肿瘤抑制基因DNA甲基化与肝癌

DNA甲基化是最早发现的基因表观修饰方式之一.肿瘤抑制基因(tumor suppressor genes,TSGs)DNA甲基化是肝癌的常发事件,该文从转录调节因子、负调控转录因子、细胞周期调控因子、信号通路的调节因子、DNA损伤修复因子等方面综述了TSGs DNA甲基化状态在肝癌发生、发展中的作用,对肝癌早期诊断、分期、预后、个体化治疗具有重要意义.     

GPX3 DNA甲基化在恶性肿瘤发生发展中作用的研究进展

近年来,肿瘤表观遗传学的改变已成为癌症治疗和预防极具吸引力的靶点,其最常见和最广泛的表观遗传途径是DNA甲基化.研究发现,谷胱甘肽过氧化物酶3(glutathione peroxidase 3,GPX3)作为一种硒蛋白在体内具有抗病毒和防治多种恶性肿瘤的作用,GPX3的mRNA和(或)蛋白表达下调甚至沉默与GPX3基因...     

DNA甲基化在肺癌中的研究进展

表观遗传学主要是研究基因表达的变化,而这种变化能够通过减数分裂遗传,但不涉及有关基因碱基序列的改变.随着肿瘤发病机制研究在分子生物学方面的逐步深入,发现DNA甲基化、非编码RNA表达调控和组蛋白的修饰等表观遗传学(epigenetics)改变引起的调控机制异常与人体多种恶性肿瘤的发生发展过程有着密切联系.研究表明抑癌基因启动子区高度DNA甲基化状态在恶性肿瘤的发生发展和侵袭转移中发挥着重要作用.在肺癌发生的早期,出现很多DNA甲基化状态的改变.因此,甲基化检测可能对肺癌患者的早期诊断、治疗及预后评估具有重要意义.     

Spatial concordance of DNA methylation classification in diffuse glioma

BackgroundIntratumoral heterogeneity is a hallmark of diffuse gliomas. DNA methylation profiling is an emerging approach in the clinical classification of brain tumors. The goal of this study is to investigate the effects of intratumoral heterogeneity on classification confidence.MethodsWe used neuronavigation to acquire 133 image-guided and spatially separated stereotactic biopsy samples from 16 adult patients with a diffuse glioma (7 IDH-wildtype and 2 IDH-mutant glioblastoma, 6 diffuse astrocytoma, IDH-mutant and 1 oligodendroglioma, IDH-mutant and 1p19q codeleted), which we characterized using DNA methylation arrays. Samples were obtained from regions with and without abnormalities on contrast-enhanced T1-weighted and fluid-attenuated inversion recovery MRI. Methylation profiles were analyzed to devise a 3-dimensional reconstruction of (epi)genetic heterogeneity. Tumor purity was assessed from clonal methylation sites.ResultsMolecular aberrations indicated that tumor was found outside imaging abnormalities, underlining the infiltrative nature of this tumor and the limitations of current routine imaging modalities. We demonstrate that tumor purity is highly variable between samples and explains a substantial part of apparent epigenetic spatial heterogeneity. We observed that DNA methylation subtypes are often, but not always, conserved in space taking tumor purity and prediction accuracy into account.ConclusionOur results underscore the infiltrative nature of diffuse gliomas and suggest that DNA methylation subtypes are relatively concordant in this tumor type, although some heterogeneity exists.     

Hybrid Capture–Based Genomic Profiling of Circulating Tumor DNA from Patients with Advanced Non–Small Cell Lung Cancer

Introduction

Genomic profiling informs selection of matched targeted therapies as part of routine clinical care in NSCLC. Tissue biopsy is the criterion standard; however, genomic profiling of blood-derived circulating tumor DNA (ctDNA) has emerged as a minimally invasive alternative.

Epigenetic silencing of multiple genes in primary CNS lymphoma

Epigenetic silencing of functionally important genes is important in the development of malignancies and is a source of potential markers for molecular detection. Primary central nervous system lymphoma (PCNSL) is an increasingly common tumor that has not been extensively examined for changes in promoter region methylation. We examined 14 tumor suppressor genes in 25 cases of PCNSL using methylation-specific PCR. Methylation was observed in DAPK (84%), TSP1 (68%), CRBP1 (67%), p16(INK) (4a) (64%), p14(ARF) (59%), MGMT (52%), RARbeta2 (50%), TIMP3 (44%), TIMP2 (42%), p15(INK) (4b) (40%), p73 (28%), hMLH1 (12%), RB1 (8%) and GSTP1 (8%). Promoter methylation of p14(ARF), p16(INK) (4a) and MGMT was correlated with loss of expression by immunohistochemical staining. The methylation of many of these genes in PCNSL is similar to that reported in other high-grade B-cell lymphomas. All 25 cases of PCNSL had methylation of at least 2 genes. Methylation of DAPK, p16(INK) (4a) or MGMT was found in 96% of the tumors, suggesting simple marker strategies to detect circulating methylated DNA in serum that might facilitate early tumor detection. Our study provides insight into the epigenetic alterations in PCNSL and provides potential biomarkers of disease.     

Papillary Tumor of the Pineal Region Rare Pediatric CNS Tumor Case Series Treated in King Fahad Medical City (KFMC)

The clinical behaviors, prognosis, and appropriate treatments of papillary tumors of the pineal region (PTPR) are not fully defined due to the rarity of these tumors. At diagnosis, PTPR may present with clinical symptoms, including headache with obstructive hydrocephalus, diplopia, vomiting, and lethargy, as well as neurological signs, including Argyll Robertson pupils and Parinaud’s syndrome due to compression of the dorsal midbrain, specifically the periaqueductal region with horizontal nystagmus. Radiological assessment of pineal region lesions is challenging, with a wide range of potential differential diagnoses. PTPR typically presents as a heterogeneous, well-circumscribed mass in the pineal region, which might contain cystic areas, calcifications, hemorrhages, or protein accumulations. Here, we report three female pediatric patients with PTPR treated in King Fahad Medical City (KFMC) in Saudi Arabia. Histological and immunohistochemical diagnosis was confirmed by analysis of genome-wide DNA methylation profiles. This case series expands on the available reports on the clinical presentations of PTPR and provides important information on the responses to different treatment modalities.     

Genome‐wide DNA methylation profiling shows molecular heterogeneity of anaplastic pleomorphic xanthoastrocytoma

In the revised World Health Organization classification 2016, anaplastic pleomorphic xanthoastrocytoma (PXA) has been newly defined as a variant of the PXA entity. Furthermore, some anaplastic PXA were reported to have extremely poor prognosis which showed a type of pediatric glioblastoma (GBM) molecular profile. Recent integrated molecular classification for primary central nervous system tumors proposed some differences between histological and molecular features. Herein, in a genome‐wide molecular analysis, we show an extreme aggressive anaplastic PXA that resulted in a pediatric GBM molecular profile. A full implementation of the molecular approach is the key to predict prognosis and decide the treatment strategy for anaplastic PXA.     

The presence of a methylation fingerprint of Helicobacter pylori infection in human gastric mucosae

Aberrant DNA methylation is deeply involved in human cancers, but its inducers and targets are still mostly unclear. Helicobacter pylori infection was recently shown to induce aberrant methylation in gastric mucosae, and produce a predisposed field for cancerization. Here, we analyzed the presence of target genes in methylation induction by H. pylori and the mechanism for the gene specificity. Noncancerous gastric mucosae were collected from 4 groups of individuals (with and without a gastric cancer, and with and without current H. pylori infection; N = 11 for each group), and methylation of promoter CpG islands of 48 genes that can be methylated in gastric cancer cell lines was analyzed by methylation‐specific PCR. In total, 26 genes were consistently methylated in individuals with current or past infection by H. pylori, whereas 7 genes were not methylated at all. In addition, 14 genes were randomly or intermediately methylated in individuals with gastric cancers and the remaining 1 gene was methylated in all the cases. The methylation‐susceptible genes had significantly lower mRNA expression levels than the methylation‐resistant genes. H. pylori infection did not induce mRNA and protein expression of DNA methyltransferases; DNMT1, DNMT3A or DNMT3B. Gene specificity was present in the induction of aberrant DNA methylation by H. pylori infection, and low mRNA expression, which could precede methylation, was one of the mechanisms for the gene specificity. These findings open up the possibility that a methylation fingerprint can be used as a novel marker for past exposure to a specific carcinogenic factor. © 2008 Wiley‐Liss, Inc.     

Classification,Pathogenesis and Molecular Pathology of Primary CNS Lymphomas

Classification, pathogenesis and molecular pathology of primary central nervous system lymphomas (PCNSL) pose major clinico-pathologic problems. Application of histopathologic classification schemes developed for nodal lymphomas, i.e. the Kiel Classification and the Working Formulation, is not reliable and clinically not relevant for PCNSL. The more recent REAL Classification will simplify subtyping of PCNSL, but reliability and clinical significance remain to be determined. There are virtually no experimental data on whether PCNSL develop outside of the brain, or whether they arise from polyclonal lymphoproliferations within the brain. Mutations of oncogenes and tumor suppressor genes have been analyzed in only a few tumors, and their type and frequency is essentially unknown. In conclusion, compared with both neuroectodermal brain tumors and nodal lymphomas, and given the increasing incidence and clinical impact of PCNSL, there is a remarkable lack of histopathologic consensus as well as a surprising shortage of pathogenetic and molecular genetic information.     

The expression of hematopoietic progenitor cell antigen CD34 is regulated by DNA methylation in a site‐dependent manner in gastrointestinal stromal tumours

The anatomic site‐dependent expression of hematopoietic progenitor cell antigen CD34 is a feature of gastrointestinal stromal tumours (GISTs). The basis for the differential CD34 expression is only incompletely understood. This study aimed at understanding the regulation of CD34 in GISTs and clarification of its site‐dependent expression. Two sample sets of primary GISTs were interrogated including 52 fresh‐frozen and 134 paraffin‐embedded and formalin‐fixed specimens. DNA methylation analysis was performed by HumanMethylation450 BeadChip array in three cell lines derived from gastric and intestinal GISTs, and differentially methylated CpG sites were established upstream of CD34. The methylation degree was further quantified by pyrosequencing, and inverse correlation with CD34 mRNA and protein abundance was revealed. The gene's expression could be activated upon induction of DNA hypomethylation with 5‐aza‐2′‐deoxycytidine in GIST‐T1 cells. In patient samples, a strong inverse correlation of DNA methylation degree with immunohistochemically evaluated CD34 expression was documented. Both CD34 expression and DNA methylation levels were specific to the tumours' anatomic location and mutation status. A constant decrease in methylation levels was observed ranging from almost 100% hypermethylation in intestinal GISTs from duodenum to hypomethylation in rectum. CD34 was heavily methylated in gastric PDGFRA‐mutant GISTs in comparison to hypomethylated KIT‐mutant counterparts. Next to CD34 hypermethylation, miR‐665 was predicted and experimentally confirmed to target CD34 mRNA in GIST‐T1 cells. Our results suggest that CD34 expression in GISTs may undergo a complex control by DNA methylation and miR‐665. Differential methylation and expression of CD34 in GISTs along the gastrointestinal tract axis and in tumours that harbour different gain‐of‐function mutations suggest the origin from different cell populations in the gastrointestinal tract.     

循环肿瘤DNA的研究进展

循环肿瘤DNA(circulating tumor DNA,ctDNA)是由肿瘤细胞释放到循环系统中的基因组小片段,其来源于机体所有荷瘤部位,具有含量极低、个体间差异大、半衰期短、匀质性、携带有肿瘤特异性遗传学/表观遗传学变异等特点,与肿瘤的发展进化、休眠耐药、转移复发等密切相关,正逐渐成为肿瘤学分子研究领域的热点.目前针对ctDNA的检测方法和平台种类繁多,主要分为基于PCR和二代测序(next-generation sequence,NGS)的方法,二者各有利弊,需要根据研究目的灵活选择并建立统一标准.随着相关技术的发展和法规的不断完善,ctDNA可作为癌症筛查、早期诊断、个体化治疗及预后评估理想的血源性生物标志物,在肿瘤的精准医疗中必将大放异彩.     

Potential of DNA methylation in rectal cancer as diagnostic and prognostic biomarkers

Background:

Aberrant DNA methylation is more prominent in proximal compared with distal colorectal cancers. Although a number of methylation markers were identified for colon cancer, yet few are available for rectal cancer.

Methods:

DNA methylation differences were assessed by a targeted DNA microarray for 360 marker candidates between 22 fresh frozen rectal tumour samples and 8 controls and validated by microfluidic high-throughput and methylation-sensitive qPCR in fresh frozen and formalin-fixed paraffin-embedded (FFPE) samples, respectively. The CpG island methylator phenotype (CIMP) was assessed by MethyLight in FFPE material from 78 patients with pT2 and pT3 rectal adenocarcinoma.

Results:

We identified and confirmed two novel three-gene signatures in fresh frozen samples that can distinguish tumours from adjacent tissue as well as from blood with a high sensitivity and specificity of up to 1 and an AUC of 1. In addition, methylation of individual CIMP markers was associated with specific clinical parameters such as tumour stage, therapy or patients'' age. Methylation of CDKN2A was a negative prognostic factor for overall survival of patients.

Conclusions:

The newly defined methylation markers will be suitable for early disease detection and monitoring of rectal cancer.     

Expert panel consensus recommendations on the use of circulating tumor DNA assays for patients with advanced solid tumors

Comprehensive genomic profiling is increasingly used to facilitate precision oncology based on molecular stratification. In addition to conventional tissue comprehensive genomic profiling, comprehensive genomic profiling of circulating tumor DNA has become widely utilized in cancer care owing on its advantages, including less invasiveness, rapid turnaround time, and capturing heterogeneity. However, circulating tumor DNA comprehensive genomic profiling has some limitations, mainly false negatives due to low levels of plasma circulating tumor deoxyribonucleic acid and false positives caused by clonal hematopoiesis. Nevertheless, no guidelines and recommendations fully address these issues. Here, an expert panel committee involving representatives from 12 Designated Core Hospitals for Cancer Genomic Medicine in Japan was organized to develop expert consensus recommendations for the use of circulating tumor deoxyribonucleic acid‐based comprehensive genomic profiling. The aim was to generate guidelines for clinicians and allied healthcare professionals on the optimal use of the circulating tumor DNA assays in advanced solid tumors and to aid the design of future clinical trials that utilize and develop circulating tumor DNA assays to refine precision oncology. Fourteen clinical questions regarding circulating tumor deoxyribonucleic acid comprehensive genomic profiling including the timing of testing and considerations for interpreting results were established by searching and curating associated literatures, and corresponding recommendations were prepared based on the literature for each clinical question. Final consensus recommendations were developed by voting to determine the level of each recommendation by the Committee members.     

Clinical validation of methylation biomarkers for optimal detection of high-grade vulvar intraepithelial neoplasia

The precursor lesions of vulvar squamous cell carcinoma (VSCC) include human papillomavirus (HPV)-associated and HPV-independent squamous neoplasia with a varying cancer risk. Our study aimed to validate the accuracy of previously identified DNA methylation markers for detection of such high-grade vulvar intraepithelial neoplasia (VIN). A large clinical series of 751 vulvar lesions, originally diagnosed as high-grade VIN, were reassessed and categorized into HPV-associated or HPV-independent vulvar disease categories. Together with 113 healthy vulvar controls, all samples were tested for 12 methylation markers with quantitative multiplex methylation-specific PCR (qMSP). Performance of individual markers and selection of an optimal marker panel for detection of high-grade VIN was determined by logistic regression analysis. SST was the best-performing individual marker (AUC 0.90), detecting 80% of high-grade VIN cases, with excellent detection of HPV-independent VIN (95%), known to have the highest cancer risk. Merely 2% of controls tested methylation positive for SST. Selection of a marker panel, including ZNF582, SST and miR124-2, resulted in a comparably high accuracy for detection of high-grade VIN (AUC 0.89). In conclusion, we clinically validated the accuracy of 12 DNA methylation markers for detection of high-grade VIN. SST, as a sole marker or in a panel, provides an optimal diagnostic tool to distinguish high-grade VIN in need of treatment, particularly HPV-independent VIN, from low-grade or reactive vulvar lesions. These findings warrant further prognostic validation of methylation biomarkers for cancer risk stratification of patients with VIN.