancer cell aggregation. As such, an ovarian cancer cell line, OV-90, was cultured in adherent and non-adherent conditions using stable isotope labeling with amino acids in cell culture (SILAC). Anchorage-dependent cells (OV-90AD) were grown in tissue culture flasks, whereas anchorage-independent cells (OV-90AI) were grown in suspension using the hanging-drop method. Cellular proteins from both conditions were then identified using LC-MS/MS, which resulted in the quantification of 1533 proteins. Of these, 13 and 6 proteins were up-regulated and down-regulated, respectively, in aggregate-forming cells compared with cells grown as monolayers. Relative gene expression and protein expression of candidates were examined in other cell line models of aggregate formation (TOV-112D and ES-2), which revealed an increased expression of calcium-activated chloride channel regulator 1 (CLCA1). Moreover, inhibitor and siRNA transfection studies demonstrated an apparent effect of CLCA1 on cancer cell aggregation. Further elucidation of the role of CLCA1 in the pathogenesis of ovarian cancer is warranted. Background: Cancer spheroid formation is a hallmark during ovarian cancer progression. Results: Quantitative proteomics were used to delineate proteomic alterations during aggregate formation. Conclusion: CLCA1 is elevated in ovarian cancer cell line aggregates. Significance: CLCA1 and chloride channels may serve as novel therapeutic targets for ovarian cancer treatment.