Epigenetic silencing and tumor suppressor gene of HAND2 by targeting ERK signaling in colorectal cancer

Background: The screening biomarkers for early detection of colorectal cancer (CRC) is lacking. The aim is to identify epigenetic silenced genes and clarify its roles and underlying mechanism in CRC. We conducted integrative analyses of epigenome-wide Human Methylation 450 K arrays and transcriptome to screen out candidate epigenetic driver genes with transcription silencing. Methylated silencing HAND2 were identified and verified in large CRC cohort. The mechanism of HAND2 expression by promoter inhibition were clarified both in vitro and vivo assays. Cell biofunctional roles of HAND2 methylation was investigated in CRC cells. HAND2 reconstitution were constructed by lentivirus plasmid and tumor xenograft model of HAND2 were built subcutaneously. Genomic mRNA analysis by RNA-sequencing and subsequent GSEA analysis were performed to identify potential target of HAND2 and qPCR/WB was conducted to identify the results. Results: We firstly reported high frequency of HAND2 methylation in promoter in CRC and hypermethylation was negatively correlated with expression silencing and leaded to poor survival in several CRC cohort patients. 5-Aza treatment to demethylated HAND2 could revert its expression in CRC cells. Functionally, HAND2 reconstitution can inhibit cell proliferation, invasion and migration in vitro. In tumor xenograft, HAND2 reconstruction significantly repressed tumor growth when compared to control vector. Thousands of aberrant expressed genes were observed in the heatmap of RNA-sequencing data. HAND2 reconstitution could bind to ERK and reduce its phosphorylation by CoIP assay. These above results showed HAND2 reconstitution perturbed the activation of MAPK/ERK signaling by reduction of ERK phosphorylation. Conclusions: HAND2 is one tumor suppressor by targeting ERK signaling and one potential epigenetic driver gene in CRC.

Automated summary

This study identified HAND2 as a potential epigenetic driver gene in colorectal cancer (CRC). The researchers found a high frequency of HAND2 methylation in CRC patients, and this methylation was negatively correlated with gene silencing and poor survival. Demethylation treatment was able to restore HAND2 expression in CRC cells. Functional experiments showed that HAND2 reconstitution could inhibit cell proliferation, invasion, and migration in vitro. In a tumor xenograft model, HAND2 reconstruction significantly suppressed tumor growth. Furthermore, HAND2 was found to target the ERK signaling pathway and reduce ERK phosphorylation. Overall, this study suggests that HAND2 is a tumor suppressor gene that acts by targeting the ERK signaling pathway and has potential as an epigenetic driver gene in CRC.