Abstract 3023: RNA-Seq identifies neuropilin-2 as a novel oncogene in esophageal squamous cell carcinoma.
Authors: Fung Tsun Ming , Tong Man , Kwan Pak Shing , Chan Yuen Piu , Guan Xin-Yuan , Ma Stephanie 
Published: August 2014

Esophageal squamous cell carcinoma (ESCC), the major histologic subtype of esophageal cancer, is one of the most common and deadliest cancers in the world, with survival rates of less than 15%. Like many other solid tumors, the pathogenesis of ESCC is believed to be a multistep process with accumulation of numerous genetic alterations involving de-regulation of both tumor-suppressor genes and oncogenes. An understanding of the molecular events that result in the development and progression of the disease may lead to treatments that will increase survival rates of ESCC patients. Recent advances in sequencing technologies offer the opportunity to characterize the cancer genome at unprecedented depth and sensitivity. RNA-Seq has proved an effective means of precisely quantifying the changing expression levels of each transcript. In this study, we performed integrative RNA-Seq analysis on 3 paired patient-derived non-tumor and ESCC clinical specimens and discovered a number of commonly differentially expressed genes, of which included a significantly de-regulated neuropilin (NRP) family including neuropilin-1 (NRP1) and neuropilin-2 (NRP2). NRP is a single-spanned membrane glycoprotein. Members of the family have previously been shown to play an important role in various cancer types and to be a co-receptor of vascular endothelial growth factors. Yet to date, its role in ESCC has not been explored. Up-regulation of both NRP1 and NRP2 in ESCC was subsequently validated in the original 3 pairs of clinical samples used for RNA-Seq as well as in a larger cohort of samples (n = 40) by qPCR. Given that NRP2 de-regulation in ESCC is more significant, we chose to first focus our study on this member of the family. Immunohistochemical studies likewise found NRP2 to be up-regulated in ESCC as compared with its non-tumor counterparts in a tissue microarray. Consistently, expression studies in a panel of esophageal cell lines found NRP2 to be expressed at a significantly higher level in 7 of the 8 ESCC cell lines examined compared with immortalized normal esophageal cell lines, NE1 and NE3. Functional studies in ESCC cells, KYSE180 and KYSE140, with NRP2 stably repressed by lentiviral-based shRNA approach impaired proliferation and metastatic abilities in vitro, as demonstrated by foci formation and chemotaxis migration assays, respectively. Moreover, conditioned medium collected from NRP2 repressed cells displayed a reduced ability to induce capillary tube formation in HUVEC cells. These results suggest that NRP2 plays an important role in regulating cell proliferation, migration and angiogenesis in ESCC. Additional work on the mechanism by which NRP2 drives ESCC and the effectiveness of the use of a NRP2 neutralizing antibody against ESCC is currently being studied. We believe with further study, NRP2 may provide a prognostic biomarker for ESCC outcome prediction and a novel therapeutic target for ESCC treatment.

Citation Format: Tsun Ming Fung, Man Tong, Pak Shing Kwan, Yuen Piu Chan, Xin-Yuan Guan, Stephanie Ma. RNA-Seq identifies neuropilin-2 as a novel oncogene in esophageal squamous cell carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3023. doi:10.1158/1538-7445.AM2013-3023