Inactivation of p16, RUNX3, and HPP1 occurs early in Barrett's-associated neoplastic progression and predicts progression risk
Inactivation of p16, RUNX3, and HPP1 occurs early in Barrett's-associated neoplastic progression and predicts progression risk
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DOI：
10.1038/sj.onc.1208598
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发表时间：
2005-06-09
期刊：
影响因子：
8
通讯作者：
Meltzer, SJ
中科院分区：
文献类型：
--
作者：
Schulmann, K;Sterian, A;Meltzer, SJ
Barrett食管（BE）患者发生食管腺癌（EAC）的风险增加。临床肿瘤进展的风险因素，如年龄和食管BE段的长度，已被确定。然而，需要改进预测进展风险增加的分子生物标志物以改进风险评估和分层。阳离子采用实时定量甲基化特异性PCR技术，对77例EAC、93例BE和64例正常食管（NE）标本中10个基因（HPP 1、RUNX 3、RIZ 1、CRBP 1、3-OST-2、APC、TIMP 3、p16、MGMT、p14）的启动子区进行甲基化检测。然后在20个发育异常标本中分析了BE和EAC之间甲基化频率显着差异的基因子集。除p14外，所有10个基因在EAC中均频繁甲基化，其中RUNX 3、HPP 1、CRBP 1、RIZ 1和OST-2代表EAC和/或BE中的新甲基化靶点。与EAC相比，BE中p16、RUNX 3和HPP 1的甲基化频率增加。此外，这些甲基化的增加发生在早期，在BE和低度异型增生（LGD）之间的界面。为了证明超甲基化的沉默效应，我们选择了EAC细胞BIC 1，其中HPP 1启动子天然甲基化，并使它们经受5-氮杂-2 '-脱氧胞苷（Aza-C）处理。实时RT-PCR结果显示，Aza-C处理3天后，HPP 1 mRNA水平增加，甲基化HPP 1 DNA水平降低。然后在一项回顾性纵向研究中检测了6个基因（APC、TIMP 3、CRBP 1、p16、RUNX 3和HPP 1）亚组的超甲基化，该研究包括从53例接受监测内镜检查的BE患者中获得的99份BE和9份LGD标本。只有高度异型增生（HGD）或EAC被定义为进展终点。采用考克斯比例风险模型比较两组患者：8例进展者（P）和45例非进展者（NP），以确定年龄、BE片段长度和甲基化事件的相对进展风险。多变量分析显示，只有p16基因的高甲基化（比值比（OR）1.74，95%置信区间（CI）1.33 - 2.20），RUNX 3（OR 1.80，95%CI 1.08 - 2.81）和HPP 1（OR 1.77，95%CI 1.06 - 2.81）与疾病进展风险增加独立相关，而年龄、BE段长度和TIMP 3、APC或CRBP 1的高甲基化不是独立风险因素。在综合分析中，风险可检测到，但不早于肿瘤进展前2年。BE或LGD中p16、RUNX 3和HPP 1的高甲基化可能是BE进展为HGD或EAC的独立风险因素。这些发现对风险分层有影响。阳离子，早期EAC检测，以及适当的内镜监测时间间隔。
Patients with Barrett's esophagus ( BE) are at increased risk of developing esophageal adenocarcinoma (EAC). Clinical neoplastic progression risk factors, such as age and the length of the esophageal BE segment, have been identified. However, improved molecular biomarkers predicting increased progression risk are needed for improved risk assessment and strati. cation. Using real-time quantitative methylation-specific PCR, we screened 10 genes (HPP1, RUNX3, RIZ1, CRBP1, 3-OST-2, APC, TIMP3, p16, MGMT, p14) for promoter hypermethylation in 77 EAC, 93 BE, and 64 normal esophagus ( NE) specimens. A subset of genes manifesting significant differences in methylation frequencies between BE and EAC was then analysed in 20 dysplastic specimens. All 10 genes except p14 were frequently methylated in EACs, with RUNX3, HPP1, CRBP1, RIZ1, and OST-2 representing novel methylation targets in EAC and/or BE. p16, RUNX3, and HPP1 displayed increasing methylation frequencies in BE vs EAC. Furthermore, these increases in methylation occurred early, at the interface between BE and low-grade dysplasia (LGD). To demonstrate the silencing effect of hypermethylation, we selected the EAC cells BIC1, in which the HPP1 promoter is natively methylated, and subjected them to 5-aza-2'-deoxycytidine (Aza-C) treatment. Real-time RT-PCR indicated increased HPP1 mRNA levels after 3 days of Aza-C treatment, as well as decreased levels of methylated HPP1 DNA. Hypermethylation of a subset of six genes ( APC, TIMP3, CRBP1, p16, RUNX3, and HPP1) was then tested in a retrospective longitudinal study of 99 BE and nine LGD specimens obtained from 53 BE patients undergoing surveillance endoscopy. Only high-grade dysplasia (HGD) or EACwere defined as progression end points. Two patient groups were compared: eight progressors ( P) and 45 nonprogressors (NP), using Cox proportional hazards models to determine the relative progression risks of age, BE segment length, and methylation events. Multivariate analyses revealed that only hypermethylation of p16 ( odds ratio ( OR) 1.74, 95% confidence interval (CI) 1.33 - 2.20), RUNX3 ( OR 1.80, 95% CI 1.08 - 2.81), and HPP1 ( OR 1.77, 95% CI 1.06 - 2.81) were independently associated with an increased risk of progression, whereas age, BE segment length, and hypermethylation of TIMP3, APC, or CRBP1 were not independent risk factors. In combined analyses, risk was detectable up to, but not earlier than, 2 years preceding neoplastic progression. Hypermethylation of p16, RUNX3, and HPP1 in BE or LGD may represent independent risk factors for the progression of BE to HGD or EAC. These findings have implications regarding risk strati. cation, early EAC detection, and the appropriate endoscopic surveillance interval for patients with BE.