Section of Experimental Oncology, Cancer Research UK Centre, Leeds Institute of Molecular Medicine, St James's University Hospital, Beckett Street, Leeds LS97TF, UK.
Br J Cancer. 2013 Apr 2;108(6):1368-77. doi: 10.1038/bjc.2013.104.
Necdin (NDN) expression is downregulated in telomerase-immortalised normal human urothelial cells. Telomerase-immortalised normal human urothelial cells have no detected genetic alterations. Accordingly, many of the genes whose expression is altered following immortalisation are those for which epigenetic silencing is reported.
NDN expression was examined in normal tissues and tumour cell lines by quantitative real-time PCR and immunoblotting. Immunohistochemistry was performed on urothelial carcinoma (UC). Urothelial carcinoma and UC cell lines were subject to HumanMethylation27 BeadChip Array-based methylation analyses. Mutation screening was performed. The functional significance of NDN expression was investigated using retroviral-mediated downregulation or overexpression.
NDN protein was widely expressed in normal tissues. Loss of expression was observed in 38 out of 44 (86%) of UC cell lines and 19 out of 25 (76%) of non-UC cell lines. Loss of NDN protein was found in the majority of primary UC. Oncomine analysis demonstrated downregulation of expression in multiple tumour types. In UC, tumour-specific hypermethylation of NDN and key CpG sites where hypermethylation correlated with reduced expression were identified. Six novel mutations, including some of predicted functional significance, were identified in colorectal and ovarian cancer cell lines. Functional studies showed that NDN could suppress colony formation at low cell density and affect anchorage-independent growth and anoikis in vitro.
NDN is a novel tumour suppressor candidate that is downregulated and hypermethylated or mutated in cancer.
端粒酶永生化的正常人体尿路上皮细胞中 Necdin（NDN）的表达下调。端粒酶永生化的正常人体尿路上皮细胞未检测到遗传改变。因此，许多在永生化后表达发生改变的基因是那些报道了表观遗传沉默的基因。
通过定量实时 PCR 和免疫印迹法检查正常组织和肿瘤细胞系中的 NDN 表达。对尿路上皮癌（UC）进行免疫组织化学染色。对尿路上皮癌和 UC 细胞系进行 HumanMethylation27 BeadChip 芯片阵列甲基化分析。进行突变筛选。使用逆转录病毒介导的下调或过表达研究 NDN 表达的功能意义。
NDN 蛋白在正常组织中广泛表达。在 44 个 UC 细胞系中的 38 个（86%）和 25 个非 UC 细胞系中的 19 个（76%）中观察到表达缺失。在大多数原发性 UC 中发现 NDN 蛋白丢失。Oncomine 分析表明在多种肿瘤类型中表达下调。在 UC 中，鉴定了 NDN 及其关键 CpG 位点的肿瘤特异性高甲基化，其中高甲基化与表达降低相关。在结直肠癌和卵巢癌细胞系中发现了六个新突变，包括一些具有预测功能意义的突变。功能研究表明，NDN 可以抑制低细胞密度下的集落形成，并影响体外的锚定非依赖性生长和失巢凋亡。
NDN 是一种新的肿瘤抑制候选基因，在癌症中下调、高甲基化或突变。
