The Preclinical Study Of Long Noncoding RNA HOTAIR Drives The Malignancy Of Esophageal Carcinoma And Acts As A Potential Prognosis Marker

BACKGROUND:HOX transcript antisense RNA (HOTAIR), which is expressed from the homebox C gene (HOXC) locus, is capable of reprogramming chromatin organization and promoting cancer cell metastasis and can simultaneously bind the Polycomb Repressive Complex 2 (PRC2), which enhances H3K27 trimethylation, and the LSDl-CoREST-REST complex, which is critical for H3K4 demethylation. Clinically, the overexpression of HOTAIR is a powerful predictor of the tumor progression and overall survival in patients with diverse cancers. The relationship between HOTAIR and esophageal squamous cell carcinoma (ESCC), however, remains unclear. We investigated the role of HOTAIR in the pathogenesis of ESCC.METHODS:We used quantitative real-time PCR (qRT-PCR) to determine the level of HOTAIR in ESCC cell lines and 100 ESCC samples from patients; 56 adjacent non-neoplastic tissues were used as controls. We measured the effect of HOTAIR knockdown and overexpression in ESCC cell lines using colony formation assays, anchorage-independent growth assays, the CCK-8 assay, transwell migration and invasion assays, and Annexin Ⅴ-binding assays. We analyzed the growth of ESCC xenograft tumors in nude mice. Changes in the gene expression and methylation levels in ESCC cell lines were analyzed using gene expression microarrays and the Infinium HumanMethylation450K BeadChip assay, respectively.RESULTS:The levels of HOTAIR were increased in ESCC cell lines and patient samples compared with the controls; the expression levels correlated with the disease stage and survival time. The knockdown of HOTAIR in the KYSE510 and KYSE180 ESCC cell lines using shRNAs reduced the ability of the cells to form foci, migrate, and invade the extracellular matrix in culture, altered cell cycle progression, and increased the sensitivity of the cells to apoptosis. The HOTAIR knockdown reduced cancer cell metastasis in vivo, and the tumors formed by HOTAIR-silenced ESCC cells were smaller, both in size and weight, than the tumors and metastases formed by the shRNA-vector control cells in a mouse xenograft model. The results of the gene microarray study showed that HOTAIR reprogrammed the gene expression profile of ESCC cells, and the Gene Ontology (GO) analysis revealed an enrichment in genes that are important for tumorigenesis, such as genes involved in cell migration and the regulation of the cell cycle. Comparing the gene expression profiles and DNA methylation analysis between the KYSE180 and KYSE180_HOTAIR cells revealed that only a small proportion of the methylation changes were correlated with gene expression changes.CONCLUSIONS:HOTAIR is upregulated in ESCC cell lines and patient samples and promotes ESCC cell proliferation and tumor metastasis in mice. The knockdown of HOTAIR resulted in significant changes in gene expression, and data analysis suggested that HOTAIR-mediated gene regulation plays a critical role in ESCC progression and is a novel epigenetic molecular target for treating ESCC patients.