Exosomal HIF1α supports invasive potential of nasopharyngeal carcinoma-associated LMP1-positive exosomes

13301甲第4339号博士（医学）金沢大学博士論文本文Full 以下に掲載：oncogene 33(37) pp.4613-4622 2014. nature publishing group. 共著者：Aga M, Bentz GL, Raffa S, Torrisi MR, Kondo S, Wakisaka N, Yoshizaki T, Pagano JS, Shackelford J

上咽頭癌のリンパ球浸潤におけるヒト内在性レトロウイルスの関与

上咽頭癌組織と細胞株でHERVの発現を認めたため、HERVがRE-1 likeモチーフを多く含むことから転写抑制因子であるRE1-silencing transcription factor (REST/NRSF)の発現に着目して研究を行った。RESTを強制発現させると癌幹細胞の表現型で見られるようにCD24の発現が抑制され、さらにマトリゲルを通過する細胞の数が増えた。以上のことから、RESTは転写抑制因子として上咽頭癌細胞のCICに関わっており、上咽頭癌の浸潤能に寄与することが示唆された。In the present study we showed a panel of LMP1 expressing cell lines had high expression of REST compared with not. The enhancement of REST by LMP1 was regulated by activation of REST promoter, not blockade of ubiquitin-proteasome pathway. Furthermore, we demonstrated that REST works as transcriptional suppressor of CSC marker in NPC cells. Moreover, it enhances invasive potential of NPC cells. The series of findings indicate that REST functions as transcriptional repressor and involved with invasive potential of NPC expressing EBV oncogene.研究課題/領域番号:16K20235, 研究期間(年度):2016-04-01 - 2019-03-3

Upregulation of special AT-rich-binding protein 1 by Epstein–Barr virus latent membrane protein 1 in human nasopharyngeal cells and nasopharyngeal cancer

A global regulator of chromatin remodelling and gene expression, special AT-rich-binding protein 1 (SATB1) has been implicated in promotion of growth and metastasis of a number of cancers. Here, we demonstrate that the principal oncogene of Epstein–Barr virus (EBV), latent membrane protein 1 (LMP1) upregulates SATB1 RNA and protein expression in human nasopharyngeal cell lines. Silencing of endogenously expressed SATB1 with specific short hairpin RNA decreases cell proliferation and resistance to apoptosis induced by growth factor withdrawal. Additionally, we provide evidence that LMP1-mediated expression of Survivin, a multifunctional protein involved in promoting cell growth and survival, is mediated at least in part by SATB1 in human nasopharyngeal cells. Finally, we show that SATB1 protein levels are elevated in tissue samples from patients with nasopharyngeal carcinoma (NPC), and are directly correlated with the expression of LMP1. Taken together, our results suggest that SATB1 functions as a pro-metastatic effector of LMP1 signalling in EBV-positive NPC

Inhibition of UCH-L1 Deubiquitinating Activity with Two Forms of LDN-57444 Has Anti-Invasive Effects in Metastatic Carcinoma Cells

Normally ubiquitin C-terminal hydrolase L1 (UCH-L1) is expressed in the central nervous and reproductive systems of adults, but its de novo expression has been detected in many human cancers. There is a growing body of evidence that UCH-L1 de-ubiquitinating (DUB) activity plays a major pro-metastatic role in certain carcinomas. Here we tested anti-metastatic effects of the small-molecule inhibitor of UCH-L1 DUB activity, LDN-57444, in cell lines from advanced oral squamous cell carcinoma (OSCC) as well as invasive nasopharyngeal (NP) cell lines expressing the major pro-metastatic gene product of Epstein–Barr virus (EBV) tumor virus, LMP1. To overcome the limited aqueous solubility of LDN-57444 we developed a nanoparticle formulation of LDN-57444 by incorporation of the compound in polyoxazoline micellear nanoparticles (LDN-POx). LDN-POx nanoparticles were equal in effects as the native compound in vitro. Our results demonstrate that inhibition of UCH-L1 DUB activity with LDN or LDN-POx inhibits secretion of exosomes and reduces levels of the pro-metastatic factor in exosomal fractions. Both forms of UCH-L1 DUB inhibitor suppress motility of metastatic squamous carcinoma cells as well as nasopharyngeal cells expressing EBV pro-metastatic Latent membrane protein 1 (LMP1) in physiological assays. Moreover, treatment with LDN and LDN-POx resulted in reduced levels of pro-metastatic markers, a decrease of carcinoma cell adhesion, as well as inhibition of extra-cellular vesicle (ECV)-mediated transfer of viral invasive factor LMP1. We suggest that soluble inhibitors of UCH-L1 such as LDN-POx offer potential forms of treatment for invasive carcinomas including EBV-positive malignancies