Molecular Intervention in Thoracic Malignancies

目前的治疗方式无法影响肺癌和食管癌的自然史，以及这些恶性肿瘤的患病率，强调需要更基本的了解呼吸消化道致癌的分子机制。在以前的研究中，我们已经利用重组病毒载体，以证明恢复细胞周期蛋白D1，p16和p53在肺癌和食管癌细胞中的表达的影响。已知这些破坏G1限制点控制的突变发生在多步骤呼吸消化道癌变过程的早期，因此它们是干预已确定的恶性肿瘤及其前驱病变的适当靶点。最近的努力集中在评价可能达到分子终点而没有基因治疗策略的复杂性和局限性的药理学试剂依赖于病毒载体。我们已经观察到，合成黄酮，flavopiridol，介导深刻的细胞周期阻滞和凋亡的食管癌细胞，无论组织学或肿瘤抑制基因的状态，和预处理的食管癌细胞与flavopiridol显着提高其敏感性紫杉醇。这些数据为最近批准的方案提供了依据，该方案评价了食管癌患者中单独使用flavopiridol与flavopiridol联合紫杉醇的疗效。在已建立的癌症中的有效性的文件可能支持评价flavopiridol用于高危个体的呼吸消化道癌症的化学预防。大约30%的肺癌和食管癌过表达erbB-2/p185癌基因，其被认为介导对化疗剂如顺铂或紫杉醇的耐药性。erbB-2/p185基因功能的下调可以通过反义技术、酪氨酸激酶抑制剂或与p185蛋白的细胞外结构域结合的单克隆抗体来实现。最近，我们已经观察到在用安莎霉素类似物17-烯丙基氨基格尔德霉素（17-AA GA）处理的肺癌和食管癌细胞中pl 85蛋白的快速、剂量依赖性消耗。通过17-AA GA减少p185表达显著增强了紫杉醇在这些细胞中的细胞毒性;具体地，与单独暴露于紫杉醇的细胞相比，在用17-AA GA/紫杉醇处理的细胞中观察到紫杉醇敏感性增加3至超过40倍。这种增强效应是在纳摩尔浓度的17-AA GA下实现的。流式细胞术分析显示，联合药物治疗诱导明显的G2/M期阻滞和随后的细胞凋亡内的药物暴露72小时。这些临床前数据支持17-AA GA和紫杉醇在胸部肿瘤患者中的评价。在另外的研究中，我们试图利用去甲基化剂和组蛋白去乙酰化酶抑制剂来诱导肿瘤抑制基因如p16和ARF的表达，这些基因在胸部肿瘤中经常被启动子超甲基化灭活。有趣的是，这些实验与我们实验室中的平行研究一致，表明5氮杂-2脱氧胞苷（地西他滨）诱导肺癌细胞中NY-ESO-1和法师-3癌症-睾丸抗原表达。我们的研究已经表明，相对于正常的人支气管上皮细胞、成纤维细胞或EBV转化的淋巴细胞，肺癌细胞更容易诱导癌症睾丸抗原表达--这可能是由于已知在恶性转化期间发生的全基因组去甲基化。一项旨在评估地西他滨介导的肺癌患者肿瘤抗原和肿瘤抑制基因表达诱导的I期试验目前正在开放供患者招募。一项旨在评估Depsipeptide介导的肺癌患者基因诱导和细胞凋亡的II期研究将在不久的将来开放，近20%的癌症患者死于孤立的肺转移，我们部分的额外努力已被导向评估局部药物输送到肺部。建立了绵羊离体肺灌流模型，完成了紫杉醇的药代动力学研究。将使用该模型评价通过非病毒技术递送的其他药物和基因构建体。这些动物实验为在不可切除的肺恶性肿瘤患者中使用紫杉醇和中度高温进行隔离肺灌注的分析提供了依据。NCI-IRB已经审查了胸部肿瘤科提交的以下临床方案，预计在不久的将来将开放供患者招募：第一章食管内连续3天单独输注Flavopiridol与Flavopiridol联合输注3小时紫杉醇的随机II期研究癌症患者。2）地西他滨介导的肺癌患者中肿瘤抗原和肿瘤抑制基因表达的诱导的I期研究。3)肺癌患者静脉输注4小时Depsipeptide（FR 901228/NSC 630176）介导的基因诱导的II期研究。4）不可切除的肺恶性肿瘤患者使用紫杉醇和中度高温进行隔离肺灌注的I期研究。

The inability of current treatment modalities to impact on the natural history of lung and esophageal cancers, together with the prevalence of these malignancies, underscores the need for more fundamental appreciation of molecular mechanisms of aerodigestive tract carcinogenesis. In previous studies we have utilized recombinant viral vectors to demonstrate the impact of restoration of cyclin D1, p16, and p53 expression in lung and esophageal cancer cells. These mutations, which disrupt G1 Restriction Point control are known to occur early during multistep aerodigestive tract carcinogenesis, hence they are appropriate targets for intervention in established malignancies as well as their precursor lesions.In conjunction with Dr. Dao Nguyen, Senior Investigator, Thoracic Oncology Section, recent efforts have focused on the evaluation of pharmacologic agents which might achieve molecular end points without the complexities and limitations of gene therapy strategies dependent upon viral vectors. We have observed that the synthetic flavone, flavopiridol, mediates profound cell cycle arrest and apoptosis in esophageal cancer cells irrespective of histology or tumor suppressor gene status, and that pretreatment of esophageal cancer cells with flavopiridol markedly enhances their sensitivity to paclitaxel. These data have provided the rationale for a recently-approved protocol evaluating flavopiridol alone versus flavopiridol with paclitaxel in esophageal cancer patients. Documentation of efficacy in established cancers may support the evaluation of flavopiridol for the chemoprevention of aerodigestive tract cancers in high risk individuals.Approximately 30% of lung and esophageal cancers overexpress the erbB-2/p185 oncogene which is believed to mediate resistance to chemotherapeutic agents such as cisplatin or paclitaxel. Downregulation of erbB-2/pl85 gene function can be achieved by antisense techniques, tyrosine kinase inhibitors, or monoclonal antibodies that bind to the extracellular domain of the pl85 protein. Recently, we have observed rapid, dose-dependent depletion of pl85 protein in lung and esophageal cancer cells treated with the ansamycin analogue 17-allylamino geldanamycin (17-AA GA). Reduction of p185 expression by 17-AA GA markedly enhanced the cytotoxicity of taxol in these cells; specifically, a 3 to over 40-fold increase in paclitaxel sensitivity was observed in cells treated with 17-AA GA/paclitaxel as compared to cells exposed to paclitaxel alone. This enhancement effect was achieved at nanomolar concentrations of 17-AA GA. Flow cytometry analysis revealed that combination drug treatment induced pronounced G2/M arrest and subsequent apoptosis within 72 hours of drug exposure. These preclinical data support the evaluation of 17-AA GA and paclitaxel in patients with thoracic neoplasms.In additional studies we have attempted to utilize demethylating agents and histone deacetylase inhibitors to induce expression of tumor suppressor genes such as p16 and ARF which are frequently inactivated by promoter hypermethylation in thoracic neoplasms. Interestingly, these experiments have converged with parallel studies in our laboratory demonstrating induction of NY-ESO-1 and MAGE-3 cancer-testis antigen expression in lung cancer cells by 5 Aza-2 deoxycytidine (decitabine). Our studies have shown that cancer testis antigen expression is more readily induced in lung cancer cells relative to normal human bronchial epithelial cells, fibroblasts, or EBV transformed lymphocytes--perhaps due to genome-wide demethylation which is known to occur during malignant transformation. A phase I trial designed to evaluate decitabine-mediated induction of tumor antigen and tumor suppressor gene expression in lung cancer patients is currently open for patient accrual. A Phase II study designed to evaluate Depsipeptide mediated gene induction and apoptosis in lung cancer patients will be open in the near future.Nearly 20% of all cancer patients succumb to isolated pulmonary metastases, and additional efforts in our section have been directed toward the evaluation of regional drug delivery to the lungs. A sheep model of isolated lung perfusion has been established , and pharmacokinetic studies using paclitaxel have been completed. Additional pharmacologic agents and gene constructs delivered via nonviral techniques will be evaluated using this model. These animal experiments have provided the the rationale for analysis of isolated lung perfusion utilizing paclitaxel and moderate hyperthermia in patients with unresectable pulmonary malignancies.The following clinical protocols submitted by the Thoracic Oncology Section have been reviewed by the NCI-IRB and are expected to be open for patient accrual in the near future:1)Randomized Phase II Study of Continuous Three Day Flavopiridol Infusion Alone vs. Flavopiridol In Combination With Three Hour Taxol Infusion inEsophageal Cancer Patients.2)Phase I Study of Decitabine Mediated Induction of Tumor Antigen and Tumor Supressor Gene Expression in Lung Cancer Patients. 3) Phase II Study of Gene Induction Mediated by Four-Hour Intravenous Depsipeptide (FR901228/NSC630176) Infusion in Lung Cancer Patients.4)Phase I Study of Isolated Lung Perfusion with Paclitaxel and Moderate Hyperthermia in Patients with Unresectable Pulmonary Malignancies.