胸苷酸合成酶在膀胱癌组织中的表达及意义

目的探讨膀胱癌（BC）中胸苷酸合成酶（TS）的表达对膀胱癌患者预后的影响。方法采用免疫组化S-P法检测50例膀胱癌石蜡标本中的TS蛋白表达情况。结果TS蛋白在膀胱癌与正常膀胱黏膜中的表达有显著性差异（P＜0．01），与肿瘤低分化、临床病理分期晚期、淋巴结转移密切相关（P＜0．05）。结论TS在膀胱癌中的表达增强可能是膀胱癌患者一个重要的预后指标。     

胃癌组织中胸苷酸合成酶、胸苷磷酸化酶mRNA表达及其临床意义

背景与目的:化学治疗在胃癌治疗中扮演了重要的作用。通过肿瘤组织中某些基因表达的水平来选择化疗药物已成为今后化疗的方向。胸苷酸合成酶(TS)、胸苷磷酸化酶(TP)是氟尿嘧啶(5-FU)体内的关键代谢酶,其表达水平影响恶性肿瘤患者接受5-FU化疗后的预后。本研究探讨胃癌组织中TS、TP mRNA表达水平与预后的关系。方法:采用实时定量RT-PCR技术检测51例胃腺癌组织TS、TP mRNA表达水平。结果:胃癌组织TS、TP mRNA表达水平的中位数分别为0.94和21.20,TS高表达组和低表达组之间无瘤生存期和总生存期差异有显著性(P<0.05),TP高表达组和低表达组之间总生存期差异有显著性(P<0.05),但无瘤生存期之间差异无显著性(P>0.05)。TS、TP mRNA表达与年龄、性别、淋巴结转移、组织学分级及临床分期均无相关性(P>0.05)。结论:检测TS、TP mRNA表达水平对接受5-FU为基础治疗方案的胃癌患者的预后有很好的预测价值。     

胸苷酸合成酶的表达及其与子宫内膜癌的相关性研究

目的　探讨子宫内膜癌(EMC)中胸苷酸合成酶(TS)的表达对子宫内膜癌患者预后的影响。方法　采用免疫组化SP法检测48例子宫内膜癌石蜡标本中的TS蛋白表达情况。结果　TS蛋白在子宫内膜癌与正常内膜中的表达有显著性差异(P <0 .0 1) ,TS表达与肿瘤低分化(P <0 .0 5 )、深层浸润(P <0 .0 5 )、手术-病理分期晚期(P <0 .0 5 )、淋巴结转移(P <0 .0 5 )密切相关。结论　TS在子宫内膜癌组织中的表达增强,可能是子宫内膜癌患者一个重要的预后指标。     

结直肠癌组织胸苷酸合成酶的表达及临床意义

目的:探讨了胸苷酸合成酶(TS)在结直肠癌组织中的表达及其与临床病理特征和预后的关系。方法:用免疫组化SP法检测72例结直肠癌组织中TS的表达,按TS表达水平将患者分为2组:高表达组和低表达组,用Kaplan-Meier法分析TS的表达与临床病理特征和预后的相关性。用多因素COX模型分析影响患者预后的相关因素。结果:在72例结直肠癌组织中TS表达从15%~95%不等,平均(49.79±25.05)%。TS的表达随Dukes分期的增加而增加,但无统计学意义(P=0.145)。早期(Dukes A~B)结直肠癌的预后明显优于晚期结直肠癌(Dukes C、D),P<0.0001。TS低表达者预后优于高表达患者。治疗前血清CEA、CA19-9水平正常者预后较好,CEA、CA19-9升高者预后较差。多因素分析显示Dukes分期、TS表达状况、CEA是影响预后的因素。结论:结直肠癌治疗前Dukes分期是影响预后的主要因素。TS的表达状况、CEA也是影响预后的参考指标。     

胸苷酸合成酶基因多态性与消化道肿瘤个体化治疗

消化道肿瘤的发生率在我国位居第一,而多数患者确诊时即为中晚期,已失去手术的机会,故化疗已成为此类患者的主要治疗手段。虽然以氟尿嘧啶（5-Fu）为基础的联合化疗可以延长胃癌、结直肠癌等晚期消化道肿瘤的生存期,但是接受此类方案中至少50％患者对化疗无效,而且同一化疗方案对不同患者的疗效可从完全缓解到无效,产生差别的主要原因在于个体间对化疗药物敏感性不同所致,近年来的各项研究逐渐表明,TS基因的多态性与患者对化疗药物的敏感性密切相关。     

胸苷酸合成酶、胸苷酸磷酸化酶的表达与培美曲塞时辰化疗的疗效分析

目的构建肺腺癌A549细胞的裸鼠皮下移植瘤模型,研究按昼夜不同时辰给予培美曲塞是否能提高疗效和降低不良反应,分析胸苷酸合成酶（TS）mRNA和胸苷酸磷酸化酶（TP）mRNA表达高低与培美曲塞疗效以及不良反应的关系。方法 常规方法体外培养肺腺癌A549细胞。待肿瘤体积生长至0.5~1.5 cm³,随机将荷瘤鼠分为四组,每组5只,其中三组严格按照每昼夜24 h的三个时间点给药即:早晨7点（光照后0 h）、中午15点（光照后8 h）、晚间23点（光照后16 h）进行腹腔给药,第四组为对照组,腹腔给予同体积的0.9%氯化钠溶液。培美曲塞按照150 mg/kg应用,每天一次,连续4天。以后每周测量瘤体积,裸鼠体重,观察裸鼠的行为状态、精神以及食欲变化。实验结束时称量剥离的裸瘤重量。应用实时荧光定量PCR（Real-Time PCR）检测肿瘤组织中TS mRNA和TPmRNA的表达水平。结果7 h给药组裸鼠体重增长较快（P=0.041）,化疗不良反应较小,瘤体积生长最慢（P=0.032）,抑瘤率最大（P=0.000）。7 h给药组的肿瘤组织中TS mRNA相对表达量最低,TP mRNA的相对表达量对高,差异有统计学意义（P<0.01）。结论培美曲塞对肺腺癌裸鼠移植瘤有明显的抑制作用,疗效及不良反应与时辰给药有关,7 h给药组疗效最佳,化疗不良反应最小;TS和TP可以作为培美曲塞的疗效预测因子,为培美曲塞的个体化临床用药提供参考。     

胸苷酸合成酶预测胃癌化疗敏感性的研究

目的 探讨胃癌组织中TS的表达及其对胃癌DDP+5-Fu化疗敏感性的预测价值.方法 60例胃癌均接受2个周期DDP+5-Fu化疗.化疗方案:DDP 15～20 ms/(m2·d),静脉滴注,d1～5;5-Fu 375～500 mg/(m2·d),静脉滴注8h,d1～5.间隔4周后,进行第2个周期化疗.采用免疫组织化学S-P法检测胃癌组织中TS的表达.结果 60例胃癌中TS高表达者29例,化疗有效率34.5%(10/29),TS低表达者的化疗有效率为64.5%(20/31),两者比较差异具有统计学意义(P<0.05).结论 胃癌组织中TS表达水平对胃癌DDP+5-Fu化疗的敏感性具有一定预测价值.     

胸苷酸合成酶基因3’-UTR多态性与晚期胃癌化疗敏感性关系的研究

目的：研究胸苷酸合成酶（TS）基因3’-UTR多态性与胃癌对5-Fu化疗敏感性的关系。方法：收集经病理学确诊的晚期胃癌106例，所有病例化疗前抽静脉血，提取白细胞DNA，用PCR-RFLP技术检测TS3’-UTR基因型。所有患者均经5-FU为基础的化疗方案治疗。以WHO实体瘤疗效评定标准和毒性评定标准评价疗效和毒性。结果：（1）106例胃癌患者中TS＋6／＋6bp、＋6／-6bp、-6／-6bp基因型频度分别为7．6％、44．3％和48．1％，化疗的总有效率为35．9％。（2）TS＋6／＋6bp、＋6／-6bp、-6／-6bp基因型组化疗的有效率分别为0％（0／8）、40．4％（19／47）和37．3％（19／51）。TS-6／-6bp基因型组和＋6／-6bp组化疗的有效率均显著高于＋6／＋6bp组（Fisher双侧精确检验，P值分别为0．0452和0．0404）。TS-6／-6bp基因型组Ⅱ度以上毒副反应的发生率高于＋6bp／＋6bp基因型组，但其差异无统计学意义。结论：TS基因3’-UTR多态性与晚期胃癌对5-FU为基础的化疗敏感性相关，TS基因型检测有助于指导晚期胃癌的化疗。     

胸苷酸合成酶和谷胱甘肽-S-转移酶在食管癌及其他肿瘤中的应用

胸苷酸合成酶(TS)是抗肿瘤药5-氟尿嘧啶作用的分子靶点,谷胱甘肽-S-转移酶(GST)对顺铂具有解毒作用.研究发现TS、GST的表达与食管癌患者的预后有关,TS和GST表达越高的患者,预后相对较差.TS、GST可为有针对性地选择化疗药物提供依据并成为新的预后因子.     

Polymorphisms in the Thymidylate Synthase Gene and Risk of Colorectal Cancer

To evaluate the relationship between polymorphisms (28 bp repeated sequences in 5’-UTR and 6-bp ins/del in 3’-UTR) in then thymidylate synthetase gene (TS) and risk of colorectal, colon and rectal cancers, weconducted a case-control study with 315 cases of colorectal cancer and 439 population-based controls in Jiangsuprovince, China. TS genotypes were identified using PCR–RFLP (restriction fragment length polymorphism)methods. Odds ratios (ORs) were estimated with an unconditional logistic regression model. We found that thedistributions of 5’-UTR genotypes in TS were significantly different between controls and male colon cases (χ2=8.25, P = 0.016). Compared with 3R/3R genotype, individuals with the 2R allele were at an increased risk ofcolon cancer (age-, BMI-, smoking- and alcohol drinking-adjusted OR=1.98, 95%CI: 1.11-3.53) among men. Inccontrast, the 6-bp ins/del polymorphism at the TS 3’- UTR did not influence risk of the colorectal, colon andrectal cancers. When combined genotypes for both TS 5’-UTR and 3’-UTR polymorphisms were evaluated,individuals with the 5’-UTR 2R allele had a OR of 3.61 (95%CI: 1.38-9.49) for colon cancer among men withthe 3’-UTR –6bp/-6bp genotype. These results show that the polymorphism of the 28 bp repeated sequences inTS 5’-UTR could influence susceptibility to colon cancer and that there was a coordinated effect between TS3’-UTR and 5’-UTR polymorphisms in increasing risk of colon cancer among Chinese men.     

瘦素在非肌层浸润性膀胱癌中的表达及其意义

[目的]探讨瘦素在非肌层浸润性膀胱癌中的表达情况,组织并分析其与膀胱癌患者临床病理特征及预后的关系。[方法]收集112例膀胱移行细胞癌组织和20例正常膀胱组织的病理切片,采用免疫组化方法检测膀胱癌和正常膀胱组织中瘦素的表达。[结果]正常膀胱组织中瘦素的表达水平明显低于肿瘤组织（P=0．032）。在低级别、高级别膀胱癌中,瘦素的阳性表达率分别为43．8％、75．8％,差别有统计学意义（辟0．001）。本组随访6-102个月,其中67例出现复发。复发组与未复发组中瘦素的阳性表达率分别为70．1％和46．7％,差别有统计学意义（P=0．013）。但瘦素表达与患者年龄、性别、肿瘤数目、大小及分期无明显相关性（P〉0．05）。[结论]瘦素表达在膀胱癌组织中明显高于正常膀胱组织,瘦素的检测有助于膀胱癌诊断及预后评估。     

Expression of Thymidylate Synthase in Human Non-small Cell Lung Cancer

5-Fluorouracil (5-FU) has been used worldwide, and the correlation between its effects and thymidylate synthase (TS) expression has been reported in gastrointestinal malignancy. But the significance of TS expression for 5-FU-based chemotherapy has rarely been reported in non-small cell lung cancer (NSCLC). We investigated surgically resected specimens of 23 consecutive patients with previously untreated NSCLC. We used immunohistochemistry and western blot analysis with anti-TS polyclonal antibody to evaluate the existence of TS, and fluorodeoxyuridine-5'-monophosphate (FdUMP) binding assay to evaluate the enzymic activity of TS. We found that 14 samples (60.9%) were positive immunohistochemically, and that the results of immunohistochemistry closely reflected the enzymic activity measured by FdUMP binding assay (ranging from 1.8 to 56.9 pmol/g protein). These results seem to support our experience that 5-FU and its derivatives are clinically significantly effective as a postoperative adjuvant chemotherapy against NSCLC.     

A Novel Approach to Cloning and Expression of Human Thymidylate Synthase

Thymidylate synthase (TS) catalyzes the transfer of a methyl group from methylenetetrahydrofolate to dUMPto form dTMP. It is a primary target in the chemotherapy of colorectal cancers and some other neoplasms. Inorder to obtain pure protein for analysis of structure and biological function, an expression vector TS-pET28b(+) was constructed by inserting wild-type human thymidylate synthase (hTS) cDNA into pET28b (+). Then anexpression strain was selected after transformation of the recombined plasmid into Rosetta (DE3). Fusion proteinwith His-tag was efficiently expressed in the form of inclusion bodies after IPTG induction and the content wasapproximately 40.0% of total bacteria proteins after optimizing expression conditions. When inclusion bodieswere washed, dissolved and purified by Ni-NTA under denatured conditions, the purity was up to 90%. OnSDS-PAGE and West-blotting, the protein band was found to match well with the predicted relative molecularmass-36kDa. Bioactivity was 0.1 U/mg. The results indicated that high-level expression of wild-type hTS cDNAcan be achieved in prokaryotes with our novel method, facilitating research into related chemotherapy.     

Ki67蛋白在104例非肌层浸润性膀胱癌中表达及其意义

[目的]探讨Ki67蛋白表达与非肌层浸润性膀胱癌患者临床病理特征及预后的关系。[方法]收集104例膀胱移行细胞癌和20例正常膀胱组织的病理切片,采用免疫组化法检测Ki67蛋白的表达,分析Ki67蛋白表达与患者临床特征的关系。[结果]膀胱正常组织中Ki67蛋白的表达阳性率明显低于膀胱癌组织（20.0%vs 60.6%,P〈0.001）。G1级、G2级、G3级膀胱癌中,Ki67蛋白表达阳性率分别为36.1%、64.9%和83.9%,差异有统计学意义（P〈0.001）。Ki67蛋白表达与膀胱癌的病理分级、复发及进展有关（P〈0.05）;与患者的年龄、性别、肿瘤数目及肿瘤大小无明显相关性（P〉0.05）。Ki67阴性患者的无复发生存率及无进展生存率优于Ki67阳性患者（P〈0.05）。[结论]Ki67在膀胱癌中高表达,Ki67蛋白的检测有助于判断肿瘤恶性分化程度及监测术后复发及进展。     

Polymorphisms in Thymidylate Synthase and Methylenetetra-hydrofolate Reductase Genes and the Susceptibility to Esophageal and Stomach Cancer with Smoking and Drinking Habits

Thymidylate synthetase (TS) and methylenetetrahydrofolate reductase (MTHFR) are major enzymes in the ‍metabolism of folates, involved in DNA ‘breaks’, instability and hypomethylation.To investigate the possible relations ‍between the TS 3’-UTR and MTHFR C677T polymorphisms and environmental factors impacting on risk of ‍esophageal and stomach cancers, we conducted a case-control study in a high incidence region of China for these ‍cancers. We recruited 138 esophageal and 155 stomach cancer cases, and 223 controls. The TS 3’-UTR and MTHFR ‍C677T genotypes were detected by RFLP assay, using PCR products. The frequency of the -6 bp homozygous TS 3’- ‍UTR genotype was 37.7 % in controls, higher than in Caucasians, although the present distribution was not in ‍Hardy-Weinberg equilibrium. Ever-smoking with the -6 bp/-6 bp TS genotype elevated the ORs (2.61, 1.24-5.49; ‍3.54, 1.60-7.82) for cases of esophageal and stomach cancers, respectively, when compared with never-smoking with ‍the +6 bp/+6 bp and +6 bp/-6 bp genotypes. No combination between the TS and MTHFR genotypes gave increased ‍ORs. The present results suggest that TS polymorphism may modify the risk of esophageal and stomach cancer with ‍smoking, pointing to the necessity for further investigations with information on folate and methionine intake with ‍a larger population. ‍     

Thymidylate synthase as a determinant of pemetrexed sensitivity in non-small cell lung cancer

Background:

Although a high level of thymidylate synthase (TS) expression in malignant tumours has been suggested to be related to a reduced sensitivity to the antifolate drug pemetrexed, no direct evidence for such an association has been demonstrated in non-small cell lung cancer (NSCLC). We have now investigated the effect of TS overexpression on pemetrexed sensitivity in NSCLC cells.

Methods:

We established NSCLC cell lines that stably overexpress TS and examined the effects of such overexpression on the cytotoxicity of pemetrexed both in vitro and in xenograft models. We further examined the relation between TS expression in tumour specimens from NSCLC patients and the tumour response to pemetrexed by immunohistochemical analysis.

Results:

The sensitivity of NSCLC cells overexpressing TS to the antiproliferative effect of pemetrexed was markedly reduced compared with that of control cells. The inhibition of DNA synthesis and induction of apoptosis by pemetrexed were also greatly attenuated by forced expression of TS. Furthermore, tumours formed by TS-overexpressing NSCLC cells in nude mice were resistant to the growth-inhibitory effect of pemetrexed observed with control tumours. Finally, the level of TS expression in tumours of non-responding patients was significantly higher than that in those of responders, suggestive of an inverse correlation between TS expression and tumour response to pemetrexed.

Conclusion:

A high level of TS expression confers a reduced sensitivity to pemetrexed. TS expression is thus a potential predictive marker for response to pemetrexed-based chemotherapy in NSCLC patients.     

Correlation between Thymidylate Synthase Genotype and Susceptibility to Gastric Carcinoma

Objective  To investigate the correlation between polymorphism of the 5′-untranslated region (5′-UTR) of thymidylate synthase genes, as well as the lifestyle, and the susceptibility of gastric carcinoma. Methods  A case-control study, with 60 cases of gastric carcinoma and 170 cases of general risk population-based controls from Nantong, Jiangsu province, China, was conducted. The epidemiological data, such as living habits of the cancer patients, were collected. DNA of peripheral blood leukocytes was obtained from all of the subjects. The TS 5′-UTR tandem repeat genotype was detected using polymerase chain reaction (PCR). Results  There were three TS 5′-UTR genotypes in the group of gastric cancer cases (2R/2R, 2R/3R and 3R/3R) and six TS 5′-UTR genotypes in the group of the controls (2R/2R, 2R/3R, 3R/3R, 2R/4R, 2R/5R and 3R/4R). The genotypic frequencies were respectively 5.0%, 43.3% and 51.7% in the gastric cancer group. Compared with the parameters in the control group, i.e., 4.7%, 31.7%, 60.6%, 1.2%, 1.2% and 0.6%. There were no significant differences between the two groups. Compared with the 3R/3R-genotype individuals who where non-smokers, drank alcohol twice or less each week, drank tea and did not intake pickled food (PF), the risk of gastric cancer significantly went up in the 2R/2R or 2R/3R-genotype people who had habits of smoking, drinking alcohol more than twice each week, no tea drinking but with frequent intake of PF. The adjusted ORs were as follows, 3.79 (95% CI: 2.45∼8.64), 3.41 (95% CI: 1.21∼8.47), 5.99 (95% CI: 3.01∼14.7), and 3.61 (95% CI: 1.81∼8.78). Conclusion  There is an obvious correlation between the polymorphisms of TS 5′-UTR genotypes and the lifestyle of individuals in the development of gastric carcinoma. The work was supported by a grant from Nantong Municipal Bureau of Public Health, Jiangsu Province, China (2006 [No.29]).