p53基因增加肿瘤放射治疗敏感性的机制

在肿瘤放疗过程中,p53基因的正常功能对肿瘤细胞的凋亡和提高放射敏感性起了关键性作用。野生型p53基因通过激活或抑制一系列基因,使肿瘤细胞周期阻滞,抑制肿瘤细胞放射损伤的修复,促进肿瘤细胞的凋亡,以及在乏氧环境中对促进肿瘤细胞凋亡至关重要,增强了肿瘤细胞对放疗的敏感性。     

p53基因在宫颈癌发病中的意义及对放射敏感性的影响

放疗在宫颈癌治疗中具有重要地位,但放疗抵抗导致局部未控仍常见,因此预测放射敏感性对制定个体化治疗策略是非常有益的。影响放射敏感性的因素有很多,p53状态是重要因素之一,p53基因上下游信号分子的调控亦能影响宫颈癌放射敏感性。本文分析了p53基因在不同宫颈癌中的差异,并探讨p53调控放射敏感性的可能方式。     

PUMA——非常有前景的肿瘤基因治疗靶点

PUMA(p53上调凋亡调控因子)是2001年发现的Bcl-2蛋白家族中的一员,是p53的下游基因,因为具有强大的促凋亡作用而备受关注.肿瘤组织中的PUMA可被化疗药物诱导而表达增加,同时,PUMA诱导多种肿瘤细胞凋亡,抑制肿瘤细胞增殖,而且,PUMA与有化疗和放疗有协同作用,增加肿瘤细胞对放化疗的敏感性.PUMA的作用与p53状态无关,是非常有前景的肿瘤基因治疗靶点.     

p53基因改变对人体鼻咽癌放射敏感性的影响

很久以来,肿瘤的病理类型、分化程度、乏氧等在临床上被看作是影响肿瘤放射敏感性的重要因素。近来一些文献报道细胞内某些癌基因(如ras、myc、raf等)的改变至少对肿瘤细胞的放射敏感性有影响。作者对既往在中国医科院肿瘤医院接受过放疗的部分鼻咽癌治疗前标本中p53基因的表达情况进行了检测,以了解p53基因改变对体内肿瘤放射敏感性的影响。1　材料和方法　　随机调取154例鼻咽癌患者治疗前活检的福尔马林固定、石蜡包埋标本,采用p53单克隆抗体DO-1(鼠抗人)和免疫组化染色试剂盒LSAB(日本DAKO公司)检测肿瘤组织中p53基因过量表达。其中…     

p21/WAF-1/CIP-1与宫颈癌放射敏感性的关系

已知凋亡受多种基因调控，其中p21／WAF-1／CIP-1是近年来研究较多的基因，又简称p21。它通过作用于周期蛋白．周期蛋白依赖性激酶(cyclin-CDK)复合物及增殖细胞核抗原(proliferating cell nuclear antigen，PCNA)引起肿瘤细胞生长抑制及调控细胞凋亡，从而影响肿瘤的放射敏感性。笔者搜集了30例宫颈癌病例，通过研究细胞凋亡与肿瘤放射敏感性以及p21与细胞凋亡的关系，探讨p21对宫颈癌放射敏感性的作用。     

E2F基因与肿瘤放射敏感性研究进展

目的 肿瘤放射敏感性是影响肿瘤放射治疗效果的重要原因.在众多影响放射敏感性的因素中,细胞周期、细胞凋亡和DNA损伤修复发挥重要作用.E2F基因家族通过编码E2Fs转录因子家族,调控细胞周期、细胞凋亡等生命活动.本研究旨在总结E2F基因与肿瘤细胞放射敏感性的关系的研究进展.方法 应用PubMed、CNKI等数据库,以“E2F,放疗敏感性,辐射敏感性”等为关键词,检索1990-12-2017-05的中英文文献,共检索到英文文献1 560篇,中文文献634篇.纳入标准:(1)E2F的结构其对细胞周期,细胞凋亡,DNA损伤调控的相关研究;(2)肿瘤细胞辐射敏感性影响因素的临床、基础研究;(3)E2F基因与肿瘤放射敏感性关系的基础研究.排除标准:(1)讨论与辐射敏感性因素关系不紧密的研究;(2)探究不与E2F基因相互作用的基因和细胞因子的研究.符合分析的文献126篇,72篇纳入分析.结果 E2F基因通过PRb/E2F通路调控G1/S期转变,进而调控细胞周期进程.细胞DNA损伤后,E2F1通过p53依赖型和非p53依赖型方式诱导细胞凋亡,成为肿瘤放射治疗的潜在靶点.结论 探究E2F基因与肿瘤放射敏感性的关系将可能成为未来提高肿瘤放射敏感性的重要思路.     

腺病毒ElA基因研究进展

腺病毒(Ad)早期基因(E1A)具有双向调控多种基因转录的作用,并且作为新发现的一种抗癌基因而受到广泛关注.E1A通过提高p53的表达,抑制HER-2/neu的转录来促进肿瘤细胞的凋亡,同时其可以提高CTL细胞、NK细胞、巨噬细胞的杀伤效应,提高肿瘤细胞对化疗、放疗的敏感性.本文就E1A的研究进展做一综述.     

乏氧诱导因子-1对肿瘤放射敏感性影响的机制

肿瘤的放射敏感性与放射治疗疗效密切相关,而肿瘤内乏氧又被认为对放射敏感性有重要的影响。乏氧诱导因子-1（HIF-1）是调节细胞适应乏氧的关键转录因子,通过维持肿瘤细胞代谢、促进肿瘤血管形成、影响细胞增殖和凋亡而影响放射敏感性。通过抑制HIF-1和（或）其下游基因可以提高肿瘤放射敏感性,是一条具有潜力的放疗增敏途径。     

乏氧诱导因子-1对肿瘤放射敏感性影响的机制

肿瘤的放射敏感性与放射治疗疗效密切相关,而肿瘤内乏氧又被认为对放射敏感性有重要的影响.乏氧诱导因子-1(HIF-1)是调节细胞适应乏氧的关键转录因子,通过维持肿瘤细胞代谢、促进肿瘤血管形成、影响细胞增殖和凋亡而影响放射敏感性.通过抑制HIF-1和(或)其下游基因可以提高肿瘤放射敏感性,是一条具有潜力的放疗增敏途径.     

宫颈癌放疗前后肿瘤细胞凋亡及其相关基因的变化

目的:探讨宫颈癌组织放疗前后肿瘤细胞凋亡及凋亡调节基因p53、bcl-2和bax蛋白表达的变化及意义.方法:选择未经治疗的宫颈癌患者20例为试验对象,采集放疗前和放疗10Gy后宫颈癌组织标本,用原位DNA切口末端标记(TUNEL)法检测凋亡细胞;单克隆抗体免疫组化SABC法检测细胞凋亡相关基因p53、bcl-2和bax的蛋白表达水平.结果:1)在宫颈癌放疗前后,肿瘤细胞凋亡阳性率和平均凋亡指数分别为25.00%和0.11%、75.00%和3.40%,放疗前后有显著性差异(P＜0.01);2)放疗前肿瘤细胞凋亡相关基因p53、bax和bcl-2蛋白表达阳性率分别为55%、10%和20%,而放疗后分别为25%、60%和25%,放疗后p53蛋白表达减少,bax蛋白明显增加,bcl-2蛋白无明显变化.结论:放射治疗诱导宫颈癌肿瘤细胞凋亡,可能与凋亡调节基因bax基因表达的诱导密切相关.     

抑癌基因p53对人胃癌细胞系放射敏感性的作用

目的评价野生型抑癌基因（正常）ｐ５３对人胃癌细胞系放射敏感性的控制作用。方法用流式细胞仪分析４Ｇｙ照射后８和２４小时４种不同ｐ５３状态的人胃癌细胞系细胞周期分布和凋亡的反应。以４Ｇｙ细胞存活份数和１０Ｇｙ照射后的肿瘤生长曲线比较４种细胞的放射敏感性。结果照射４Ｇｙ后８小时和２４小时的ｐ５３正常的ＢＧＣ８２３细胞出现强烈的Ｇ１期阻滞（分别占原细胞总数的６７．９％和６１．１％）,比无照射的该细胞Ｇ１期比例有显著的增高（Ｐ＜０．０５）,并出现明显的预示凋亡的亚Ｇ１峰（ＳｕｂＧ１）,凋亡细胞比例分别达１３．０％和１５．３％;同样条件下其他３种ｐ５３异常的细胞Ｇ１期比例没有显著的变化（Ｐ＞０．０５）,都没有出现亚Ｇ１峰,凋亡细胞比例均为０．０％。ｐ５３正常的ＢＧＣ８２３细胞４Ｇｙ的存活份数明显低于其它３种细胞（Ｐ＜０．０５）;且细胞移植肿瘤１０Ｇｙ照射后比其它３种的生长受到更明显抑制（Ｐ＜０．０５）。结论以上的结果证实了野生型ｐ５３基因促进了照射后肿瘤细胞的Ｇ１期阻滞和凋亡,从而明显地提高肿瘤细胞的放射敏感性。     

Down-regulation of Wilms’ tumor 1 expression in glioblastoma cells increases radiosensitivity independently of p53

The Wilms’ tumor 1 (WT1) gene is overexpressed in human glioblastoma and correlates with wild-type p53 status. In other cell types, WT1 inhibits p53-mediated apoptosis in response to DNA damaging agents. However, neither this interaction nor the relationship between WT1 and radiosensitivity has been studied in glioblastoma. To study this interaction, we generated LN-229 glioma cell lines (p53 mutant) stably expressing WT1 isoforms and induced apoptosis by transfecting with different doses of wild-type p53 plasmid expression vector. Constitutive expression of WT1 did not protect against exogenous p53-mediated apoptosis. Likewise, WT1 expression did not protect against endogenous p53-mediated cell death induced by radiotherapy in U87MG cells, which contain functional wild-type p53. We then tested the efficacy of WT1 siRNA in inhibiting WT1 expression and its effect on radiosensitivity. In T98G and LN-18 glioma cells, which possess p53 mutations, WT1 siRNA decreased WT1 protein to almost undetectable levels by 96-h post-transfection. Furthermore, WT1 siRNA transfection caused a significantly larger decrease in viability following irradiation than was seen in untransfected cells in both cell lines after treatment with ED₅₀ of ionizing radiation. In conclusion, WT1 overexpression did not protect against p53-mediated apoptosis or ionizing radiation induced cell death. WT1 siRNA increased the radiosensitivity of two human glioma cell lines independently of p53. Anti-WT1 strategies may, therefore, prove useful in improving the response of glioblastoma to radiotherapy, thus potentially improving patient survival.     

吉西他滨联合X射线照射对肺癌细胞系放射增敏作用的初步研究

目的 探讨吉西他滨(GEM)是否对非小细胞肺癌具有放射增敏作用,并对GEM的放射增敏机制进行初步探讨.方法 用克隆形成分析法观察GEM对p53基因突变的人肺腺癌细胞系(973细胞)的放射增敏效应.流式细胞术观察照射前后973细胞周期分布和细胞凋亡,分析其与p53基因突变是否为放射增敏机制.结果 10 nmol/L GEM照前、照后给药均具有极轻微放射增敏作用;100 nmol/L GEM照前、照后给药时均具有明显放射增敏作用,且照前给药组的增敏作用明显强于照后给药组.p53基因突变影响细胞周期再分布及细胞凋亡,但与GEM的放射增敏作用无关.结论 100 nmol/L GEM具有明显放射增敏作用,p53基因突变、细胞周期再分布及细胞凋亡不是GEM放射增敏作用的主要机制.     

EFFECT OF ADENOVIRUS—MEDIATED p53 GENE TRANSFER ON APOPTOSIS AND RADIOSENSITIVITY OF HUMAN

To evaluate the effect of adenovirus-mediated p53 gene(Adp53) on apoptosis and radiosensitivity of human gastric carcinoma cell lines.Methods:Recombinant adenovirus expressing wild-type p53 lines with different p53 genetic status.p53 protein expression was detected by immunohistochemistry assay and western blot assay.Cell survival was assessed using a clonogenic assay.TUNEL assay was used in determination of apoptosis.Four human gastric carcinoma cells infected with Adp53 were irradiated with 4Gy and cell cycle distribution and Sub-G1 peak were assayed by flow cytometry.Results:G2/M arrest,apoptosis and inhibition of tumor cell proliferation were induced by infection at Adp53 at 100 MOI which caused high transfer rate of wild-type p53 and strong expression of p53 protein in four human gastric carcinoma cells.The radio-enhancement ratio of Adp53 at 4Gy were3.0 for W cell,3.6 for M cell,2.2 for neo cell and 2.5 for 823 cell in vitro.Conclusion :This study demonstrated that Adp53 transfer increased cellular apoptosis and radiosensitivity of human gastric carcinoma cell lines in vitro independently on cellular intrinsic p53 status thus supporting the combination of p53 gene therapy with radiotherapy in clinical trials.     

p53-dependent G2 arrest associated with a decrease in cyclins A2 and B1 levels in a human carcinoma cell line

In vivo transfer of wild-type (wt) p53 gene via a recombinant adenovirus has been proposed to induce apoptosis and increase radiosensitivity in several human carcinoma models. In the context of combining p53 gene transfer and irradiation, we investigated the consequences of adenoviral-mediated wtp53 gene transfer on the cell cycle and radiosensitivity of a human head and neck squamous cell carcinoma line (SCC97) with a p53 mutated phenotype. We showed that ectopic expression of wtp53 in SCC97 cells resulted in a prolonged G1 arrest, associated with an increased expression of the cyclin-dependent kinase inhibitor WAF1/p21 target gene. A transient arrest in G2 but not in G1 was observed after irradiation. This G2 arrest was permanent when exponentially growing cells were transduced by Ad5CMV-p53 (RPR/INGN201) immediately after irradiation with 5 or 10 Gy. Moreover, levels of cyclins A2 and B1, which are known to regulate the G2/M transition, dramatically decreased as cells arrived in G2, whereas maximal levels of expression were observed in the absence of wtp53. In conclusion, adenoviral mediated transfer of wtp53 in irradiated SCC97 cells, which are mutated for p53, appeared to increase WAF1/p21 expression and decrease levels of the mitotic cyclins A2 and B1. These observations suggest that the G2 arrest resulted from a p53-dependent premature inactivation of the mitosis promoting factor.     

Expression of P53, P21/WAF/CIP,BCL-2, BAX,BCL-X,and BAK In Radiation-Induced Apoptosis in Testicular Germ Cell Tumor Lines

Purpose: Testicular germ cell tumors (TGCTs) represent one of the few tumor types that are curable by antineoplastic therapy, probably due to the high sensitivity of this neoplasm to induction of apoptosis by chemotherapeutic agents and/or ionizing radiation. Here, we tested cell susceptibility to radiation-induced apoptosis in a panel of TGCT cell lines and attempted to correlate this with the known potentially relevant molecular determinants (p53 gene status and Bcl-2 family proteins) of apoptosis.Methods and Materials: Induction of apoptosis by γ-radiation was morphologically recognized in NT2, NCCIT, S2, and 2102 EP using Hoechst/PI staining and additionally confirmed by Western blot analysis of PARP cleavage. The p53 gene status was estimated by sequence analysis. Expression of p21/WAF/CIP was determined by Northern blot analysis and immunoblotting was used to monitor p53, Bax, Bcl-2, Bcl-x, and Bak protein levels. In vitro colony formation was studied to establish clonogenic survival curves.Results: NT2 and NCCIT appeared to be susceptible for radiation-induced apoptosis, contrasting 2102 EP and S2 which were highly resistant. Sequence analysis showed that NT2, S2, and 2102 EP are homozygous for wild-type p53 (wtp53), whereas NCCIT contains mutant p53 (mtp53). NT2 and 2102 EP cells showed radiation-induced p53 upregulation, while NCCIT (mtp53) and S2 (no p53 protein) cells did not. Consistently, γ-radiation-induced DNA damage resulted in a p53-dependent transactivation of the p21/WAF/CIP gene in NT2 and 2102 EP, but not in mtp53-containing NCCIT cells and p53 nonexpressing S2 cells. Constitutive expression of Bax, Bcl-2, Bcl-x, and Bak was not affected by radiation and showed no correlation with cell susceptibility to radiation-induced apoptosis. A discrepancy was found between apoptosis and reproductive death.Conclusions: The present study revealed that: i) the presence of wtp53 may not be absolutely required for the hypersensitivity for radiation-induced apoptosis in TGCT cell lines, ii) the molecular mechanism underlying the unique radiosensitivity was independent of the expression of Bcl-2 family proteins, and iii) cell susceptibility to apoptosis induction is not sufficiently informative to predict intrinsic radiosensitivity as determined by clonogenic survival.     

Nutlin-3, an Antagonist of MDM2, Enhances the Radiosensitivity of Esophageal Squamous Cancer with Wild-Type p53

Murine double minute 2 (MDM2) negatively regulates the activity of the p53 protein and plays a vital role in cell cycle arrest, apoptosis, and senescence mediated by p53. Nutlin-3, an antagonist of MDM2, is frequently used in anti-cancer studies. In many human tumors, nutlin-3 stabilizes p53 status and enhances p53 expression in cells with wild-type p53. However, the effect of nutlin-3 combined with radiotherapy on esophageal squamous cancer (ESCC) has not been reported. In this study, we examined whether nutlin-3 increases the radiosensitivity of ESCC in vitro and in vivo.We chose two cell lines, ECA-109 (wild-type p53) and TE-13 (p53 mutated), for the following experiments. Cell proliferation and clonogenic survival experiments showed that nutlin-3 inhibits the cell growth and colony formation of ECA-109 cells in a dose-dependent manner. Flow cytometry analysis showed that the apoptosis rate of ECA-109 cells co-treated with nutlin-3 and irradiation(IR) was significantly increased compared with cells treated with irradiation or nutlin-3 alone. Western blotting detected the expression of apoptosis-associated proteins in ECA-109 cells in response to nutlin-3 and irradiation. These effects were not evident in TE-13 cells. Xenograft mouse models indicated that nutlin-3 suppresses tumor growth and promotes radiosensitivity in the ESCC cell line ECA-109 in vivo. We have demonstrated that co-treatment of nutlin-3 with irradiation can significantly inhibit the growth and improve the radiosensitivity of ESCC cells with wild-type p53. The study suggests that nutlin-3 may be a potent therapeutic agent in conjunction with radiotherapy in ESCC.