Anoctamin 5 regulates the cell cycle and affects prognosis in gastric cancer

Background: Anoctamin 5 (ANO5)/transmembrane protein 16E belongs to the ANO/ transmembrane protein 16 anion channel family. ANOs comprise a family of plasma membrane proteins that mediate ion transport and phospholipid scrambling and regulate other membrane proteins in numerous cell types. Previous studies have elucidated the roles and mechanisms of ANO5 activation in various cancer types. However, it remains unclear whether ANO5 acts as a plasma membrane chloride channel, and its expression and functions in gastric cancer (GC) have not been investigated.

Aim: To examine the role of ANO5 in the regulation of tumor progression and clinicopathological significance of its expression in GC.

Methods: Knockdown experiments using ANO5 small interfering RNA were conducted in human GC cell lines, and changes in cell proliferation, cell cycle progression, apoptosis, and cellular movement were assessed. The gene expression profiles of GC cells were investigated following ANO5 silencing by microarray analysis. Immunohistochemical staining of ANO5 was performed on 195 primary tumor samples obtained from patients with GC who underwent curative gastrectomy between 2011 and 2013 at our department.

Results: Reverse transcription-quantitative polymerase chain reaction (PCR) and western blotting demonstrated high ANO5 mRNA and protein expression, respectively, in NUGC4 and MKN45 cells. In these cells, ANO5 silencing inhibited cell proliferation and induced apoptosis. In addition, the knockdown of ANO5 inhibited G₁-S phase progression, invasion, and migration. The results of the microarray analysis revealed changes in the expression levels of several cyclin-associated genes, such as CDKN1A, CDK2/4/6, CCNE2, and E2F1, in ANO5-depleted NUGC4 cells. The expression of these genes was verified using reverse transcription-quantitative PCR. Immunohistochemical staining revealed that high ANO5 expression levels were associated with a poor prognosis. Multivariate analysis identified high ANO5 expression as an independent prognostic factor for 5-year survival in patients with GC (P = 0.0457).

Conclusion: ANO5 regulates the cell cycle progression by regulating the expression of cyclin-associated genes and affects the prognosis of patients with GC. These results may provide insights into the role of ANO5 as a key mediator in tumor progression and/or promising prognostic biomarker for GC.