非小细胞肺癌组织p53蛋白Ser15磷酸化表达的研究

[目的]探讨人非小细胞肺癌组织中突变型p53蛋白15位丝氨酸位点（p53Ser15）的磷酸化状况,p53Ser15磷酸化、p53蛋白和血清p53抗体水平之间的关联以及与非小细胞肺癌临床病理特征和预后的关系。[方法]应用酶联免疫吸附实验（ELISA）检测40例非小细胞肺癌组织、相应正常肺组织及10例肺良性疾病组织中p53Ser15磷酸化、p53蛋白以及血清p53抗体水平。随访观察40例非小细胞肺癌组的转移复发情况。[结果]p53Ser15磷酸化、p53蛋白水平在非小细胞肺癌组高于正常肺组织组和肺部良性疾病组,差异有统计学意义（P〈0.05,P〈0.01）。非小细胞肺癌组p53Ser15磷酸化、p53蛋白及血清p53抗体水平两两之间均有正相关性（P〈0.05）。非小细胞肺癌组中p53抗体阳性率为42.5%,在鳞癌中的阳性率高于腺癌和腺鳞癌（P〈0.05）。随访期内5例出现复发或远处转移的病例p53Ser15磷酸化、p53蛋白表达水平高于未复发转移病例（P〈0.05）。[结论]突变型p53蛋白在人体内也可如野生型p53一样被磷酸化,而且这种磷酸化有助于突变型p53蛋白的稳定。p53Ser15磷酸化及p53蛋白高水平表达可能会有助于肿瘤进展更快。     

非小细胞肺癌p73基因突变与表达的研究

[目的]研究p73基因在非小细胞啼癌中的突变及表达情况，分析其与非小细胞肺癌的临床分期、组织学分型的相关性，探讨p73与肺癌发生、发展的关系．[方法]应用催化信号放大（CSA）法检测40例非小细胞肺癌患者癌组织及癌旁组织p73蛋白表达；酚-氯仿-异戊醇法提取基因组DNA，应用PCR—SSCP法（单链构象多态性分析）检测p73基因突变，并对出现异常条带样本进行DNA测序分析．[结果]40例非小细胞肺癌组织中p73阳性表达率52．5％（21／40）明显高于癌旁肺组织（0）（P〈0．005）。p73阳性表达率与肿瘤临床分期、组织学分型有相关性（P〈0．01）。有3例p73基因第2位外显子序列存在遗传多态性，碱基T→C置换、[结论]p73在非小细胞肺癌组织中存在过度表达及遗传多态性，D73可能参与调控非小细胞肺癌的发生、发展过程．     

非小细胞肺癌中p53蛋白表达及其临床意义

目的：为探讨非小细胞肺癌中p53蛋白表达与肺癌组织学类型、分化程度和淋巴结转移的关系。方法：应用免疫级化技术（ABC法），以p53单克隆抗体检测53例非小细胞肺癌组织中p53基因的表达，组织经微波抗原修复后进行免疫染色。结果采用X2检验。结果：p53蛋白在53例非小细胞肺癌中表达阳性率为64．2％，p53蛋白表达与非小细胞肺癌的组织学类型无关（P＞0.05）；与肺鳞癌分化程度有关（P＜0.01）；肺腺癌中与淋巴结转移有关（p＜0.05）。结论：p53基因的表达异常与不同类型的肺癌发生发展相关，提示P53基因在肺癌的形成、分化和转移过程中起不同的调节作用，p53蛋白过度表达则预后差，检测p53蛋白对判断非小细胞肺癌预后具有应用价值。     

非小细胞肺癌FHIT基因蛋白表达缺失及与突变型p53表达相关性

[目的]了解非小细胞肺癌组织(NSCLC)中脆性组氨酸三联体(FHIT)基因蛋白(Fhit)表达缺失及其与突变型p53基因蛋白(p53)表达的相关性.[方法]应用免疫组织化学方法了解NSCLC组织中Fhit及p53的表达.[结果]62.2%(56/90)的NSCLC组织显示Fhit表达缺失,同时48.9%(44/90)有突变型p53表达;两者的异常表达在非鳞组中明显相关(P＜0.001),而在鳞癌组中无相关性(P=0.079);另外,两者的异常表达在非吸烟组中明显相关(P=0.006),而在吸烟组中无相关性(P=0.113).[结论]Fhit表达缺失和突变型p53表达是NSCLC中常见事件;而且两者在non-SqCC和非吸烟的肺癌者中有明显的相关性.     

Maspin在非小细胞肺癌中的表达及与p53的相关性

目的探讨maspin在非小细胞肺癌中的表达及其与p53的关系。方法对99例非小细胞肺癌组织maspin及p53蛋白的表达情况通过免疫组化法进行检测,分析其临床病理意义及相关性。结果 maspin蛋白表达与肿瘤病理类型、肿瘤分化程度、淋巴结状态及临床分期相关（P〈0.05）。而p53蛋白的表达则与临床分期、淋巴结状态相关（P〈0.05）。maspin与p53蛋白的表达呈负相关（γ=-0.180）,但无统计学意义（P=0.075）。结论 Maspin及p53蛋白的表达在非小细胞肺癌的发生、发展过程中起重要作用,但本研究未发现两者具有统计学意义的相关性。     

非小细胞肺癌中PTEN及p53的表达及其临床意义

目的研究非小细胞肺癌（NSCLC）中PTEN及p53的表达及其临床意义。方法采用免疫组化S-P法,对61例NSCLC手术切除标本进行PTEN及突变型p53蛋白表达研究,分析其与NSCLC患者的病理类型及临床分期、预后的关系。结果（1）非小细胞肺癌中PTEN的表达明显低于对照组（P〈0.05）;而p53蛋白则明显增高。（2）PTEN表达与组织学类型、临床分期和淋巴结转移显著相关;p53表达与性别和淋巴结转移显著相关。（3）非小细胞肺癌标本中PTEN与p53的表达呈显著负相关（rs=-0.282,P〈0.05）。（4）Kaplan-Meier生存分析显示,PTEN表达阴性组患者平均生存期为36.5月,阳性组为71.0月;p53表达阴性组患者平均生存期为70.1月,阳性组为46.0月,经Logrank检验均显示P〈0.05。结论PTEN的表达缺失及野生型p53基因的突变,可能是导致NSCLC发生的重要原因之一,可以作为肿瘤基因治疗的靶点;PTEN及p53的表达与NSCLC患者的预后关系密切,可能是肿瘤预后的标志物;PTEN与p53的表达呈明显的负相关,两者是否通过同一途径,或者两条途径如何相互影响,需进一步研究。     

p53蛋白在非小细胞肺癌中表达及与其临床特征、同步放化疗疗效的关系分析

目的：检测p53蛋白在非小细胞肺癌患者中的表达水平,并探讨其表达与临床特征的相关性及同步放化疗疗效的关系。方法采用免疫组化方法,检测100例非小细胞肺癌组织中p53表达水平,结合显微图像分析仪测定p53免疫组化强度,以阳性单位为定量分析单位。分析p53表达水平与患者临床病理特征及同步放化疗疗效的关系。结果 p53蛋白在非小细胞肺癌中阳性表达率为48％;p53表达与淋巴结转移、远端转移和TNM分期均无显著相关性;非小细胞肺癌p53蛋白表达者放化疗效果显著低于p53蛋白表达阴性患者,且p53蛋白表达与患者生存期呈负相关。结论 p53表达与否影响非小细胞肺癌患者化疗效果,与生存期直接相关,临床治疗时可考虑将其作为治疗靶点。     

p53、p16、FHIT基因蛋白在非小细胞肺癌中的表达及意义

目的研究非小细胞肺癌中p53、p16、FHIT基因蛋白的表达及意义.方法应用免疫组化S-P法检测62例非小细胞肺癌标本中p53、p16、FHIT的表达和组织学分型、分期等临床病理参数之间的关系.结果p53与肺癌分期、淋巴结转移及年龄有显著性差异,与分型、吸烟史、性别无关;p16与肺癌分期、淋巴结转移有显著性差异,与分型、年龄、性别、吸烟史均无关.FHIT与肺癌的分型、吸烟史、性别有显著性差异,与分期、年龄、淋巴结转移无关.结论p53、p16基因改变在非小细胞肺癌的发生、发展中起重要作用,p53、p16基因蛋白异常表达可作为评价肺癌预后的有效指标,而FHIT基因表达缺失是常见事件.     

肺癌病人痰液标本p53基因突变临床流行病学分析

[目的]探讨痰液标本p53基因突变检测用于肺癌高危人群筛查和肺癌早期诊断方面的意义。[方法]应用PCR鄄SSCP对63例肺癌患者和30例肺良性疾病的肺组织和痰液细胞中p53基因5～8外显子的突变进行了检测,并采用临床流行病学的方法进行分析。[结果]肺癌病人组肺癌组织中p53基因突变率为49.2%(31/63),对照组组织中突变率为3.3%(1/30),两组差别有统计学意义(P<0.001)。肺癌组痰液标本中p53突变率为23.8%(15/63);肺良性疾病患者痰液中未发现p53基因突变。以痰液标本p53基因突变为诊断指标,诊断的敏感度为45.2%,特异度为96.8%,标化阳性预告值为93.5%,标化阴性预告值为63.9%,标化一致性为71.0%。[结论]检测痰液中p53基因突变能间接反映其在肺组织中的改变,该方法可作为肺癌高危人群筛查以及肺癌诊断的无创性手段之一。     

重组人p53腺病毒逆转非小细胞肺癌顺铂耐药的体外研究

目的 为重组人p53腺病毒应用于临床非小细胞肺癌顺铂耐药患者提供实验依据.方法 采用CCK-8试剂检测半数抑制浓度(IC50);应用流式细胞仪检测细胞周期分布及凋亡发生率;采用功能分类基因芯片检测药物作用前后的基因变化.结果 顺铂联合重组人p53腺病毒后,H1299细胞顺铂的半数抑制浓度下降至5 μg/mL(P<0.05),细胞的凋亡率上升为10.73%(P<0.05).重组人p53腺病毒单药作用后,H1299细胞内p53基因大量表达.在两药联合作用下,导入的p53基因激活了由Fas/Apo-1/CD95系统、TNFR超家族、Bcl-2/Bax等所介导的凋亡通路,同时也上调了部分细胞周期调控基凶.结论 重组人p53腺病毒介导的基因治疗有希望成为治疗临床非小细胞肺癌顺铂耐药患者的一种有效的辅助手段.     

Clinical update of Ad-p53 gene therapy for lung cancer

These preliminary Phase I and II gene therapy trials in NSCLC have demonstrated that Ad-p53 gene transfer is associated with low toxicity and evidence of antitumoral activity at the locoregional site. Effort to enhance antitumoral efficacy with chemotherapy and radiation therapy have not increased Ad-p53 toxicity and appear to be feasible. Randomized Phase III studies are now needed to determine the potential of Ad-p53 to improve overall survival in selected subsets of NSCLC patients. Future gene therapy research is required to develop systemic delivery systems and to overcome p53 tumor resistance. It is hoped that these efforts will ultimately lead to a novel mode of therapy to complement conventional chemotherapy, radiation therapy, and surgical treatment strategies.     

Late resistance to adenoviral p53-mediated apoptosis caused by decreased expression of Coxsackie-adenovirus receptors in human lung cancer cells

Adenovirus-mediated wild-type p53 gene transfer induces apoptosis in a variety of human cancer cells. Although clinical trials have demonstrated that a replication-deficient recombinant adenovirus expressing the wild-type p53 gene (Ad-p53) is effective in suppressing growth of non-small cell lung cancer (NSCLC), we often experienced late resistance to this treatment. To elucidate the mechanism of late resistance to Ad-p53 in human lung cancer cells, we generated 5 different resistant variants from p53-susceptible H1299 NSCLC cells by repeated infections with Ad-p53. We first examined the transduction efficiency of adenoviral vector by Ad-LacZ transduction followed by X-gal staining in parental and 5 resistant H1299 cell lines. Their sensitivity to viral infection decreased in correlation with the magnitude of resistance, and Ad-p53-mediated tumor suppression could be restored by dose escalation of Ad-p53 in the resistant variants. The expression of Coxsackie and adenovirus receptor (CAR) and alphaV integrins, which are cellular receptors for attachment and internalization of the virus, respectively, was next investigated in these cell lines. Flow cytometry revealed that alphaVbeta3 and alphaVbeta5 integrin expression was consistent, while p53-resistant cell lines showed that diminished CAR expression correlated with the magnitude of the resistance. Our results demonstrated that decreased CAR expression could be one of the mechanisms of late resistance to Ad-p53, which may have a significant impact on the outcome of adenovirus-based cancer gene therapy.     

PCR—SSCP法检测非小细胞肺癌p53基因的突变

目的 分析原发性非小细胞肺癌（ＮＳＣＬＣ）中ｐ５３基因５～８外显子点突变发生及意义。方法 动用聚合酶链反应－单链构象多态性分析方法（ＰＣＲ－ＳＳＣＰ分析法）进行检测。结果 发现８例肺良性疾病均无点突变发生,４０例ＮＳＣＬＣ中１９例（４７．５％）有点突变发生,点突变发与性别,年龄,组织学类型和组织发级无明显相关性（Ｐ〉０．０５）,但与临床分期和淋巴结累及状态明显相关（Ｐ〈０．０１,Ｐ〈０．０５）。Ｘ     

p53 Gene Therapy

Based on the frequent inactivation of the p53 gene product in human malignancy, and its functional involvement in tumor suppression, cell cycle control and apoptosis, p53 was identified as an attractive target for somatic gene therapy strategies in cancer. Several pilot and phase I studies explored the intratumoral injection of viral expression vectors encoding the p53 cDNA in patients with advanced cancer. These studies confirmed the safety and feasibility of this approach. Further, vector-specific transgene expression and surrogate markers for biological activity of the transgene were demonstrated. Local tumor regression, or stabilization of tumor growth, were observed in some studies, and were interpreted as evidence for clinical activity. No formal assessment of in vivo transduction efficacy and vector distribution, following intratumoral injection of p53 expression vectors, was performed. The instillation of adenoviral p53 expression vectors into cavitary organs, such as the peritoneum or the bladder, has been studied as an alternative mode of application. Intravesical p53 vector installation, in combination with a transduction-enhancing agent, proved to be safe, feasible and biologically active in a phase I study of patients with bladder cancer. Transgene expression, following vector installation, was superior to intratumoral vector injection in this trial, and effective vector distribution and penetration were achieved by the former treatment. The publication of results from the studies of intraperitoneal vector installation in patients with ovarian cancer is still awaited. To date, only one published phase II study has formally assessed the clinical efficacy of adenovirus-mediated p53 gene transfer in patients with advanced non-small cell lung cancer (NSCLC). No clinically relevant benefit of local p53 gene therapy was identified in patients undergoing systemic chemotherapy when local responses of individual tumor lesions treated with intratumoral vector were compared. The results from additional efficacy studies performed in patients with NSCLC or head and neck cancer are pending. In summary, current studies failed to translate the preclinical success of intratumoral p53 vector injection into the clinic. A major obstacle is the unknown, and probably inefficient, vector delivery following intratumoral injection. In contrast, an effective delivery can be achieved by intravesical instillation of a p53 expression vector in patients with bladder cancer. These results merit further investigation in phase II studies. Recent improvements in vector technology, including targeting techniques, tissue-specific transgene expression andtumor-selective vector replication, will certainly allow investigators to study the efficacy of p53 gene therapy in a next wave of carefully conducted clinical trials.     

Adenovirus-mediated p53 gene therapy has greater efficacy when combined with chemotherapy against human head and neck, ovarian, prostate, and breast cancer

Purpose: Adenovirus-mediated p53 gene therapy for cancer is currently undergoing phase I/II clinical trials. The drug used in our clinical trials (p53 Ad; ACN53; SCH58500) consists of a replication-deficient, type 5 adenovirus vector expressing human wildtype p53 tumor suppressor under the control of the cytomegalovirus promoter. In preclinical models, p53 Ad has therapeutic efficacy against a wide range of human tumor types containing nonfunctional p53, both in vitro and in vivo. Results from early clinical trials using p53 gene therapy by itself support optimism for the future of this therapeutic approach. However, it is likely that many phase II/III trials will incorporate an arm comparing traditional chemotherapy against chemotherapy combined with p53 gene therapy. Therefore, it is important to study possible interactions between p53 Ad and chemotherapeutic drugs in preclinical models before starting the clinical trials. Methods: Proliferation of tumor cells was quantitated after incubation with various combinations of p53 Ad and chemotherapeutic drugs. Human tumor xenografts in scid mice were dosed with intraperitoneal or intratumoral p53 Ad with or without chemotherapeutic drugs and the tumor burden after therapy monitored. Results: p53 Ad combined with cisplatin, doxorubicin, 5-fluorouracil, methotrexate, or etoposide inhibited cell proliferation more effectively than chemotherapy alone in SCC-9 head and neck, SCC-15 head and neck, SCC-25 head and neck, SK-OV-3 ovarian, DU-145 prostate, MDA-MB-468 breast, and MDA-MB-231 breast tumor cells. No obvious dependence on dosing schedule was observed. Greater anticancer efficacy was also demonstrated in four human tumor xenograft models in vivo. Of particular significance, there was enhanced efficacy using the three drug combination of p53 Ad, cisplatin, and paclitaxel in an ovarian cancer model. Conclusion: These results support the combination of p53 gene therapy with chemotherapy in clinical trials. Received: 5 October 1998 / Accepted: 3 December 1998     

Promising developments in targeted therapies for non-small-cell lung cancer

Despite advances in chemotherapy, radiation therapy, and surgery, the overall survival for patients with lung cancer remains poor. Thus, novel therapeutic approaches are warranted. As knowledge of the molecular abnormalities and dysregulated cellular processes contributing to the pathogenesis and progression of lung cancer has been acquired, intense interest has been directed at developing agents that target these abnormalities. New agents targeting aberrant receptor tyrosine kinases, the Ras oncoprotein, mediators of metastases and angiogenesis, and the tumor suppressor gene p53 have, among other agents, shown promise in preclinical studies. Early clinical trials with these agents in patients with advanced malignancies suggest preliminary evidence of clinical activity and possible applications in non-small-cell lung cancer (NSCLC). Ongoing clinical trials will help clarify the settings in which these agents are of greatest therapeutic value, the optimal schedule of administration, toxicities associated with chronic administration, and hopefully, provide additional insight into the biology of lung cancer. Selected clinical trials will be presented to highlight the use of rationally designed, targeted therapies for patients with NSCLC.     

驱动基因突变非小细胞肺癌免疫治疗的研究进展

随着肺癌靶点的发现和药物研发,靶向治疗改善了驱动基因突变非小细胞肺癌(NSCLC)的临床预后。同时,免疫检查点抑制剂在驱动基因阴性NSCLC中也取得了良好的疗效。虽然部分驱动基因突变患者从对应靶向治疗中明显获益,但对免疫治疗反应欠佳。在大部分免疫治疗临床研究和日常实践中,EGFR/ALK等驱动基因突变阳性的NSCLC患者也被排除在外,或者仅占少数。免疫治疗如何应用于驱动基因突变患者,以及如何在靶向治疗、化疗及免疫治疗中选择最佳治疗方案,制定最优治疗策略,对改善晚期驱动基因突变NSCLC患者预后至关重要。本文对不同基因突变肿瘤免疫微环境的特点及免疫治疗在不同基因突变的NSCLC患者中的应用进行简要综述。     

非小细胞肺癌CT与相关基因表达的研究

目的：研究非小细胞（ＮＳＣＬＣ）肺癌ＣＴ征象与肺癌相关基因ｐ５３,Ｃ－ｍｙｃ,Ｋ－ｒａｓ,及Ｃ－ｅｒｂＢ２表达间的关系。方法：运用免疫组化法,检测５４例经手术病理证实的非小细胞肺癌组织中的ｐ５３,Ｃ－ｍｙｃ,Ｋ－ｒａｓ及Ｃ－ｅｒｂＢ２基因的表达,并分析其与术前ＣＴ征象间的关系。结果：ｐ５３表达与非小细胞肺癌期别及ＣＴ征象中瘤体大小,深分叶征,毛刺征,空洞征,及纵隔淋巴结转移有关,但与组织类型胸,胸