Altered expression of BTG1 is a potential biomarker for carcinogenesis and progression of GC, particularly for proximal nondiffuse and diffuse GC.

Background Patients with advanced gastric cancer (GC) have an adverse prognosis even after curative resection. Development of novel diagnostic and therapeutic approaches for GC is urgently required. Methods The expression and methylation status of DENN/MADD domain-containing protein 2D (DEN-ND2D), a member of the membrane trafficking proteins, were evaluated in 12 GC cell lines and 112 pairs of surgical specimens. Subgroup analysis based on tumor differentiation, location, and morphology was also performed. Expression and distribution of DENND2D protein were determined by immunohistochemistry. Results The majority of GC cell lines (75 %) and tissues (79 %) showed reduced expression of DENND2D mRNA compared with noncancerous gastric tissues. GC tissues showed a significantly lower mean expression level of mRNA and a higher frequency of promoter hypermethylation of DENND2D than corresponding noncancerous tissues. No significant differences in DENND2D mRNA expression and methylation status were found between GC subtypes categorized by tumor differentiation, location, and morphology. The expression patterns of DENND2D protein were confirmed to be consistent with those of DENND2D mRNA. Downregulation of DENND2D mRNA in GC tissues was significantly associated with factors related to more advanced GC and subsequent adverse prognosis. Among 72 patients who underwent R0 resection, downregulation of DENND2D mRNA in GC tissues was an independent prognostic factor and associated with early recurrence. Conclusions Our results suggested that DENND2D is a putative tumor suppressor gene regulated by promoter hypermethylation in GC. Downregulation of DENND2D can serve as a novel tumor biomarker to predict progression and early recurrence of all types of GC.

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths worldwide and its prognosis is poor. Novel targets for treating recurrence and progression along with associated biomarkers are urgently required. In this study, the expression and regulatory mechanism of DENN/MADD domain containing 2D (DENND2D) were investigated in an attempt to identify a tumor suppressor gene for HCC regulated by silencing through promoter hypermethylation. The levels of DENND2D expression in HCC cell lines and surgical specimens were determined using a quantitative polymerase chain reaction assay and the relationship between the expression levels of DENND2D mRNA and clinicopathological factors was evaluated. The expression and distribution of DENND2D were determined using immunohistochemistry. DNA methylation analysis was performed to determine the regulatory mechanisms of DENND2D expression in HCC. Most HCC cell lines (89%) and surgical specimens (78%) expressed lower levels of DENND2D mRNA compared with normal liver tissue. In contrast, there was no significant difference in the expression levels of DENND2D mRNA between normal tissues of HCC patients with and without cirrhosis. The expression patterns of DENND2D protein and mRNA were consistent. Patients with significantly lower levels of DENND2D mRNA in HCC tissues had remarkably earlier recurrences after hepatectomy and their prognosis worsened. The DENND2D promoter was methylated in eight out of nine HCC cell lines and DNA demethylation reactivated DENND2D mRNA expression. Hypermethylation of DENND2D was frequently detected in HCC tissues (75%) and was significantly associated with downregulation of DENND2D mRNA expression. DENND2D is a candidate tumor suppressor gene that is inactivated by promoter hypermethylation in patients with HCC and may serve as a novel biomarker of early recurrence of HCC.

Although the B‑cell translocation gene 1 (BTG1) plays an important role in apoptosis and negatively regulates cell proliferation, BTG1 expression in hepatocellular carcinoma (HCC) has not been evaluated. In this study expression analysis of BTG1 was conducted to clarify the role of BTG1 in the initiation of HCC carcinogenesis and progression. BTG1 mRNA expression levels were determined for HCC cell lines and 151 surgical specimen pairs using quantitative real‑time reverse transcription polymerase chain reaction (RT‑qPCR) assay. The mutational and methylation status of HCC cell lines were analyzed via high resolution melting (HRM) analysis and direct sequencing analysis to elucidate the regulatory mechanisms of BTG1 expression. The expression and distribution of the BTG1 protein in liver tissues were evaluated using immunohistochemistry (IHC). Decreased expression of BTG1 mRNA was confirmed in the majority of HCC cell lines (89%) and clinical HCC tissues (85%) compared with non‑cancerous liver tissues. Mutations or promoter hypermethylation were not identified in HCC cell lines. BTG1 mRNA expression levels were not influenced by background liver status. The pattern of BTG1 protein expression was consistent with that of BTG1 mRNA. Downregulation of BTG1 mRNA in HCC was significantly associated with shorter disease‑specific and recurrence‑free survival rates. Multivariate analysis of disease‑specific survival rates identified BTG1 mRNA downregulation as an independent prognostic factor for HCC (hazard ratio 2.12, 95% confidence interval 1.12‑4.04, P=0.022). Our results indicate that altered BTG1 expression might affect hepatocarcinogenesis and may represent a novel biomarker for HCC carcinogenesis and progression.

MAGE-D2 was associated with metastatic potential of GC and may represent a promising biomarker, both in gastric tissues and serum samples, for malignant behavior of GC.