Methylation of the calcium channel-related gene, CACNA2D3, is frequent and a poor prognostic factor in gastric cancer

Abstract

Background & aims: The calcium channel voltage-dependent alpha2delta subunit consists of 4 genes, CACNA2D1 to CACNA2D4, of which CACNA2D2 and CACNA2D3 are located on 3p21.3 and 3p21.1, respectively. Here, we examined the relation between alpha2delta subunit gene alterations and gastric carcinogenesis.

Methods: The expression and methylation status of the alpha2delta subunit genes were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and methylation-specific PCR in gastric cancers (GCs). The effects of CACNA2D3 expression were examined by cell proliferation and adhesion assays, and they predicted target gene alterations.

Results: Aberrant methylation of CACNA2D1 and CACNA2D3 mostly corresponded to their expression status in GC cell lines. CACNA2D1/3 methylation was detected in 10 (12.5%) and 24 (30%) of the 80 GC cases, respectively, but no CACNA2D2 methylation was seen in 32 cases. CACNA2D3 methylation was more frequently found in diffuse type than in intestinal type (16/38 [42.1%] vs 8/42 [19.0%]; P = .025) GCs. Among the 53 patients with advanced GCs, patients with cancers showing CACNA2D3 methylation had a significantly shorter survival time than patients without this methylation (P = .003). Exogenous CACNA2D3 expression strongly inhibited cell growth and adhesion and up-regulated p21 and p27 expression in HEK-293T and NUGC4 cells. Inverse effects were seen by CACNA2D3 small interfering RNA treatment in the CACNA2D3-positive cell lines, indicating that CACNA2D3 may have tumor suppressive functions.

Conclusions: Loss of CACNA2D3 expression through aberrant promoter hypermethylation may contribute to gastric carcinogenesis, and CACNA2D3 methylation is a useful prognostic marker for patients with advanced GC.