阳离子脂质体介导pE-CEA-cd-tk对CEA阳性肺癌的杀伤作用

目的：增强E.coli cd/HSV-1 tk自杀基因的细胞毒性，实现阳离子脂质体介导pE-CEA-cd-tk/5-FC GCV体系靶向杀伤CEA阳性肺癌，方法：PCR法分别扩增出CMV增强子，CWA启动子，cd-tk，构建真核表达载体pE-CEA-cd-tk;MTT法检测pE-CEA-cd-tk/5-FC GCV体系的体外细胞毒性，体内实验采用肺腺癌细胞SPC－A－1裸鼠皮下移植瘤模型，通过肿瘤局部或鼠尾静脉注射脂质体／pE-CEA-cd-tk复合物，腹腔注射GCV＋5－FC前体药物进行治疗。结果：阳离子脂质体介导pE-CEA-cd-tk/5-FC GCV体系体外可靶向杀伤CEA阳性肺癌细胞，这种杀伤作用存在显著的细胞差异；体内可抑制小鼠皮下肺肿瘤结节的生长，荷瘤鼠存活期延长。结论：阳离子脂质体介导pE-CEA-cd-tk/5-FC GCV体系体内外对CEA阳性肺癌均具有较强的杀伤作用，有望用于CEA阳性肺癌的治疗。     

癌胚抗原启动子控制胞嘧啶脱氨酶基因体外对结肠癌细胞专一性杀伤

目的：研究癌胚抗原（CEA）启动子是否能控制大肠杆菌胞嘧啶脱氨酶（EC－CD）基因在CEA阳性结肠癌细胞中专一性表达和杀伤结肠癌细胞。方法：构建由CEA启动子驱动的含EC－CD基因的重组腺病毒载体AdCEACD与由巨细胞病毒（CMV）启动子驱动的含EC－CD基因的腺病毒载体AdCMVCD，分别感染CEA阳性的人结肠癌细胞株Lovo细胞和CEA阴性的Hela细胞，用RT－PCR法检测受染细胞中EC－CD基因的表达，并用MTT法检测感染后细胞对5－氟胞嘧啶（5－FC）的敏感性。结果：Lovo细胞在AdCMVCD与AdCEACD感染后均有EC－CDmRNA表达，且对5－FC的敏感性明显增强，Hela细胞在AdCMVCD感染后有EC－CDmRNA表达，对5－FC的敏感性增强，而在AdCEACD感染后则没有EC－CDmRNA表达，5－FC对其亦无杀伤作用。结论：CEA启动子能够控制EC－CD基因专一性地在CEA阳性的结肠癌细胞中表达，从而实现EC－CD基因的靶向性作用。     

腺病毒介导的KDR启动子-胸苷激酶系统对血管内皮细胞的选择性杀伤作用

目的研究腺病毒介导的KDR启动子-胸苷激酶系统对血管内皮细胞的选择性杀伤作用.方法应用PCR克隆出人KDR基因启动子序列-225 bp～+127 bp,以AdEasy system为载体,构建携带受KDR启动子或CMV启动子调控tk基因表达的2种重组腺病毒质粒pAdKDR-tk和pAdCMV-tk,在293细胞中包装、扩增后,体外感染表达KDR的脐静脉血管内皮细胞系HUVEC和不表达KDR的肝癌细胞系HepG2,并给予不同浓度的丙氧鸟苷(GCV)处理,5 d后收集存活细胞并计数.结果产生的病毒滴度均为5×109pfu/ml.在MOI为100、GCV为50μg/ml条件下,AdKDR-tk转染HUVEC后细胞生存率下降(31.49%±6.42%),AdKDR-tk转染HepG2后细胞生存率为76.57%±3.49%,而转染AdCMV-tk的2细胞系生存率均显著下降,细胞生存率分别为22.24%±3.77%(HUVEC)和26.53%±6.84%(HepG2).结论KDR基因启动子可调控HSV-tk在血管内皮细胞中特异性表达,为进一步开展靶向肿瘤血管内皮的自杀基因治疗研究奠定了基础.     

AF0.3启动子调控的PNP/MeP-dR系统对AFP阳性肝癌细胞的特异杀伤作用

背景与目的:甲胎蛋白(α-fetoprotein,AFP)启动子调控下的目的基因能在AFP阳性肝癌组织中特异性表达;嘌呤核苷磷酸化酶(Escherichiacolipurinenucleosidephosphorylase,PNP)/6-甲基嘌呤-2′-脱氧核糖核苷(6-methylpurine-2-deoxyriboside,MeP-dR)自杀基因系统具有强杀瘤效应。本研究旨在探讨AF0.3启动子调控下的PNP/MeP-dR系统对AFP阳性肝癌细胞的特异性杀伤作用。方法:构建甲胎蛋白启动子AF0.3调控下PNP基因表达载体pAF0.3/PNP,导入AFP阳性肝癌细胞HepG2和阴性的肝癌细胞株SMMC7721,利用G418筛选获得稳定转染PNP基因的HepG2/0.3-PNP及SMMC7721/0.3-PNP细胞。RT-PCR检测二者在细胞中的表达。台盼蓝拒染法检测细胞增殖效应,MTT法和流式细胞仪检测两株细胞对MeP-dR的敏感性及旁观者效应,高效液相色谱法(highperformanceliquidchromatography,HPLC)检测PNP基因产物活性。结果:不论有氧还是缺氧条件,HepG2/0.3-PNP对MeP-dR均较为敏感,SMMC7721/0.3-PNP则对MeP-dR完全不敏感。在任何一种条件下,HepG2/AF0.3-PNP在混合细胞中比例达25%后,就可致明显旁观者效应;而在同样条件下,SMMC7721/0.3-PNP不导致明显的旁观者效应。HPLC结果显示,pAF0.3/PNP在HepG2细胞中可以将MeP-dR转化为6-MP,但其在SMMC7721中则不具有转化活性。结论:AF0.3启动子调控下的PNP/MeP-dR系统对AFP阳性HepG2细胞有较好的杀伤作用。     

荷CEA-rV的DC诱导的CIK对CEA阳性原代肿瘤细胞的特异杀伤作用

目的：探讨携带CEA重组基因痘苗病毒（CEA-rV）抗原的树突状细胞（DC）诱导的杀伤细胞（CIK）对CEA阳性原代肿瘤细胞的特异性杀伤作用。方法：用脐血制备脐血单个核细胞（UBMC）、DC、CIK、DC＋CIK、CEArV＋DC＋CIK和CEA-rV＋DC＋UNBC细胞。取新鲜切除的食管癌组织和肺癌组织，制备原代培养的CEA阳性肿瘤细胞作为靶细胞。用MTT法测5组效应细胞对两种靶细胞的杀伤活性。结果：1）用CEA兔抗人单克隆抗体鉴定证实两种原代培养靶细胞系CEA阳性细胞。2）流式细胞仪测定DC的分子表型，证实为DC。3）流式细胞仪测定CIK的CD3^＋CD56^＋双阳性细胞达75．4％。4）DC＋CIK和CIK相比，对两种CEA阳性肿瘤细胞的杀伤率差异无统计学意义，P〉0．05。5）CE-rV4-DC＋CIK对两种CEA阳性肿瘤细胞的杀伤率（78．80％）明显高于DC＋CIK（52．94％），P％0．05。结论：CEA—rV＋DC能明显提高CIK细胞对原代培养的CEA阳性肿瘤细胞的杀伤活性，而单纯DC不能提高CIK的杀伤活性，说明CEA—rV在提高CIK对CEA阳性肿瘤细胞的特异杀伤作用中起重要作用。     

自杀基因疗法联合化疗对癌胚抗原表达型肺肿瘤的杀伤作用

目的：探讨阳离子脂质体介导E.coli cd/HSV-1tk融合自杀基因联合长春瑞滨对癌胚抗原阳性肺癌的体内杀伤作用。方法：裸鼠皮下接种肺癌细胞SPCA－1后第5、9、13天,腹腔注射阳离子脂质体／pE-CEAcd-tk复合物,于第6－15天连续腹腔注射5－氟胞嘧碇＋丙氧鸟苷前体药物进行自杀基因治疗。于第5、13天给予长春瑞滨进行联合治疗。结果：阳离子脂质体介导融合自杀基因联合长春瑞滨使小鼠皮下的肿瘤结节的生长受到显著抑制,其平均瘤体体积显著小于单独自杀基因或化疗治疗组的平均瘤体体积（P＜0．01）。结论：阳离子脂质体介导E.coli cd/HSV-1tk融合基因联合长春瑞滨化疗在体内在有效抑制肺瘤的生长,为癌胚抗原阳性肺癌的治疗提供了1条新的途径。     

Probasin启动子的克隆及其在不同肿瘤细胞系中的表达活性

目的: 构建Probasin(PB)启动子调控的pGL3表达载体, 比较两种不同组成形式的Probasin启动子的组织特异性和活性。方法：提取大鼠前列腺组织DNA，通过PCR得到Probasin启动子（426，+28bp）序列，并通过交叠PCR得到改造的ARR2PB序列，分别插入pGL3Basic表达载体,同时构建CMV     

腺病毒介导HSV—tk自杀基因联合野生型p53基因对直肠癌细胞的杀伤作用

目的 观察重组腺病毒介导单纯疱疹病毒胸苷激酶（HSV－tk)基因联合人野生型p53基因共转染,对直肠癌细胞杀伤作用。方法 构建重组腺病毒质粒pAdCMV-Link1(tk/p53)、pAdCMV-Link(tik),pAdCMV-Link(p53),分别感染p53炎变的人直肠癌细胞SW837。进行细胞集落形成实验、细胞存活率的测定和裸鼠移植瘤治疗实验,观察HSV－tk/GCV系统与野生p53基因联合对肿瘤细胞的杀伤作用。结果 应用pAdCMV-Link1(tk/p53),pAdCMV-Link(-),pAdCMV-Link(tk),pAdCMV-Link(p53)重组腺病毒感染SW837细胞,加入GCV,各组细胞集落数分别为8、95、40、70。pAdCMV-Link(CD/53)组肿瘤细胞集落形成减少、细胞存活率显著下降（P＜0．01）。裸鼠移植肿瘤生长抑制率分别为76．5％、0．8％、55．8％、23．2％,pACMV-Link(tk/p53)重组腺病毒对肿瘤的抑制作用最强。结论 HSV－tk自杀基因与野生型p53基因共转染,对肿瘤细胞有更强的杀伤作用。     

转录靶向性KDR启动子调控双自杀基因系统对人大细胞肺癌细胞L9981的杀伤作用

背景与目的 研究KDR启动子转录调控双自杀基因系统(CDglyTK)对人大细胞肺癌细胞L9981的杀伤作用.方法 应用分子生物学技术克隆KDR基因的启动子KDRP,构建KDR启动子调控的双自杀基因(CDglyTK)真核表达质粒pcDNA3-KDRp-CdglyTK.并将其导AL9981和人肝癌细胞HepG2中.给予不同的前药(GCV和/或5-FC)处理后,应用MTT法检测各组细胞的存活率.并应用流式细胞仪检测各组细胞的细胞周期和凋亡.结果 成功的克隆KDR启动子和构建pcDNA3-KDRp-CdglyTK质粒.pcDNA3-KDRp-CdglyTK转染后,L9981细胞中均可检测到CD和TK mRNA,而HepG2细胞中则无.联合应用5-FC+GCV处理对转双自杀基因细胞(L9981)的杀伤作用显著高于单独应用5-FC或GCV(P<0.05),且二者显示了良好的药物协同作用.结论 KDR基因启动子调控双自杀基因系统可以靶向性杀伤人肺癌细胞.     

HSV-TK/GCV体系对前列腺癌PC-3m细胞杀伤作用的实验研究

目的：探讨HSV－TK／GCV体系对前列腺癌PC-3m细胞的杀伤作用。方法：应用逆转录病毒载体将HSV－TK基因转染PC－3m细胞，经RT－PCR鉴定后，MTT、流式细胞仪、电镜等方法检测丙氧鸟苷（GCV）对转染后PC－3m细胞的杀伤作用，同时以未转染PC－3m细胞为对照。结果：GCV对正常PC－3m细胞毒性较低，而对转染后PC－3m细胞有较强的细胞毒作用，但旁观者效应不明显。结论：HSV－TK／GCV体系对前列腺癌细胞具有明显杀伤作用，有必要进一步研究。     

ARE/SUZ12 dual specifically-regulated adenoviral TK/GCV system for CML blast crisis cells

Background

Treatment of blast phase chronic myeloid leukemia (BP-CML) remains a challenge, and the median survival is less than 6 months. Because effective treatments are lacking, we studied tight targeting of blast crisis CML cells using adenoviral (Ad) vectors expressing a HSV-TK system under dual control of a specific SUZ12 promoter and an antioxidant response element (ARE).

Methods

A potential SUZ12 promoter fragment was designed with bioinformatics databases and identified with a luciferase assay. Next, we cloned the ARE element of the NQO1 gene and developed Ad vectors expressing TK kinase or luciferase under the dual control of a specific SUZ12 promoter and an ARE element. An in vitro transfection assay with Ad-ARE/SUZ12-Luc was used to determine promoter activity of ARE/SUZ12 regulatory element in blast crisis CML cells. After incubating human BP-CML-derived cells with Ad-ARE/SUZ12-TK and ganciclovir, Western blot, CCK8, Immunofluorescent assays and Annexin V assays were conducted to assess the efficacy of an ARE/SUZ12 dual-specific TK/GCV system for BP-CML cell lines.

Results

Here, luciferase data confirmed significantly higher and specificer promoter activity of the ARE/SUZ12 composite component in CML blast crisis-derived cell lines (K562, KCL22, and K562/G01) compared to HepG2 cells, and Ad-AS-TK/GCV system could exhibit enhanced apoptotic effects and decreased cell viability for BP-CML cell lines. Additionally, Ad-AS-TK/GCV system altered expression of cycle-related and apoptosis-related proteins in BP-CML cell lines.

Conclusions

Thus, ARE/SUZ12 dual targeting TK/GCV system was effective in killing BP-CML cells. Moreover, efficacy and specificity of CML cell eradication were enhanced by synergistic effects of ARE/SUZ12 dual-specific regulation. We conclude that suicide gene-targeted therapy might hold promise for BP-CML treatment.

Electronic supplementary material

The online version of this article (doi:10.1186/s13046-015-0139-4) contains supplementary material, which is available to authorized users.     

肺腺癌组织特异性自杀基因治疗实验研究

目的 探讨肺腺癌组织特异性自杀基因治疗的安全性及有效性。方法 采用病毒感染法，将癌胚抗原(CEA)基因启动子所驱动的CD基因的组织特异性逆转录病毒载体(G1CEACDNa)，导入分泌CEA的肺腺癌细胞系A549细胞．研究裸鼠体内抑瘤效果；应用重组逆转录病毒裸鼠体内治疗A549肿瘤，观察G1CEACDNa／5-氟胞嘧啶(5-FC)对A549细胞致瘤裸鼠的治疗作用及毒副反应。结果 (1)将转基因的A549细胞和未转基因的A549细胞接种至裸鼠皮下．两者成瘤性无明显差异；(2)在转基因细胞致瘤裸鼠实验中，5-FC对转CEA启动子调控自杀基因的肿瘤生长具有明显的抑制作用；(3)将G1CEACDNa重组逆转录病毒上清直接注射到裸鼠成瘤部位．然后腹腔内注射5-FC同样获得明显的抑瘤效果；(4)与直接注射5-FU相比，组织特异性自杀基因治疗对骨髓的抑制明显降低。结论 组织特异性自杀基因治疗可能成为肿瘤治疗个体化的重要方法之一。     

应用AdEasier-1系统高效制备含VEGFP驱动TK自杀基因的重组腺病毒

背景及目的:自杀基因疗法的瓶颈之一是自杀基因在肿瘤细胞的特异性表达水平低。利用肿瘤特异性启动子来调控自杀基因使其在肿瘤细胞中特异性表达已成为肿瘤自杀基因研究的热点。研究证实血管内皮生长因子在绝大多数实体瘤细胞中都有过量表达而在正常组织中不表达或表达甚微,其过量表达与血管内皮生长因子启动子(vascularendothelialgrowthfactorpromoter,VEGFP)活性上调有关。为研究VEGFP可否提高TK自杀基因在肿瘤细胞的特异性表达水平,本研究应用一种高效的重组腺病毒构建系统,即AdEasier-1系统制备含VEGFP驱动TK自杀基因的重组腺病毒。方法:VEGFP自pEGFP-1-SV-VEGFP切下后插入pAdtrack载体上构建pAdtrack-VEGFP。用HindⅢ/XbaⅠ自pREP8-TK切出带polyA加尾信号的TK基因,亚克隆到pAdtrack-VEGFP中构建转移质粒pAdtrack-VEGFP-TK。PmeⅠ酶线性化转移质粒pAdtrack-VEGFP-TK,转化含腺病毒基因组质粒pAdEasy-1的细菌AdEasier-1,用25μg/ml卡那霉素筛选阳性克隆,先后进行琼脂糖电泳和PacⅠ酶切鉴定,筛选出正确重组腺病毒质粒pAdEasy-VEGFP-TK,经293细胞包装、扩增获得重组腺病毒Ad-VEGFP-TK,行酶切鉴定、PCR鉴定和测序。结果:卡那霉素抗性细菌只有两种,一种含pAdEasy-VEGFP-TK(大于33kb),另一种含pAdtrack-VE     

腺病毒介导HSV-TK/GCV系统联合放射治疗口腔鳞癌的研究

目的 探讨单纯疱疹病毒胸苷激酶基因（HSV－TK）／丙氧鸟苷（GCV）自杀基因系统对放射治疗的增敏作用。方法 口腔鳞癌细胞（Tca8113细胞系）经HSV－TK／GCV系统治疗后给予放射治疗，采用LQ和单击多靶（SHMT）模型分析细胞存活曲线参数。结果 细胞存活曲线分析显示：单纯放射治疗组其α为0.1074,β为0.0158,D0为2.2576,Dq为3.5413；与单纯放射治疗组比较，HSV－TK／GCV治疗组α（0.2127)和α：β（9.496)值大，D0（1.4526)和Dq(2.2257)值小，其放射增敏率（SER）为1.55。结论 HSV－TK／GCV系统具有放射增敏作用，可提高放射治疗对口腔鳞癌的治疗疗效。     

Construction of double suicide genes system controlled by MDR1 promoter with targeted expression in drug-resistant glioma cells

The multiple drug resistance protein (MDR1) is frequently overexpressed in human glioma. The aim of this study is to clone the MDR1 promoter from C6/ADR, construct the double suicide genes expressive vector controlled by MDR1 promoter, and explore its targeted expression in C6/ADR cells. MDR1 promoter from C6/ADR genomic DNA, which was linked with T vector, was amplified by using Polymerase chain reaction (PCR). After cut by NdeI and HindIII, MDR1 promoter was cloned into pcDNA3-TK (thymidine kinase) plasmid. The cytosine deaminase (CD) gene from pcDNA3-CD-TK plasmid was directly cloned into the above vector to construct pcDNA3-MDR1-promoter-CD-TK vector. Then this vector was transfected into C6 and C6/ADR cells respectively by liposome. After selection by G418, the tumor cell lines were stably established. Then these cell lines were examined through PCR and RT-PCR to respectively detect the integration and expression of TK and CD genes. The results showed the length and sequence of MDR1 promoter amplified by PCR were confirmed by DNA sequencing. The pcDNA3-MDR1-promoter-CD-TK expression vectors were constructed successfully. PCR indicated the double suicide genes were integrated into C6 and C6/ADR cells. RT-PCR reveled that CD and TK genes expressed in C6/ADR/CD-TK cells, whereas not in C6/CD-TK cells. In conclusions, construction of expressive vector containing double suicide genes controlled by MDR1 promoter with targeted expression in C6/ADR will provide a sound basis for targeted gene therapy for multidrug resistance (MDR) glioma.     

Exogenous wt-p53 enhances the antitumor effect of HSV-TK/GCV on C6 glioma cells

Objective To study on the antitumor effect of combining wt-p53 gene with suicide gene therapy (HSV-tk+GCV) for malignant gliomas. Methods AdCMV-p53 was transfected into C6 glioma cells at MOI of (Multiplicity of infection) 0(G100), 10(TPG1), 100(TPG2), then AdCMV-tk was transducted to C6 glioma cells of G100, TPG1 and TPG2, respectively, at MOI of 100. The C6 glioma cells tranfected with both AdCMV-p53 and AdCMV-tk were exposed to various concentration of GCV. The cell survival rate was measured by MTT assay in vitro. Rat glioma model was established by injecting 5 × 10⁵ C6 glioma cells into right caudate nucleus of SD rats. AdCMV-p53 and AdCMV-tk were injected into glioma on day 5 and 6, respectively. On day 7, ganciclovir (GCV) was administrated intraperitoneally at 15 mg/kg/day for 14 days. The survival time of all rats was observed. The growth of intracerebral tumors was monitored dynamically by enhanced MRI. Cell apoptosis was evaluated by TUNEL method. Expression of HSV-tk gene was identified by in situ hybridization and expression of exogenous p53 gene was detected with Western blotting. Results In vitro, wt-p53 significantly enhanced antitumor effect of HSV-tk/GCV. The concentration of GCV for ID50 of TPG2 cells (0.001 μg/ml GCV) was 10 times lower than that for the cells of tk-GCV group (MOI = 100), while the concentration of GCV for ID100 of TPG2 (0.01 μg/ml GCV) and TPG1(0.1 μg/ml GCV) was 100 and 10 times lower than that for the cells of tk-GCV group (MOI = 100), respectively. Apoptosis of C6 glioma cells also could be induced by transfection with wt-p53 gene slightly. For in vivo study, the survival time of tumor-bearing rats treated with HSV-TK/GCV or wt-p53 combined with HSV-TK/GCV was significantly prolonged and the intracerebral tumors were regressed and disappeared earlier in the combined gene therapy group than those in the HSV-TK/GCV therapy group as shown in enhanced MRI. However, only half dose of GCV for the rats treated with both wt-p53 and HSV-TK/GCV was needed to obtain the same efficacy as those rats treated with HSV-TK/GCV alone. These results indicate that the transfection of wt-p53 potentiates the effect of HSV-TK/GCV therapy. Conclusions The combination of HSV-tk/GCV system with wt-p53 gene transduction is optimal for clinical therapeutic trials of suicide gene therapy for malignant gliomas.     

HSV—tk／GCV系统对神经胶质瘤的自杀基因治疗研究

目的在ｉｎｖｉｔｒｏ和ｉｎｖｉｖｏ水平建立致死神经胶质瘤的ＨＳＶ－ｔｋ／ＧＣＶ自杀基因系统,并验证该系统的有效性。方法用携带单纯疱疹病毒胸苷激酶（ＨＳＶ－ｔｋ）基因的重组逆转录病毒,感染大鼠神经胶质瘤细胞Ｃ６,筛选稳定表达ｔｋ的克隆细胞Ｃ６／ｔｋ;经生长抑制试验,比较Ｃ６／ｔｋ细胞对三种核苷类似物ＧＣＶ、ＢＶｄＵ和ＡＣＶ的敏感性;Ｃ６／ｔｋ细胞在裸鼠皮下成瘤,用ＧＣＶ进行治疗并观察疗效。结果ｔｋ基因整合入Ｃ６细胞并在Ｃ６／ｔｋ细胞中稳定表达;生长抑制试验表明,ＧＣＶ是最为有效的原药,Ｃ６／ｔｋ细胞对ＧＣＶ高度敏感,ＩＣ５０＜０．２μｍｏｌ／Ｌ,而野生型Ｃ６细胞和转染空载病毒载体的Ｃ６／０细胞对ＧＣＶ的ＩＣ５０≥１００μｍｏｌ／Ｌ,相差５００多倍。裸鼠ｉｎｖｉｖｏ实验得到相应结果,治疗组肿瘤受到明显抑制和杀伤。结论在ｉｎｖｉｔｒｏ和ｉｎｖｉｖｏ水平,表达ｔｋ基因的肿瘤细胞均被ＧＣＶ有效杀伤,这表明ＨＳＶ－ｔｋ／ＧＣＶ自杀基因系统有可能成为基因治疗脑瘤的有效方法。     

pGL3-hTERT-tk/GCV对胃癌细胞的促凋亡作用

目的:探讨重组质粒pGL3hTERTtk/GCV对胃癌细胞的促调亡作用。方法：以基因工程方法构建重组质粒pGL3hTERTtk和相应的荧光报告质粒pGL3hTERTtkLuc+;脂质体LipofectamineTM2000瞬时转染胃癌细胞系SGC7901并用GCV干预,荧光显微镜观察细胞形态变化和转染效率,TUNEL标记和流式细胞术观察转染后胃癌细胞的凋亡;以上实验均以正常肝细胞L02为对照。结果：经鉴定,重组质粒pGL3hTERTtk中tk片段的长度为1 100 bp。荧光素酶标记的阳性、阴性对照及治疗报告质粒pGL3hTERTtkLuc+均能有效转染高表达端粒酶活性的胃癌细胞SGC7901,转染效率为(8.2±114)%。重组质粒转染胃癌细胞后与GCV共育4 d,细胞的凋亡率为(60.0±1.56)%;被pGL3hTERTtk转染的肿瘤细胞细胞周期发生了变化,处于细胞周期早期的细胞大量凋亡,早期凋亡率为（47.1±1.35）%。〖HT5W〗结论：〖HT5"SS〗pGL3hTERTtk/GCV对胃癌细胞有强烈的杀伤作用,但不影响正常细胞的生长,有潜在临床应用前景。