结直肠癌APC、K-ras、p53基因突变检测

目的:探讨结直肠癌中APC、K-ras、p53基因突变模式。方法:应用酚/氯仿法提取48例结直肠癌组织及其相应正常黏膜组织的DNA,用聚合酶链反应(PCR)、单链构象多态性分析(SSCP)和DNA测序等方法检测APC基因第15外显子突变密集区(mutation cluster region,MCR)区段、K-ras和p53基因的突变。结果:APC、K-ras和p53基因的突变率分别为37.5%(18/48)、43.8%(21/48)和35.4%(17/48)。48例结直肠癌组织中,有42例发生APC、K-ras或p53基因突变,突变率高达87.5%(42/48),其中仅有APC、K-ras或p53 1种基因发生突变的发生率分别为16.7%(8/48)、25.0%(12/48)和20.8%(10/48)。单独1种基因发生突变的总发生率为62.5%(30/48)。APC和p53,APC和K-ras或p53和K-ras 2种基因均有突变的发生率分别为6.3%(3/48)、10.4%(5/48)和4.2%(2/48)。APC、K-ras和p53 3种基因均发生突变的发生率为4.2%(2/48)。2种和3种基因均发生突变的总发生率为25%(12/48)。结论:结直肠癌的发生、发展并不完全遵循由正常结直肠黏膜上皮细胞向腺瘤和侵袭性癌转化的过程中,及依次发生“APC→K-ras→p53→DCC”突变累积这一经典的结直肠癌发生发展模式,可能存在其他结直肠癌发病机制。     

肺癌患者组织和痰液中p53基因、K-ras基因突变

 目的　探讨p53、K-ras基因在肺癌患者癌组织及相应痰液中改变情况及其联合检测在肺癌早期诊断中的价值。方法　对59例肺癌组织和14例肺部良性组织及相应痰液,应用PCR-SSCP-银染法检测了p53基因第5～8外显子突变情况;应用PCR-RFLP法对K-ras基因突变进行了检测。结果　p53基因在肺癌组织中突变率为37.3%,K-ras基因在肺腺癌突变率为48.0%,其它类型肺癌突变率仅为8.8%。相应痰液中两基因突变率分别为33.8%和44.0%,与组织中的突变率无明显差异,P<0.01。良性组织及相应痰液中两基因均无突变。吸烟患者的突变率(48.7%,68.5%)明显高于非吸烟患者的突变率(15.0%,11.1%),P<0.01;两基因的联合检测在肺癌的早期诊断中的价值(54.2%)明显优于单基因的检测,P<0.05。结论　痰液和组织中的基因突变率基本相似,即痰液中脱落细胞的分子遗传学改变能反映肺组织情况。因此以痰液为目标多基因的联合检测可能有助于肺癌的诊断。     

p53基因突变在胃癌发生中的作用

胃癌的发生发展是多基因参与、多步骤进行的过程.而p53基因突变作为一种重要抑癌基因失活方式,在其中发挥了重要作用.现综述近年来在胃癌中发现的p53基因突变类型、方式、地区差异性及与多种因素之间的关系等方面研究进展.     

胃癌组织p53基因突变点检测意义

目的 进一步明确胃黏膜癌变进程中基因变异的规律。方法 用PCR单链构象多态性（SSCP）方法对18例胃癌组织以及正常胃黏膜中p53基因突变情况进行分析。结果 p53基因突变10例（10／18）占55．6％,其中第5外显子突变4例,第7外显子突变3例,第6外显子突变2例,第8例显子突变1例。结论 p53基因结构和产物的变化可以作为肿瘤相关标志,对评估胃癌病人的预后和指导临床治疗有一定的作用。     

肝硬化及肝癌p53基因突变的实验研究

目的：研究肝硬化及肝癌p53基因的突变情况。方法：选择80例肝硬化、肝癌标本,分别以PCR－SSCP法,双链DNA序列测定法研究其p53基因外显子的突变情况及突变位点。结果：62例肝癌标本p53总突变率为19．4％,其中,早、中、晚期突变率分别为10．5％、15．0％、35．0％;18例肝硬化标本p53总突变率为5．6％;第7外显子的突变发生在249位密码子第3号碱基上,为G：C→T：A的转换突变。结论：p53基因突变发生在肝细胞发生形态学改变之初,随着肝癌的进展逐渐积累,突变率呈上升趋势,故p53基因突变很可能是启动癌变过程的重要因素之一。     

抑癌基因p53在胃癌组织的表达及临床意义

目的：探讨ｐ５３基因在胃癌的表达与临床、病理因素及预后的关系,探索ｐ５３基因异常表达在胃癌发生过程中的作用。方法：使用抗ｐ５３蛋白单克隆抗体ＤＯ－７,对９９例人胃癌组织ｐ５３的表达进行免疫组化研究。结果：４８例胃癌组织ｐ５３基因表达阳性（４９％）。ｐ５３基因表达与胃癌的大体类型、分化工、淋巴结转移等指标均无明显关系。在１０例不典型增生的癌旁组织中,２例呈阳性反应;在４例有肠上皮化生的癌旁组织中及其     

肝硬化及肝癌p53基因突变的实验研究

目的 :研究肝硬化及肝癌p53基因的突变情况。方法 :选择 80例肝硬化、肝癌标本 ,分别以PCR SSCP法 ,双链DNA序列测定法研究其p53基因外显子的突变情况及突变位点。结果 :62例肝癌标本p53总突变率为 19 4 % ,其中 ,早、中、晚期突变率分别为 10 5%、15 0 %、35 0 % ;18例肝硬化标本p53总突变率为 5 6% ;第 7外显子的突变发生在 2 4 9位密码子第 3号碱基上 ,为G :C→T :A的颠换突变 ;第 8外显子的突变发生在 2 73位密码子第 1号碱基上 ,为C :G→T :A的转换突变。结论 :p53基因突变发生在肝细胞发生形态学改变之初 ,随着肝癌的进展逐渐积累 ,突变率呈上升趋势 ,故p53基因突变很可能是启动癌变过程的重要因素之一。     

胃癌组织中p53基因突变及p53和mdm2蛋白表达的研究

目的：探讨mdm2和p53基因异常在胃癌发生发展中的作用以及两者相关性。方法：应用免疫组化技术检测58例胃癌组织以及相应癌旁组织中mdm2和p53蛋白的表达；PCR-SSCP银染技术检测p53基因exon5～8突变情况。结果：胃癌组p53和mdm2蛋白阳性率分别为86．21％（50／58）和29．31％（17／58），癌旁组织组p53和mdm2蛋白均为阴性，胃癌组p53和mdm2蛋白阳性率明显高于癌旁组织组，两两间差异有统计学意义，P=0．0000、P=0．0001。胃癌组织中p53和mdm2蛋白表达率与肿瘤大小、组织学类型、分化程度、淋巴结转移以及患者年龄等无显著相关性，P〉0．05。mdm2蛋白阳性表达与p53蛋白过表达呈显著正相关,X^2=11．1839，P=0．0008，r=0．4391。2例胃癌组织检测到p53基因突变，突变均位于exon5，58例相应癌旁组织均禾检测到p53基因突变。结论：mdm2和p53蛋白异常表达与胃癌发生有关，p53基因突变可能并非胃癌组织中p53蛋白异常累积的主要原因，mdm2蛋白在胃癌发生发展中的作用可能与p53蛋白密切相关。     

直肠癌p53、K-ras基因突变与临床病理的相关性研究

目的　探索p5 3、k ras基因同时突变对直肠癌的恶性行为升级的作用及其临床意义。方法　用PCR SSCP方法检测直肠癌细胞p5 3、K ras基因突变 ,分析该基因突变与临床病理因素及预后的关系。结果　直肠癌细胞p5 3、K ras基因突变与临床病理因素无关 ,生存率也无统计学差异。结论　直肠癌细胞p5 3、K ras基因突变对癌细胞恶性生物学行为升级无明显的促进作用 ,p5 3、K ras基因同时突变也无协同促癌作用 ,也不影响病人的预后。     

胃癌组织和腹腔冲洗液p53基因突变的DHPLC技术检测

目的:应用部分变性高效液相色谱(denaturing high performance liquid chromatography, DHPLC)检测胃癌组织、腹腔冲洗液中p53基因突变,并探讨该技术成为检测胃癌腹膜亚临床转移理想方法的可能性.方法: 应用DHPLC对45例胃癌组织及腹腔冲洗液中p53基因突变进行检测,并测序验证.结果:胃癌组织p53基因的突变率为20 0%(9/45),9例突变中有3例突变均未见报道;p53基因在肠型胃癌中突变率35 3%(6/17)明显高于弥漫型胃癌10 7 %(3/28),P＜0 01.腹腔冲洗液中检出p53基因突变2例,检出率为22 2%(2/9),经测序证实分别位于第5 外显子(AAG>AGG,Lys132Arg) 和第6外显子(CTG>CCG,Leu188Pro),均与原发癌组织中相同,这2例患者腹腔冲洗液细胞学检测均为阴性.结论:DHPLC可应用于胃癌患者原发癌灶及腹腔冲洗液中p53基因突变的检测,而且如有效地联合检测多个指标,则可能成为预测胃癌腹膜亚临床转移的理想方法.     

Tumor Protein p53 and K-ras Gene Mutations in Peruvian Patients with Gallbladder Cancer

Background: Recent studies have shown that genetic alterations are associated with the effect of patient geographiclocation on gallbladder cancer development. Peru has a high incidence of gallbladder cancer, but causative factorshave not yet been identified. We examined the frequency of mutations in TP53 and K-ras genes in Peruvian patientswith gallbladder cancer, and compared this with data from Bolivia, Hungary, Chile, and Japan, which have a highgallbladder cancer incidence. Methods: DNA was extracted from formalin-fixed paraffin-embedded gallbladder tissuesections of 30 gallbladder cancer patients (9 men and 21 women) obtained using microdissection. Mutations in exons5 to 8 of TP53 and codons 12, 13, and 61 of K-ras were examined using direct sequencing. Results: TP53 mutationswere observed in 10 (33.3%) of patients, but K-ras mutations were absent. Nine (90%) TP53 mutations were pointmutations (7 missense and 2 silent mutations), and the most frequent substitution was a G:C to A:T transition. G:C toA:T transitions at the CpG site or G:C to T:A transversions were found in one patient each. No significant differenceswere found in the frequency of TP53 and K-ras mutations among patients in the 5 countries. Conclusions: Our findingssuggest that endogenous mechanisms and exogenous carcinogens may affect the carcinogenic process in Peruviangallbladder cancer patients, similar to that in Bolivian patients. Further studies with a larger sample size are neededto clarify these findings.     

High Frequency of TP53 but not K-ras Gene Mutations in Bolivian Patients with Gallbladder Cancer

Although genetic characteristics are considered to be a factor influencing the geographic variation in the prevalence of gallbladder cancer (GBC), they have not been well studied in Bolivia, which has a high prevalence rate of GBC. The purpose of this study was to examine the frequency of TP53 and K-ras mutations in Bolivian patients with GBC and to compare them with our previous data obtained in other high-GBC-prevalence countries, namely Japan, Chile, and Hungary. DNA was extracted from cancer sites in paraffin-embedded tissue from 36 patients using a microdissection technique. TP53 mutations at exons 5 to 8 and K-ras mutations at codons 12, 13 and 61 were examined using direct sequencing techniques. The data obtained were compared with those in the other high-GBC-prevalence countries. Of the 36 patients, 18 (50.0%) had a TP53 mutation (one mutation in each of 17 patients and three mutations in one patient), and only one (2.8%) had a K-ras mutation. Of the 20 TP53 mutations, 12 were of the transition type (60.0%). This rate was significantly lower than that in Chile (12/12, P<0.05). In addition, three mutations were of the CpG transition type (15.0%), which is a feature of endogenous mutation. All three were found in the hot spot region of the TP53 gene. In contrast, G:C to T:A transversion was found in Bolivia, suggesting the presence of exogenous carcinogens. Our findings suggest that the development of GBC in Bolivia is associated with both exogenous carcinogens and endogenous mechanisms. The identification of an environmental risk factor for GBC is needed to confirm these findings.     

血清糖链抗原、粘附分子及K-ras基因突变水平在胰腺癌中的表达及其临床意义

目的 探讨血清糖链抗原、粘附分子及K-ras基因突变联合检测在胰腺癌诊断中的应用价值,及其在胰腺癌中的表达意义.方法 收集109例胰腺癌患者(Ⅰ期14例、Ⅱ期33例、Ⅲ期39例、Ⅳ期23例;淋巴结转移56例,无淋巴结转移53例)、78例胰腺炎患者和98例健康体检者血清.采用化学发光法检测血清CA50、CA125、CA199、CA242水平,采用双抗夹心法酶联免疫吸附试验法检测ICAM、VCAM、ALCAM、CEACAM水平,采用聚合酶链反应法检测K-ras基因突变情况.结果 胰腺癌组CA50、CA125、CA199、CA242、VCAM、ALCAM、CEACAM水平和K-ras突变率明显高于胰腺炎组和健康对照组(P＜0.05);胰腺炎组VCAM、ALCAM、CEACAM水平明显高于健康对照组(P＜0.05),而胰腺炎组和健康对照组CA50、CA125、CA199、CA242水平和K-ras突变率比较,差异无统计学意义(P＞0.05).联合检测[logit (P)=0.114×CA199+0.235×CEACAM+ 0.082×K-ras]对胰腺癌的诊断价值最大,AUC达0.931,显著高于各个指标单独检测,其敏感性和特异性分别达94.5％和86.9％.VCAM、ALCAM、CEACAM水平随胰腺癌分期的增加,水平不断升高(P＜0.05);有淋巴结转移的胰腺癌患者粘附分子VCAM、ALCAM、CEACAM水平均明显高于无淋巴结转移胰腺癌患者(P＜0.05).结论 糖链抗原、粘附分子和K-ras基因突变联合检测能够大大提高胰腺癌的诊断效能,其中粘附分子表达水平与胰腺癌的分期和淋巴结转移密切相关.     

肺癌术后支气管切缘p53基因突变的临床研究

预测肺癌手术切除的效果和局部复发情况。方法滞酶链一单链构象多态吕手术切除标本的无癌支气管切缘和病灶癌组织ｐ５３基因突变的情况。结果１０例支气管切缘有Ｐ５３基因突变,阳性率２５．０％（１０／４０）。２１例病灶癌组织有Ｐ５３基因突变,阳性率５２．５％（２１／４０）。术后全部病例随访３－１４个月。在１０例支气管切缘Ｐ５３阳性者中,有４例发生支气管残端癌复发（复发率４０．０％）,而在３０例阴性者中,无１例     

Clinicopathological and p53 Gene Alteration Comparison between Young and Older Patients with Gastric Cancer

Background: Differences in clinicopathological characteristics of gastric cancer (GC) between young and olderpatients are controversial and a matter of debate. Determining the statistical significance of clinicopathologicalinformation with respect to age might provide clues for better management and treatment of GC. Materials andMethods: A total of 103 Indian GC patients were enrolled for study and specimens were classified according to theAJCC-TNM system. Patients were grouped into two age-wise categories, young patients (<40 years; n=13) andolder patients (≥40 years, n=90). The clinicopathological features of both groups were retrospectively examinedand compared. p53 alterations were analyzed by polymerase chain reaction-single strand conformationalpolymorphism and immunohistochemistry methods at gene and protein levels respectively. The cases wereconsidered p53 over-expressed if it was present in more than 25% of the tumor cells and p53 alterations wascorrelated with the clinicopathological characteristics of the patients as well as etiological factors for GC in bothgroups. Results: We found significant association of young patients with cancer stage (p=0.01), and very strongassociation with histology grade (p=0.064) and poorly differentiated (p=0.051) state of GC. However, neitheryoung nor elderly patients showed associations with location, gender, etiological factors and p53 expressionand alteration. Overall the male-to-female ratio of GC patients was 3.12 and the value was higher in the young(5.5) than in the older group (2.91). Conclusions: Clinicopathological features of GC like cancer stage, celldifferentiation and histological grades were significantly different among young and old age cohorts. We observeda male predominance among the young group that decreased significantly with advancing age. More awarenessof GC onset is required to detect cancer at an early stage for successful treatment.     

胃癌患者p53基因甲基化的研究

作者应用限制性内切酶ＨｐａⅡ和ＭｓｐⅠ酶切胃癌组织及正常胃组织ＤＮＡ，经ＰＣＲ扩增ｐ５３基因第５外显子，琼脂糖凝胶电泳分析其电泳图谱，比较胃癌组织及正常胃组织ｐ５３基因第５外显子特定序列５′－ＣＣＧＧ－３′位点甲基化差异。结果显示：１５例胃癌组织中１２例ｐ５３基因第５外显子出现高甲基化状态，而１０例正常胃组织为低甲基化状态，结果提示，ｐ５３基因高甲基化状态与胃癌发生有关。     

COX-2与p53在非小细胞肺癌中的表达及临床意义

[目的]探讨COX-2和p53蛋白在非小细胞肺癌（NSCLC）中的表达和临床意义。[方法]采用免疫组织化学法检测60例NSCLC组织中COX-2与p53蛋白的表达情况。[结果]COX-2与p53蛋白在NSCLC中阳性表达率分别为56．67％和61．67％。COX-2的表达水平与肺癌的病理类型相关（P=0．009）。COX-2阳性表达的Ⅲ期NSCLC患者中位生存期（MST）和3年生存率均显著低于COX-2阴性表达患者；但p53阳性、阴性表达患者MST和3年生存率无显著性差异。Cox逐步回归分析显示，TNM分期和肿瘤组织COX-2表达水平是NSCLC预后独立因素。[结论]COX-2的高表达与NSCLC患者预后不良相关。     

肺癌患者GSTM1基因型与p53基因突变的相关关系研究

目的:探索肺癌患者Ⅱ相代谢酶主要亚型GSTM1的基因型与p53基因突变的关系.方法:应用双重PCR和PCR-SSCP(单链构像多态性分析)技术,检测了63例肺癌患者和47例健康对照中GSTM1基因缺失及肺癌组织p53基因突变的情况,并分析其相关关系.结果:肺癌组GSTM1基因缺失率为71.4%(45/63),对照组为51.1%(24/47),差异有显著统计学意义,OR为2.40(95%CI为1.09-5.29).p53基因突变率在63例肺癌组织中为49.2%(31/63),在15例对照组织中仅为6.6%(1/18).58.1%肺癌病例同时存在p53基因突变和缺失GSTM1基因(P＜0.05).结论:GSTM1基因缺失与p53基因突变相关,GSTM1基因缺失可能增加p53基因突变的几率,从而导致肺癌患病危险性的增加.     

Prognostic Significance of p53 in Gastric Cancer: a Meta-Analysis

Background: Gastric cancer is one of the frequently seen cancers in the world and it is the second most commonreason for death due to cancer. The prognostic role of expression of p53 detected by immunohistochemistry ingastric cancer remains controversial. This meta-analysis aimed to explore any association between overexpressionand survival outcomes. Materials and Methods: We systematically searched for studies investigating therelationships between expression of p53 detected by immunohistochemistry and prognosis of gastric cancerpatients. Study quality was assessed using the Newcastle-Ottawa Scale. After careful review, survival datawere extracted from eligible studies. A meta-analysis was performed to generate combined hazard ratios foroverall survival and disease-free survival. Results: A total of 4.330 patients from 21 studies were included inthe analysis. Our results showed tissue p53 overexpression in patients with gastric cancer to be associated withpoor prognosis in terms of overall survival (HR, 1.610; 95% CI, 1.394 -5.235; p:<0.001). Pooled hazard ratiofor disease free survival showed that p53 positivity or negativity were not statitistically significant (HR, 1.219;95%CI, 0.782-1.899; p:0.382). Conclusions: The present meta-analysis indicated overexpression of p53 detectedby immunohistochemistry to be associated with a poor prognosis in patients with gastric cancer.