基于生物信息学技术对结直肠癌相关lncRNA、miRNA和mRNA的筛选及其部分生物学功能的分析

[摘要] 目的：筛选TCGA数据库中结直肠癌（CRC）差异表达的lncRNA、miRNA和mRNA,探讨其与CRC预后的关系及相关生物学功能。方法：从TCGA数据库中下载CRC样本的RNA测序（RNA-Seq）数据及miRNA-Seq 数据并进行分析,利用R程序筛选出差异表达的lncRNA、miRNA和mRNA。通过miRcode、TargetScan 和miRTarbase 3 个数据库对差异表达的RNA之间的关系进行分析整合,构建CRC中的lncRNA-miRNA-mRNA ceRNA网络。用Kaplan-Meier 法分析ceRNA网络中的lncRNA、miRNA和mRNA表达量与患者生存预后的关系,后用GSEA功能富集分析软件分析出参与CRC发生发展的信号通路。结果：共鉴定出CRC中614 个差异表达的lncRNA、244 个差异表达的miRNA、12 672 个差异表达的mRNA;构建了由139 个lncRNA、37 个miRNA和228 个mRNA组成的ceRNA 网络;发现有58 个lncRNA、23 个miRNA、150 个mRNA与CRC的预后相关。GSEA富集分析结果显示,mRNA主要参与Notch、Hedgehog 和TGF-β 等信号通路。结论：成功构建CRC 相关ceRNA 网络,并筛选出与CRC预后相关的lncRNA、miRNA和mRNA,为后续深入开展CRC的临床研究及基础实验研究提供有价值的前期基础。     

乳腺癌竞争内源性RNA网络构建与分析

目的:通过对癌症基因组图谱(The Cancer Genome Atlas，TCGA)数据库中的乳腺癌数据进行综合分析，寻找在乳腺癌中差异表达的长链非编码RNA(long non-coding RNA，lncRNA）、微小RNA（microRNA，miRNA）和信使RNA（messenger RNA，mRNA），构建乳腺癌竞争内源性RNA（competing endogenous RNA，ceRNA）网络，识别和预测ceRNA网络在乳腺癌中的作用。方法:下载TCGA数据库中乳腺癌组织样本基因表达谱数据，寻找差异表达的lncRNA、miRNA和mRNA，通过miRcode、miRTarBase、TargetScan和miRDB四个数据库对差异表达的RNA之间的关系进行分析，构建以上调和下调的miRNA为中心的ceRNA网络。采用Kaplan-Meier法分析乳腺癌ceRNA网络中的lncRNA、miRNA和mRNA表达量与患者生存预后的关系，采用基因富集分析法对乳腺癌ceRNA网络中mRNA基因功能和调控通路进行分析。结果:在乳腺癌中异常表达的350个lncRNA、185个miRNA和2 928个mRNA中，分别有23个lncRNA、27个miRNA、70个mRNA参与构建乳腺癌的ceRNA网络。Kaplan-Meier生存分析显示，lncRNA MAGI2-AS3（P=0.01）、GRIK1-AS1（P=0.03）以及miRNA hsa-miR-301b（P=0.01）、hsa-miR-503（P=0.04）和mRNA CCNE1（P=0.01）、KPNA2（P=0.02）、FLI1（P=0.02）、TGFBR2（P=0.02）这8个基因与乳腺癌患者的生存预后显著相关。70个mRNA主要被富集到20条通路上，其中有15条通路与肿瘤有关，最显著的通路为“Pathways in cancer”。结论:ceRNA网络在乳腺癌中发挥着重要的作用，多种差异表达基因与乳腺癌预后相关，可能成为潜在的肿瘤诊断标志物和治疗靶点。     

喉鳞状细胞癌的转录组数据分析及生物标志物筛选

目的通过数据挖掘分析长链非编码RNA(lncRNA)、微小RNA(miRNA)和信使RNA(mRNA)在喉鳞状细胞癌中的相互作用机制。方法从肿瘤基因图谱(TCGA)中获取喉鳞状细胞癌的lncRNA、miRNA、mRNA表达谱数据及相关临床数据,从TCGA和GEO数据库中获取喉鳞状细胞癌的DNA甲基化数据。进行蛋白相互作用(PPI)、基因文本(GO)、京都基因和基因组百科全书(KEGG)信号通路分析和Kaplan-Meier生存分析,构建竞争性内源RNA(ceRNA)网络和转录因子调控网络。结果LINC00278、LINC00689、MYHAS、miRNA-99a和miRNA-301a属于低风险基因(HR﹤1);MNX1-AS1、LINC02575、HOXB-AS4、LSAMP-AS1、LINC02086、IGFL2-AS1、LINC02253、CASC20、miRNA-383、miRNA-196a-2、miRNA-196a-1、miRNA-100和miRNA-4652属于高风险基因(HR﹥1)。ceRNA网络中有11个lncRNA(LINC02576、LINC02086、AC020659.1、LINC00528、LINC00689、HOXB-AS4、MNX1-AS1、LINC00278、AC010624.1、AC016773.1、MYHAS)、5个miRNA(has-miRNA-206、has-miRNA-573、has-miRNA-3662、has-miRNA-133b、has-miRNA-449a)和12个mRNA(STC2、PAX3、NETO2、EIF5A2、ADPRHL1、SYNM、ACTA1、GPR156、CLIC5、BMP3、HNF4A、FOXL2)。MYHAS的共表达mRNA(cor-mRNA)主要富集在钙信号通路、环磷酸鸟苷(cGMP)-cGMP依赖性蛋白激酶(PKG)信号通路和催产素信号通路等。转录因子网络中有9个转录因子(MYOD、RSRFC4、TAL1BETAITF2、YY1、P300、PAX3、PAX5、ZID和OLF1)。喉鳞状细胞癌DNA甲基化分析得到75个甲基化位点,对应高甲基化基因56个,低甲基化基因15个。结论ceRNA网络并不能完全阐释lncRNA、miRNA和mRNA在喉鳞状细胞癌发生中的作用机制,lncRNA、miRNA和mRNA在喉鳞状细胞癌发生过程中既相互联系又相互独立。筛选出的lncRNA、miRNA和mRNA为试验验证缩小了范围并提供了理论基础。MYHAS在喉鳞状细胞癌中发挥重要作用,是一个潜在的生物学靶点。     

通过竞争性内源RNA调控网络筛查肝细胞癌预后相关分子

目的：通过构建肝细胞癌（hepatocellular carcinoma，HCC）的竞争性内源RNA(competing endogenous RNAs，ceRNA）调控网络，寻找与HCC发生发展及预后相关的分子。方法：从TCGA数据库下载HCC转录组数据，通过“Perl”语言处理转化得到mRNA、lncRNA和miRNA的表达谱矩阵。通过“Edge R”包提取3种RNA的差异表达基因（differential expression genes，DGEs），其阈值为|log FC|>2.0 且P<0.01。通过数据库比对得到差异lncRNA-差异miRNA、差异miRNA-差异mRNA关系对，导入至Cytoscape 软件构建ceRNA调控网络图。整理3 种DGEs相关的生存数据，通过“Survival”包及Kaplan-Meier Plotter 分析软件进行生存分析，绘制DGEs的生存曲线，分析获取预后相关基因。结果：成功构建HCC的lncRNA相关ceRNA调控网络，通过该网络分析DGEs 间的相互作用和调控关系，获取3 条lncRNA-miRNA-mRNA 调控关系对，其中1 条调控通路（CCDC26-hsa-mir-141-EPHA2）符合ceRNA 理论。预后分析显示，14 个mRNA高表达组患者生存率低于低表达组，可作为HCC不良预后的生物标志物；1 个lncRNA（TSPEAR-AS1）和2 个mRNA（CPEB3 和PROK2）低表达组患者生存率低于高表达组（P<0.05 或P<0.01），可能是HCC的保护性基因。结论：通过HCC lncRNA 相关的ceRNA调控网络筛查到高表达的14 个mRNA可能为HCC不良预后相关分子，低表达的1 个lncRNA和2 个mRNA可能为HCC良好预后相关分子，研究结果为HCC治疗和预后测评提供了参考依据。     

基于竞争内源性RNA网络探讨槲皮素在肺腺癌中的作用靶点

目的:利用从肺腺癌（LUAD）数据集中筛选的差异表达基因（DEGs）、差异表达lncRNA（DElncRNA）及其上游miRNA,构建LUAD的竞争内源性RNA网络（ceRNA）,探索槲皮素可能的作用靶点。方法:从基因表达综合数据库（GEO）及肿瘤基因组图谱数据库（TCGA）中检索并筛选LUAD基因及lncRNA表达数据集,对差异表达基因（DEGs）进行富集和功能注释。使用在线数据库预测miRNA及lncRNA,建立ceRNA调控网络,并通过网络药理学分析槲皮素的药物靶点。结果:构建了AURKA与上游hsa-miR-363-3p及AP000553.1、LINC00858、AL354707.1的ceRNA调控网络,槲皮素与5DOS对接良好。结论:AP000553.1、LINC00858、AL354707.1与hsa-miR-363-3p竞争性调控AURKA,进一步影响LUAD预后,槲皮素可作用于AURKA。     

不同基因作为竞争性内源RNA参与膀胱癌调控的研究进展

竞争性内源RNA（competing endogenous RNA,ceRNA）是一种新的转录后RNA相互调控机制,越来越多的研究发现长链非编码RNA（long non-coding RNA,lncRNA）、环状RNA（circular RNA,circRNA）和假基因（pseudogenes）可以与微小RNA（microRNA,miRNA）竞争,影响靶RNA的稳定性或翻译,从而调控基因表达。近年来,不同基因作为ceRNA参与膀胱癌调控的研究已引起学者的广泛关注。为此,本文综述了lncRNA、circRNA和假基因作为ceRNA在膀胱癌中作用的研究进展,以及不同基因作为ceRNA调控膀胱癌的信号转导通路。从ceRNA分类的角度综述各类ceRNA在膀胱癌发生发展过程中的研究进展。     

ceRNA在肿瘤中的研究进展

竞争性内源RNA(competing endogenous RNA,ceRNA)是一类能与miRNA结合位点（miRNA response element,MRE）结合从而抑制miRISC形成,并能进一步增加相应mRNA的表达来达到RNA间相互沟通及转录后调控基因表达的一类非编码RNA。ceRNA主要包括长链非编码RNA（long non-coding RNA,lncRNA）、环状RNA（circular RNA,circRNA）、假基因、人工合成miRNA抑制剂及病毒miRNA抑制剂等。目前越来越多的研究表明ceRNA在肿瘤的基因调控及肿瘤细胞增殖、凋亡、细胞周期、侵袭和转移等各种生物过程中发挥重要作用。本文从ceRNA分类、调控网络及ceRNA在不同肿瘤中的研究进展作一综述。     

肺腺癌竞争内源性RNA网络的综合分析:基于肿瘤基因组图谱数据库

目的:基于肿瘤基因组图谱数据库（TCGA）架构肺腺癌（LUAD）竞争内源性RNA（ceRNA）网络，鉴定潜在的生存关联性生物标志物，并确定特异性治疗的小分子药物。方法:依据纳入研究标准，利用Edger软件筛选LUAD组织与正常组织(mRNA、lncRNA和miRNA)差异表达的基因。通过miRcode、miRDB、TargetScan和miRanda数据库对差异表达的RNA之间的关系进行分析，构建ceRNA网络。采用Kaplan-Meier方法分析ceRNA网络中的RNA表达量与生存预后的关系，通过富集分析对网络中的mRNA基因功能和调控通路进行分析。通过Cmap数据库筛选治疗LUAD的特异性小分子药物。利用D-lnc软件确定与关键的lncRNA相关的特异性小分子药物。结果:mRNAs(ELAVL2和PBK)、miRNAs(miR-13和miR-210)和lncRNAs(AP002478.1、DSCAM-AS1、LINC00269、LINC00470和LINC00483)与总生存期(OS)关系密切。喜树碱和甲萘醌有可能逆转LUAD的状态。卡铂、多西紫杉醇、帕比司他被确定为与关键lncRNA密切相关的药物。结论:ceRNA网络在LUAD中发挥重要作用，多种差异表达RNA与LUAD预后相关，可能成为潜在的肿瘤诊断标志物和治疗靶点。     

基于GEO数据库的小细胞肺癌lncRNA-miRNA-mRNA网络的构建与潜在的诊治靶点分析

目的 筛选参与小细胞肺癌(SCLC)发生发展的ncRNAs,预测其诊治的潜在靶点。方法 对人类肿瘤相关基因表达汇编(Gene Expression Omnibus, GEO)数据库中获得的表达谱芯片数据进行差异基因分析，构建了ceRNA网络，并进行功能富集分析、PPI网络构建。收集7例手术治疗的小细胞肺癌患者，采用实时定量PCR(qPCR)对部分差异基因(lncRNA、mRNA和miRNA)进行临床样本验证。结果 共有99个lncRNA、46个miRNA和73个mRNA分子参与调控网络的构建。GO和KEGG分析表明失调的mRNA主要参与小细胞肺癌的细胞癌变、增殖以及转移。PPI网络分析提示46个基因被认为是中心基因。qPCR结果显示，与癌旁组织比较，小细胞肺癌组织中lnc-ENST00000430027.3呈相对高表达(P<0.001),hsa-miR-143-5p呈相对低表达(P<0.01),FANCA呈相对高表达(P<0.01)。结论 构建“lncRNA-miRNA-mRNA”的ceRNA调控网络，可为小细胞肺癌诊治提供新的依据和靶点，为小细胞肺癌诊断和治疗的分子...     

lncRNA XIST介导的ceRNA调控网络在恶性肿瘤中作用的研究进展

lncRNA XIST是在哺乳动物中发现最早的lncRNAs之一，尤其是在X染色体失活相关疾病中起着重要作用，在胃癌、肝细胞癌、结直肠癌、胰腺癌、膀胱癌、骨肉瘤、鼻咽癌、食管鳞状细胞癌等非生殖系统相关恶性肿瘤中，其作为癌基因通过介导ceRNA调控网络，促进肿瘤细胞的增殖、迁移、侵袭和耐药等生物学行为。本文就 lncRNA XIST的分子生物学属性及介导的ceRNA调控网络在多种恶性肿瘤中作用的研究进展作一综述，以期为后续的肿瘤防治研究提供新的方向。     

Characteristics of mutations in the p53 gene in oral squamous cell carcinoma associated with betel quid chewing and cigarette smoking in Taiwanese

p53 mutations are etiologically associated with the development of oral squamous cell carcinomas (OSCCs) or are associated with exposure to specific carcinogens. In this study, we used PCR-single strand conformation polymorphism and DNA sequencing to analyze the conserved regions of the p53 gene (exons 5-9) in OSCC tumor specimens from 187 patients with varied histories of betel quid, tobacco and alcohol use. Ninety-one of the 187 OSCCs (48.66%) showed p53 gene mutations at exons 5-9. The incidence of p53 mutations was not associated with age, sex, TNM stage, status of cigarette smoking or betel quid chewing. However, alcohol drinkers exhibited a significantly higher incidence (57/101, 56.44%) of p53 mutations than non-users (39.53%, 34/86) (P = 0.02). The effect of alcohol on the incidence of p53 mutations was still statistically significant (RR = 2.24; 95% CI, 1.21-4.15) after adjustment for cigarette smoking and betel quid (BQ) chewing. G:C to A:T transitions were the predominant mutations observed and associated with BQ and tobacco use. Alcohol drinking could enhance these transitions. After adjustment for cigarette smoking and BQ chewing, alcohol drinking still showed an independent effect on G:C to A:T transitions (RR = 2.41; 95% CI, 1.01-5.74). These findings strongly suggest an important contributive role of tobacco carcinogens to p53 mutation in this series of Taiwanese OSCCs and alcohol might enhance these mutagenic effects. As safrole-DNA adducts have been detected in 77% (23/30) of the OSCC tissues from Taiwanese oral cancer patients with a BQ chewing history, we cannot rule out the possibility that safrole or other carcinogens present in the BQ may cause a similar pattern of mutagenesis. Determination of the role of safrole and other carcinogens present in BQ on the pattern of p53 gene mutation in OSCC will require further study.     

Safrole-like DNA adducts in oral tissue from oral cancer patients with a betel quid chewing history

Betel quid (BQ) chewing has been associated with an increased risk of oral squamous cell carcinoma (OSCC) and oral submucous fibrosis (OSF). Piper betel inflorescence, which contains 15 mg/g safrole, is a unique ingredient of BQ in Taiwan. Chewing such prepared BQ may contribute to safrole exposure in human beings (420 microM safrole in saliva). Safrole is a known rodent hepatocarcinogen, yet its carcinogenicity in human beings is largely undetermined. In this study, using a (32)P-post-labeling method, we have found a high frequency of safrole-like DNA adducts in BQ-associated OSCC (77%, 23/30) and non-cancerous matched tissue (NCMT) (97%, 29/30). This was in contrast to the absence (< 1/10(9) nucleotides) of such adducts in all of non-BQ-associated OSCC and their paired NCMT (P < 0.001). Six of seven OSF also exhibited the same safrole-like DNA adduct. The DNA adduct levels in OSF and NCMT were significantly higher than in OSCC (P < 0.05). Using co-chromatography and rechromatography techniques, we further demonstrated that these adducts were identical to synthetic safrole-dGMP adducts as well as DNA adducts from 1'-hydroxysafrole-treated HepG2 cells. These results suggest that safrole forms stable safrole-DNA adducts in human oral tissue following BQ chewing, which may contribute to oral carcinogenesis.     

Specific induction of the high-molecular-weight microtubule-associated protein 2 (hmw-MAP2) by betel quid extract in cultured oral keratinocytes: clinical implications in betel quid-associated oral squamous cell carcinoma (OSCC)

Betel quid (BQ) chewing, a popular habit in numerous Asian countries including India and Taiwan, has a strong correlation with an increased risk of oral squamous cell carcinoma (OSCC). While substantial efforts have been made to test the cytotoxic, genotoxic and mutagenic effects of BQ extract and its components, the disease mechanisms underlying BQ-induced oral carcinogenesis remain obscure. Here, we show that a neuronal protein, microtubule-associated protein 2 (MAP2), was induced by BQ extract in cultured normal human oral keratinocytes (NHOKs). Subsequent analyses demonstrated that such induction was more eminent and consistent in the high-molecular-weight isoform of MAP2 (hmw-MAP2) than that in its low-molecular-weight counterpart (lmw-MAP2). Furthermore, we analyzed expression of hmw-MAP2 protein in 88 oral specimens consisting of clinicopathologically pre-malignant (leukoplakia) and malignant (OSCC) lesions, along with their adjacent normal mucosa. Immunohistochemistry revealed that, with the exposure to BQ, the hmw-MAP2 was over-expressed in 41.2% (7/17) of OSCC, 11.2% (1/9) of leukoplakia and none (0/19) of normal mucosa. In contrast, expression of the hmw-MAP2 was barely detected in BQ-free OSCC. These results suggest a significant correlation between expression of the hmw-MAP2 and BQ-associated progression of oral carcinogenesis (P=0.0046). Interestingly, the hmw-MAP2 was found to preferentially express in histopathologically less differentiated OSCC (P=0.014); the percentages of positive staining in poorly, moderately and well differentiated OSCC were 62.5, 21.4 and 7.1%, respectively. However, BQ chewing appeared to have marginal correlation with such propensity. Finally, we show that the majority of hmw-MAP2-positive poorly differentiated lesions were also histopathologically invasive. Taken together, these findings suggest the possibility that the hmw-MAP2 may be a diagnostic marker for BQ-chewing lesions and a potential therapeutic target. To our knowledge, this study has provided the first clinical implication that closely links a cytoskeletal protein to BQ-associated oral cancer.     

Univariate and multivariate analysis of prognostic significance of betel quid chewing in squamous cell carcinoma of buccal mucosa in Taiwan

BACKGROUND AND OBJECTIVES: While betel quid (BQ) chewing is clearly the most avoidable risk factor of squamous cell carcinoma of buccal mucosa (BMSCC), little is known about the influence of this habit on the prognosis of BMSCC. METHODS: We surveyed 280 patients with BMSCC who were treated during an 8-year period in a cohort study to assess the independent predictive value of pretreatment BQ chewing habit on the prognosis by univariate and multivariate analysis. RESULTS: We found by univariate analysis that sex, age, clinical stage, smoking, and BQ chewing significantly affected the patients' prognosis and only age, clinical stage, and BQ chewing had significant influence on prognosis by multivariate analysis (P < 0.05). Further analysis revealed that the prognostic effect of BQ chewing changed in a dose- and time-dependent manner. The risk of death was 31.4-fold higher in heavy user (duration >30 years, daily consumption >30 quids, age of start <20 years old) when compared to those who chewed BQ to a milder degree (duration <10 years, daily consumption <15 quids, age of start > or =20 years old ) (P < 0.001). CONCLUSIONS: Pretreatment BQ chewing habit worsens the prognosis of BMSCC in Taiwan. BQ chewing is a prognostic indicator that can be used in conjunction with clinical staging to help plan the treatment for the patients.     

Risk of p53 gene mutation in esophageal squamous cell carcinoma and habit of betel quid chewing in Taiwanese

A recent report suggested that BQ (BQ) chewing significantly correlated with the occurrence of esophageal squamous cell carcinoma (ESCC) in Taiwanese. BQ chewing was shown to be associated with p53 mutation in oral cancers. However, the relationship between BQ chewing and p53 mutation in ESCC is unclear. Seventy-five primary ESCC patients were enrolled for mutational analysis of the p53 gene using polymerase chain amplification and direct sequencing of amplified product. Thirty-seven mutations of the p53 gene were detected in 45.5% (34/75) of tumor specimens. These mutations significantly clustered in exon 5 (21/37) of the p53 gene. The incidence of p53 mutations did not associate with clinicopathological characteristics or the habits of cigarette smoking or alcohol consumption. However, BQ chewers exhibited significantly higher incidence of p53 gene mutations than non-chewers (67.6% vs 32.4%, P = 0.007). After controlling the confounding factors of cigarette smoking and alcohol intake, BQ chewing still showed significant association with the incidence of p53 mutation in ESCCs (RR = 4.23; 95% CI, 1.317-13.60). The A:T to G:C transition (8/37, 21.6%) and G:C to T:A transversion (5/23, 13.5%) were the prevalent spectrum of p53 gene mutations. All A:T to G:C transitional mutations occurred in patients with the habits of BQ chewing and cigarette smoking. Noticeably, alcohol consumption could enhance this peculiar spectrum of p53 mutation in ESCC. Accordingly, p53 might be an important molecular target of BQ carcinogens in the development of ESCC in Taiwanese.     

CYP26B1 is a novel candidate gene for betel quid-related oral squamous cell carcinoma

Substantial epidemiological data suggest a role for environmental factors (for example, the use of alcohol, betel quid (BQ), and cigarettes) in the occurrence of oral squamous cell carcinoma (OSCC), but the evidence for the genes involved has been inconsistent. This study was to investigate the role of CYP26B1, together with the use of alcohol, BQ, and cigarettes, on BQ-related OSCC. The association study (247 OSCC cases and 338 controls) was conducted to examine the possible interplay between CYP26B1 polymorphisms and alcohol, BQ, and cigarettes use. Additional gene expression was evaluated between OSCC tissue and adjacent normal tissue. The genetic polymorphism AA of CYP26B1 appeared to correlate with the risk of OSCC (OR=2.26; 95% CI, 1.35-3.80). Chewing BQ multiplicatively interacted with CYP26B1 AA to increase the OSCC risk (aOR=70.04; 95% CI, 13.62-360.11). The independent risk of OSCC was observed among BQ chewers with CYP26B1 AA, and compared with chewers with the CYP26B1 CC genotype (stratified aOR=2.88; 95% CI, 1.07-7.74). Increased expression of CYP26B1 was observed in tumor tissue compared with adjacent normal tissue. The CYP26B1 gene plays a novel role in the BQ dependent pathogenesis of OSCC.     

Betel quid chewing and the risk of oral and oropharyngeal cancers: A meta‐analysis with implications for cancer control

We conducted a random‐effects meta‐analysis of 50 publications assessing the relationship between oral/oropharyngeal cancer and chewing betel quid, with (BQ+T) or without added tobacco (BQ‐T), a common practice in many parts of Asia and globally among Asian immigrants. Exposure‐response, by daily amount and years of BQ chewed, was assessed using spline models. Attributable fractions (PAF%) were calculated to estimate the public health impact if BQ were no longer chewed. The meta‐relative risk (mRR) for oral/oropharyngeal cancer in the Indian subcontinent was 2.56 (95%CI, 2.00–3.28; 15 studies) for BQ‐T and 7.74 (95%CI, 5.38–11.13; 31 studies) for BQ+T; in Taiwan, China, the mRR for BQ‐T was 10.98 (95%CI, 4.86–24.84; 13 studies). Restricting to studies that adjusted for tobacco and alcohol use had only a small effect on the risk estimates. For BQ+T in the Indian subcontinent, the mRR was much higher in women (mRR, 14.56; 95%CI, 7.63–27.76) than in men. Exposure‐response analyses showed that the risk of oral/oropharyngeal cancer increased with increasing daily amount and duration (years) of chewing BQ in India and Taiwan, China. Roughly half of oral cancers in these countries could be prevented if BQ were no longer chewed (PAF% = 53.7% for BQ‐T in Taiwan, China; PAF% = 49.5% for BQ+T in India). We demonstrate that betel quid chewing, with or without added tobacco, increases the risk of oral/oropharyngeal cancer in an exposure‐dependent manner, independently of tobacco and alcohol use. Further work is needed to explain the higher risks associated with chewing BQ‐T in Taiwan, China.     

Impact of Chronic Hepatitis B and Hepatitis C on Adverse Hepatic Fibrosis in Hepatocellular Carcinoma Related to Betel Quid Chewing

The pathogenesis of hepatocellular carcinoma (HCC) related to habitual betel quid (BQ) chewing is unclear.Risk of HCCis increased with adverse hepatic fibrosis. This study aimed to assess the impact of chronic viralhepatitis on adverse hepatic fibrosis in HCC related to BQ chewing. This hospital-based case-control study enrolled200 pairs of age- and gender-matched patients with HCC and unrelated healthy controls. Serologic hepatitisB surface antigen (HBsAg), antibodies to hepatitis C virus (anti-HCV), α-fetoprotein (AFP), and surrogatemarkers for significant hepatic fibrosis were measured. Information on substance-use habits was obtained witha questionnaire. By analysis of surrogate markers for hepatic fibrosis, the prevalence of significant hepaticfibrosis in patients chewing BQ was between 45.8% and 91.7%, whereas that for patients without BQ chewingwas between 18.4% and 57.9%. The difference was significant (P <0.05 for each surrogate marker). Multivariateanalysis indicated that cirrhosis with Child-Pugh C (odds ratio (OR) = 3.28; 95% confidence interval (CI), 1.29-8.37), thrombocytopenia (OR = 3.92, 95% CI, 1.77-8.68), AFP >400 mg/L (OR = 2.21, 95% CI, 1.05-4.66) andmale gender (OR = 4.06, 95% CI, 1.29-12.77) were independent factors associated with habitual BQ chewing.In conclusion, adverse hepatic fibrosis and severe liver damage play important roles in the pathogenesis of BQrelatedHCC, which could be aggravated by chronic hepatitis B and hepatitis C. BQ-cessation programs andprevention of chronic HBV/HCV infection are needed to prevent HCC related to BQ chewing.