Synchronous gastric and colon cancers:Important to consider hereditary syndromes and chronic inflammatory disease associations

In this editorial we comment on the manuscript,describing management and surveillance strategies in synchronous and metachronous,gastric and colon cancers.Synchronous or metachronous primary malignancies at different sites of the gastrointestinal tract pose a unique diagnostic and therapeutic challenge.Multidisciplinary services and strategies are required for the management of multiple site primary malignancies,to provide the best oncological outcomes.Although this study highlights the dual cancers in 76 sporadic cases,the authors excluded 55 patients due to combination of factors which includes;incomplete clinical data,genetic syndrome,gastric stump cancers.In addition,the authors did not elaborate if any patients presented with signet ring cell morphology,E-cadherin mutations or presence of inflammatory bowel disease.Genetic and mutational errors and epithelial field defects from chronic inflammatory diseases of the gastrointestinal tract are important when considering synchronous gastric and colonic cancers.We will briefly discuss these in this editorial.

The Role of Ras Gene Mutation in Gastric Cancer and Precancerous Lesions

Abnormality of ras gene family was studied in a total of 206 cases of gastric cancer and precancerous lesions by PCR-RFLP, PCR-SSCP and DNA sequencing. The results showed that mutation rate of H-ras 12 codon in metaplasia,atypical hyperplasia, early-stage cancer and advanced cancer was 16. 7%, 31. 2 %, 50. 0%, and 32. 2%, respectively. In the groups of superficial gastritis and normal controls, no mutation were detected in codon 12 of ras. Mutations of Hras 61 codon and N-ras 12 codon in various groups were the same as those in normal control. K-ras 12 codon mutation was detected in only 2 cases of gastric cancer by using PCR-SSCP, but it was not detected by DNA sequencing, which may be polymorphism. All H-ras 12 codon mutations were G→T mutation. There were significant difference between the groups of metaplasia, dysplasia, gastric carcinoma and normal control group (P<0.05, P<0.01, P<0.01,respectively). It was concluded that H-ras 12 codon mutation was an early event and may play an important role in gastric carcinogenesis. Although K-ras, N-ras mutation rates are high in colon cancer and leukemia, it seems to bear no relationship with gastric cancer.

Transglutaminase 2 serves as a pathogenic hub gene of KRAS mutant colon cancer based on integrated analysis

BACKGROUND Colon cancer is acknowledged as one of the most common malignancies worldwide,ranking third in United States regarding incidence and mortality.Notably,approximately 40%of colon cancer cases harbor oncogenic KRAS mutations,resulting in the continuous activation of epidermal growth factor receptor signaling.AIM To investigate the key pathogenic genes in KRAS mutant colon cancer holds considerable importance.METHODS Weighted gene co-expression network analysis,in combination with additional bioinformatics analysis,were conducted to screen the key factors driving the progression of KRAS mutant colon cancer.Meanwhile,various in vitro experiments were also conducted to explore the biological function of transglutaminase 2(TGM2).RESULTS Integrated analysis demonstrated that TGM2 acted as an independent prognostic factor for progression-free survival.Immunohistochemical analysis on tissue microarrays revealed that TGM2 was associated with an elevated probability of perineural invasion in patients with KRAS mutant colon cancer.Additionally,biological roles of the key gene TGM2 was also assessed,suggesting that the downregulation of TGM2 attenuated the proliferation,invasion,and migration of the KRAS mutant colon cancer cell line.CONCLUSION This study underscores the potential significance of TGM2 in the progression of KRAS mutant colon cancer.This insight not only offers a theoretical foundation for therapeutic approaches but also highlights the need for additional clinical trials and fundamental research to support our preliminary findings.

Distinguishing Rectal Cancer from Colon Cancer Based on the Support Vector Machine Method and RNA-sequencing Data

Colorectal cancer (CRC) is the third most commonly diagnosed cancer worldwide.Several studies have indicated that rectal cancer is significantly different from colon cancer interms of treatment, prognosis, and metastasis. Recently, the differential mRNA expression of coloncancer and rectal cancer has received a great deal of attention. The current study aimed to identifysignificant differences between colon cancer and rectal cancer based on RNA sequencing (RNA-seq)data via support vector machines (SVM). Here, 393 CRC samples from the The Cancer GenomeAtlas (TCGA) database were investigated, including 298 patients with colon cancer and 95 withrectal cancer. Following the random forest (RF) analysis of the mRNA expression data, 96 genessuch as HOXB13, PR4C, and BCLAFI were identified and utilized to build the SVM classificationmodel with the Leave-One-Out Cross-validation (LOOCV) algorithm. In the training (n= 196)and the validation cohorts (n=197), the accuracy (82. 1 % and 82.2 %, respectively) and the AUC(0.87 and 0.91, respectively) indicated that the established optimal SVM classification modeldistinguished colon cancer from rectal cancer reasonably. However, additional experiments arerequired to validate the predicted gene expression levels and functions.

Deletion and Mutation of WWOX Exons 6-8 in Human Non-small Cell Lung Cancer

Summary: To examine the deletion and point mutation of WWOX (WW domain containing oxidoreductase) exons 6-8 in human non-small cell lung cancer and their possible relationship with pathological stages, tumor tissues and the corresponding normal tissues were obtained from 44 Chinese patients who had undergone surgery for non-small cell lung cancer. RNA was extracted from each sample and deletion and mutation of WWOX exons 6-8 were analyzed by RT-PCR and DNA sequencing. Our results showed that 28 of 44 (63.6 %) lung cancer samples showed loss of WWOX exons 6-8 transcript and the deletion was detected in only 3 of 44 (6.8 %) corresponding adjacent normal tissues (P<0.05). The transcript sequencing analyses of the 16 lung cancer samples without transcript loss of WWOX exons 6-8 revealed no difference from the sequence of GenBank. Moreover, the deletion of WWOX exons 6-8 was significantly higher in the smokers when compared with the non-smokers. It is also higher in the men and squamous carcinomas than in women and adenocarcinomas (P<0.05). The deletion, however, was not found to be associated with pathological stages of the tumors. Our study documented a high incidence of deletion of WWOX exons 6-8 in non-small cell lung cancer in Chinese patients and suggested that the frequent loss of WWOX exons 6-8 might play an important role in the tumorigenesis of non-small cell lung cancer in Chinese. WWOX exons 6-8 may serves as a candidate molecular target of smoking carcinogenesis, and point mutation is not a predominant way of alteration of WWOX exons 6-8.

DETECTION OF GENE MUTATION IN SPUTUM OF LUNG CANCER PATIENT

Lungcancerisacommonmalignanttumor,whichhasahighincidenceandmortalityrate.Therefore,itisnecessarytoseekanewmethodforthediagnosis,especiallytheearlydiagnosisoflungcancer.Thedevelopmentofmolecularbiologymakesthegenediagnosisoflungcancerpossible.PCR-SSCP...

巢式PCR-RFLP检测大便K-ras基因第12位密码子点突变

用巢式PCR－RFLP分析技术，扩增大便中K－ras基因第12位密码子所处的一段外显子，根据BstNI内切酶的多态性，分析K－ras基因第12位密码子是否突变。结果在22例结肠直肠癌患者的术后石蜡包埋组织中，检出10例（45.5％）K－ras突变，该10例患者中，有8例（36．4％）患者的术前大便中检出K－ras基因突变。这种方法快速、简便、特异、敏感，且可避免大便中多种物质对PCR的干扰。对可疑人群具有实际应用价值。

应用PCR-SSCP法检测乳腺癌患者血中p53基因突变

[目的]探讨乳腺癌患者外周血p53基因突变的临床意义.[方法]应用聚合酶链反应-单链构象多态性分析（PCR-SSCP）的方法检测35例乳腺癌患者和20例良性肿瘤患者外周血中p53基因外显子5～8的突变情况.[结果]乳腺癌患者p53基因突变的检出率为40.0%（14/35）,显著高于对照组良性肿瘤患者的5.0%（1/20）（P＜0.05）.[结论]p53基因突变在乳腺癌外周血中有较高的检出率,是乳腺癌发生的早期信号.

PCR-SSCP检测肺癌EGFR基因突变

目的分析PCR-SSCP检测肺癌EGFR基因突变的敏感性。方法应用PCR-SSCP检测36例肺癌标本的突变情况并与其它文献进行比较。结果在36例肺癌标本中,有11例存在突变,突变率为30.6%(11/36),其中男性2例,女性9例。结论PCR-SSCP检测肺癌EGFR基因突变具有较高的敏感性,与其它文献报道一致。

甲基化特异PCR 方法对p16肿瘤抑制基因的研究

目的：研究肺癌、结肠癌组织中 p16 肿瘤抑制基因甲基化状态及其临床意义。方法：采用甲基化特异性聚合酶链反应（MSP）方法，特异扩增分析甲基化和非甲基化两种状态。结果：p16基因在人胚肺组织处于非甲基化状态；在人肺巨细胞癌细胞系（PLA801-D)呈甲基化状态；在检测的肿瘤标本中，60％ 肺癌组织及 70％ 结肠癌组织出现甲基化状态。结论：p16 基因的甲基化状态是肺癌、结肠癌组织中 p16 基因异常形式之一，有可能成为肿瘤分子诊断的重要标志，MSP 方法有临床普及应用前景。

应用ARMS- PCR法检测非小细胞肺癌EGFR基因突变及其临床意义

目的探讨依据ARMS法原理,采用实时荧光定量PCR法检测非小细胞肺癌EGFR基因突变的可行性并分析检测的临床意义。方法收集苏州大学附属第三医院2010年9月至2011年12月确诊为NSCLC患者标本64例,DNA提取后,采用ARMS-PCR法对标本进行EGFR基因突变检测并分析EGFR基因突变与患者临床病理参数的关系。结果本研究发现64例NSCLC患者的EGFR总突变率为23.4%,均为19号外显子缺失(19-Del)和21号外显子(L858R)突变;女性EGFR突变率高于男性,腺癌高于鳞状细胞癌。结论 EGFR基因突变检测有助于规范表皮生长因子受体酪氨酸激酶抑制剂(EGFR-TKI)在NSCLC患者中的临床应用。

PCR-SSCP法检测非小细胞肺癌中nm23基因突变的研究

目的 探讨nm2 3基因突变在人非小细胞肺癌中的作用。方法 采用PCR SSCP方法检测了 5 3例非小细胞肺癌原发灶组织和 5例肺部其他良恶性病变组织的nm2 3基因突变情况。结果 所有病例均未发生nm2 3基因的突变。结论 nm2

石棉肺癌组织中p53基因突变的PCR-SSCP及部分序列分析研究

为研究石棉致癌的分子机理，本研究首次直接分析石蜡包埋的石棉肺癌病例肺组织中抗癌基因ｐ５３的突变情况。利用聚合酶链反应－单链构象多态性（ＰＣＲ－ＳＳＣＰ）技术分析１０例石棉肺癌病例，检测到７例（８个片段）存在ｐ５３基因的突变，突变位点位于其第５、７和８外显子区内，突变率为７０％。对其中２例进行ＰＣＲ产物直接序列分析，发现其基因一级结构分别发生Ｃ→Ａ和Ｔ→Ａ转换。

突变体富集PCR法检测非小细胞肺癌病理组织EGFR基因突变

目的:建立突变体富集PCR法检测非小细胞肺癌病理组织中的EGFR基因突变。方法:选取55例细支气管肺泡癌和59例非小细胞肺癌病理组织蜡块,提取基因组DNA,采用不同的突变体富集PCR法(PCR-PAGE和PCR-RLFP)检测EGFR基因常见的19和21外显子突变,并经过直接测序验证。结果:在59例非小细胞肺癌中共检测出EGFR基因突变22例,突变率为37·3%(22/59)。55例细支气管肺泡癌中共检测出24例基因突变,突变率为43·6%(24/55)。经直接测序验证,EGFR19外显子有3种类型缺失突变。EGFR21外显子的错义突变为L858R。结论:突变体富集PCR法准确、快速、经济,便于临床筛查非小细胞肺癌病理组织中的EGFR基因突变。

应用PCR-SSCP/HMA分析食管癌组织中p73基因的突变

目的 检测 p73基因在食管癌中的突变状况 ,分析p73基因在食管癌发生、发展中的角色和作用机制。 方法 采用RT PCR ,单链PCR构象多态性 (PCR SSCP)和异源双链体迁移率 (HMA)技术检测 37例食管鳞状细胞癌肿瘤组织、癌旁组织、区域淋巴结和相应正常食管组织中p73mRNA的表达和基因的突变情况 ,并探讨与食管癌临床病理特征的关系。结果 37例食管肿瘤组织中有 2 1例 (51 8% ) p73mRNA过度表达 ,其阳性表达率显著高于相应的癌旁组织、区域淋巴结和正常组织 (P <0 0 5) ,并且与食管癌TNM分期、细胞分化程度和病理类型均无明显关系 (P >0 0 5) ;未发现食管癌组织中 p73基因有任何突变。 结论 p73基因在食管癌中的过表达和正常基因型没有能够阻止癌症的恶化 。

建立基于脱氧次黄嘌呤核苷修饰等位基因PCR方法检测EGFR基因热点突变

目的建立基于脱氧次黄嘌呤修饰等位基因特异荧光聚合酶链反应(d I-AS-PCR)方法,检测表皮生长因子受体(EGFR)基因热点突变L858R和19del(2235~2249,2236~2250)。方法设计包含扩增EGFR基因L858R和19del热点突变在内的特异性引物、荧光探针和内参体系;且特异检测基因突变的等位基因PCR引物3’-末端n-1或n-2位置采用脱氧次黄嘌呤(d I)修饰。建立标准品和质控样品,应用荧光PCR检测,并分析结果。结果 d I-AS-PCR能够在野生型背景下检测低于0.1%的突变,对50例肺癌临床样本进行检测,有9例(18%)EGFR基因发生L858R突变,14(28%)例19del基因突变。EGFR基因热点突变率为46%,其检测结果与DNA测序完全一致。结论基于d I-AS-PCR技术的特异荧光PCR检测EGFR基因热点突变,敏感性高,特异性强,可以应用于临床EGFR基因突变检测,指导个性化用药及酪氨酸激酶抑制剂(TKI)治疗耐药监测。

应用PCR-SSCP方法检测非小细胞肺癌患者血浆p53基因突变

目的探讨应用聚合酶链反应（PCR）-单链构象多态性分析（SSCP）检测非小细胞肺癌（NSCLC）患者血浆循环核酸p53基因突变检测的可行性,研究NSCLC患者血浆p53基因突变对NSCLC发生、发展和预后估计的意义.方法应用PCR扩增p53基因外显子5～8,并用SSCP分析产物突变情况,再与各生物参数进行相关研究.结果 10例健康人血浆循环核酸p53基因突变检测结果均为阴性,60例NSCLC患者血浆中p53基因5～8外显子突变测定的结果显示有28例出现突变,占47%（28/60）.p53基因突变率与术后肿瘤残留、对治疗的反应显著相关（P＜0.05）,与临床分期、年龄、性别、病理分型无关（P＞0.05）.结论血浆循环核酸p53基因突变对NSCLC的早期诊断、病情监测、预后评估都具有指导意义.PCR-SSCP分析简便、可靠,可作为检测大量标本的突变的可行性方法.

直接测序法与荧光定量PCR法检测非小细胞肺癌EGFR基因突变对比分析

目的:比较直接测序和荧光定量PCR法检测非小细胞肺癌(NSCLC)EGFR基因突变状态结果的差异。方法:分别采用荧光定量PCR(蝎形探针扩增阻滞突变系统ARMS)和直接测序法检测155例石蜡包埋NSCLC样本中EGFR基因18,19,20,21号外显子突变状态,比较检测结果的差异。结果:80例手术切除样本中,直接测序检测EGFR突变阳性者25例(31.25%),ARMS法为28例(35%),差异无统计学意义(P>0.05);75例活检样本中,直接测序检测EGFR突变阳性者19例(25.33%),ARMS法为26例(34.67%),差异具有统计学意义(P<0.05)。结论:对于手术切除样本,直接测序和ARMS法均可有效检测出EGFR基因突变状态;但对于活检组织样本,ARMS较直接测序灵敏性更高。

KRAS基因突变亚型对直肠癌新辅助治疗后肿瘤退缩的影响

目的 探讨KRAS基因突变亚型与直肠癌新辅助治疗后肿瘤退缩反应及临床病理特征的相关性。方法 收集新辅助治疗后行肿瘤根治术的局部晚期和转移性直肠癌(Ⅲ~Ⅳ期)患者的临床病理资料,分析KRAS基因突变亚型与肿瘤退缩反应以及残余肿瘤组织学分级、肿瘤浸润T淋巴细胞水平的关系。结果 共纳入95例患者,其中肿瘤退缩不良患者55例。肿瘤退缩不良患者KRAS基因第2外显子G12V突变和第4外显子A146突变的发生率高于肿瘤退缩显著患者(P<0.05)。KRAS基因G12D突变的患者中,肿瘤退缩不良者肿瘤组织学分级高于肿瘤退缩显著者(P=0.027)。肿瘤退缩不良患者中,KRAS基因G12D突变者肿瘤CD8^(+)、PD-1^(+)T淋巴细胞浸润水平高于G12V、G13D突变患者(P<0.05)。结论 KRAS基因G12V突变和A146突变提示直肠癌新辅助治疗效果不佳;对于肿瘤退缩不良的KRAS基因G12D突变患者,可通过检测CD8^(+)、PD-1^(+)T淋巴细胞浸润水平,筛选免疫治疗的潜在获益者。

PCR扩增肠癌组织p53基因失败的原因分析与解决

分析与解决PCR扩增肠癌组织p53基因时失败的问题,为如何解决PCR技术在科研应用中的问题提供一个实例.方法 经初步分析,将失败原因确定为DNA模板有问题、采用下列方法处理模板再进行PCR反应：用70%乙醇再清洗模板、用新标本重新提取模板、用PCR反应成功与失败的模板配成混合模板.比较PCR成功与失败所用模板的差异,寻找与证实失败的原因.结果 最终发现溶解DNA模板的TE缓冲液浓度过高,使PCR扩增失败,排除之后获得成功.结论 当PCR扩增失败时,不妨从最简单的反应体系的离子环境先找原因.