Tylosis is an inherited disorder characterized by abnormal palmoplantar skin thickening and a highly elevated risk of esophageal squamous cell carcinoma (ESCC). Analyses of tylosis in families have localized the responsible gene locus to a region of chromosome 17q25.1. Frequent loss of heterozygosity (LOH) in 17q25.1 was also observed in the sporadic form of ESCC. A putative tumor suppressor gene for ESCC may exist at this locus. We investigated the expression patterns of genes on 17q25.1 in tumor and corresponding normal tissues from patients with sporadic ESCC using RNA sequence analysis. For candidate genes, quantitative real-time reverse transcription-PCR (qRT-PCR), direct sequence, LOH and methylation analyses were performed using 93 clinical ESCC samples and 10 cell lines. A significant downregulation of ST6GALNAC1 was demonstrated in ESCC tissues compared to its expression in normal tissues by qRT-PCR (n=93, p<0.0001). Frequent LOH (17/27, 62.9%) and hyper‑methylation in ST6GALNAC1 were also observed in all cell lines. Our results indicated that ST6GALNAC1 was downregulated in sporadic ESCC via hyper-methylation and LOH, and it may be a candidate responsible gene for ESCC. Furthermore, recent studies suggest that multiple genes on chromosome 17q25 are involved in ESCC development.

[Background] Esophageal squamous cell carcinoma (ESCC) is one of the most malignant cancers. The molecular mechanism of ESCC development has not been fully elucidated. [Aim] The aim of this study is to identify responsible genes for ESCC development. [Materials and Methods] Firstly, we performed transcriptome sequence (RNA-seq) analysis of surgically resected tumor and corresponding normal tissues form 3 ESCC patients with different tumor locus, histrogical differentiation, and TNM stage.by RNA-seq analysis: (Case 1) upper thoracic esophagus, well differentiated SCC, TNM, Stage I; (Case 2) middle thoracic esophagus, poorly differentiated SCC, T3N0M0, Stage II; and (Case 3) lower thoracic esophagus, moderately differentiated SCC, T3N1M0, Stage III. Next, we validated the expression status of the candidate gene in 64 clinical samples and 10 ESCC cell lines (KYSE150, KYSE270, KYSE410, KYSE450, KYSE510, TE1, TE6, TE8, TE9, and TE10) using quantitative real-time PCR (qRT-PCR). [Results] In RNA-seq analysis, 34 genes showed more than 30-fold decrease in tumor samples compared to normal tissues, and the most downregulated gene was transmembrane protease serine 11B (TMPRSS11B). Amon these 34 genes, other 5 genes of TMPRSS11 family (TMPRSS11A, TMPRSS11BNL, TMPRSS11D, TMPRSS11E, and TMPRSS11F) were included. qRT-PCR analysis revealed that TMPRSS11B expression in ESCC samples were lower than those in normal samples (p &lt; 0.001). Downregulation of TMPRSS11B in tumor samples compared to corresponding normal samples were observed in 51 cases out of 64 (79.7%). Patients with expression levels of TMPRSS11B that were below the median value were assigned to the low expression group (n = 32), whereas those with expression values above the median were assigned to the high expression group (n = 32). Significant difference was not observed between two groups regarding clinicopathological data, including age, gender, TNM stage, and lymphatic/venous invasion. TMPRSS11B expression was not detected in all of ESCC cell lines and noncancerous esophageal epithelial cell line, HET1A. [Conclusion] Our results suggest that downreguration of TMPRSS11B expression occurs in early stage of the ESCC oncogenesis and that TMPRSS11B has tumor suppressive roles in ESCC development. Citation Format: Suburu Amano, Takeshi Iwaya, Satoshi Nishizuka, Kohei Kume, Chie Ito, Yuji Akiyama, Yoshihiro Shioi, Fumitaka Endo, Akira Sasaki. Downregulation of TMPRSS11B in squamous cell carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1159.

[Background] Tylosis is an autosomal dominant skin disorder that is associated with the early onset of esophageal squamous cell carcinoma (ESCC) in several families. The tylosis esophageal cancer (TOC) gene locus has been mapped to chromosome 17q25.1 using linkage analyses. This region is also frequently lost in sporadic ESCC. [Materials and Methods] Gene expression profiles on 17q25 in tumor samples from 3 ESCC patients were analyzed using RNA-seq. We validated the expression status of candidate genes in samples from 90 ESCC patients using qRT-PCR. Direct sequence, microsatellite LOH analysis, and methylation assay were also performed in the candidate gene, ST6GALNAC1. Mutation profiles of genes on 17q were also evaluated by exome sequence analysis from 144 ESCC patients. [Result] EVPL and ST6GALNAC1 were found to be significantly downregulated in tumor samples using RNA-seq. Significant downregulation of ST6GALNAC1 and EVPL expression in cancer tissues was confirmed by qRT-PCR in 90 ESCC samples. Although direct sequence of ST6GALNAC1 demonstrated missense mutations in 7 out of 46 (15%) cases, these mutations were not tumor specific. ST6GALNAC1 expression was significantly upregulated in 5-aza-dC treatment groups compared with control in the 5 ESCC cell lines. LOH in ST6GALNAC1 gene locus were identified 18 of 27 informative cases (67%). On chromosome 17q, exome sequence revealed that recurrent mutations were observed only in ZNF750 (14.5%) on 7q25.3. Recently, missense mutations in RHBDF2 were identified tylosis familys. However, RHBDF2 mutation was not observed in sporadic ESC samples by exome sequence analysis. [Conclusion] Our results suggest that ST6GALNAC1 is a putative tumor suppressor gene for ESCC. Furthermore, recent studies on tylosis families and our results on sporadic ESC suggest that multiple genes on chromosome 17q25 are involved in ESCC development. Citation Format: Takeshi Iwaya, Suburu Amano, Fumitaka Endo, Yuji Akiyama, Yoshihiro Shioi, Kohei Kume, Satoshi Nishizuka, Chie Ito, Akira Sasaki, Koshi Mimori. Multiple genes on chromosome 17q25 were involved in sporadic esophageal squamous cell carcinoma development. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1162.