Molecular Intervention in Thoracic Malignancies

肺、食道和胸膜恶性肿瘤在癌变领域通过多步骤机制发生。尽管它们的病因可能不同，但这些恶性肿瘤表现出共同的分子主题，包括通过ras通路的异常信号转导，Rb和P53抑癌通路的失活，以及端粒酶的表达，所有这些似乎都是恶性转化的关键。除了扰乱细胞周期调控外，癌基因和抑癌基因突变还通过调节黏附分子、基质金属蛋白酶以及血管内皮生长因子等前血管生成配体的表达来影响胸部肿瘤的转移表型。染色质结构和DNA甲基化的整体紊乱使多种肿瘤抑制基因沉默，从而促进了几种肿瘤睾丸抗原在胸部恶性肿瘤中的从头表达。 涉及ErbB1/2、Cyclin D、Rb、p16、p14/ARF和P53的突变是胸部恶性肿瘤及其前体病变干预的有吸引力的靶点；SV40癌蛋白可能是恶性胸膜间皮瘤分子干预的独特靶点。在已发表的研究中，我们已经证明，17-AAG及其相关化合物阻断erb B信号转导显着增强了对化疗药物的敏感性，并抑制了肺癌和食道癌细胞的转移表型。 更多的实验室工作集中在分析由DNA脱甲基剂和组蛋白脱乙酰酶(HDAC)抑制剂介导的胸部恶性肿瘤的基因诱导。我们观察到，在促进p16等抑癌基因表达的暴露条件下，5-氮杂-2‘-脱氧胞苷(DAC)介导了MAGE-3和NY-ESO-1肿瘤睾丸抗原的诱导。此外，我们还证明了DAC介导的靶基因诱导可以被HDAC抑制剂Depsi多肽FR901228(DP)增强。此外，我们已经证明，在DAC或DAC/DP连续暴露后，癌细胞可以被NY-ESO-1特异性的细胞溶解T淋巴细胞识别，并且DAC/DP连续治疗可以在癌细胞中介导深刻的促凋亡效应，但不能诱导培养的正常人支气管上皮细胞发生凋亡。这些数据为两个检查DAC和DP介导的基因诱导的积极临床试验提供了临床前基础，以及另一个计划在不久的将来开始评估胸部肿瘤患者DAC/DP序贯输注的方案。 在观察到DAC、DP和顺序的DAC/DP介导生长停滞与组织学或肿瘤抑制基因状态无关后，我们利用基因芯片技术来研究这些药物优先介导癌细胞凋亡的机制。到目前为止，这项分析揭示了这些药物对癌细胞的高度复杂的影响，没有明显的诱导模式相对于基因。已经确定了几种新的机制，并提交了与其中一些实验有关的结果以供发表。 除了我们的体外实验，我们还使用了基因芯片技术来分析肺癌患者接受地西他滨或Depsi多肽治疗后组织活检标本中的基因表达。在这些试验中，我们可以可靠地从患者的FNA和支气管镜活检标本中扩增出mRNA。由于癌症标本中基因诱导的评估可能会受到间质污染的不利影响，目前正在努力优化激光捕获显微切割和RNA扩增技术，以提高遗传分析的特异性。到目前为止，在这些试验中已经观察到几名患者有靶基因诱导的证据。 在所有死于胸外恶性肿瘤的患者中，近三分之一患有肺转移。在临床前研究中，我们通过逆行隔离肺灌流技术评价了紫杉醇的药代动力学和急性毒性，并观察到中高温显著增强了紫杉醇对癌细胞的细胞毒性。这些实验为正在进行的第一阶段研究提供了临床前理论基础。此外，我们还通过检查通过新型雾化技术给药的细胞毒剂的毒性和潜在疗效，扩展了我们在肺部恶性肿瘤的区域治疗方面的努力。 由胸科肿瘤科提交的以下临床方案可供患者应计： 1)第一阶段地西他滨诱导肺癌、食道癌、胸膜癌患者肿瘤抗原和抑癌基因表达的研究。 2)肺癌患者静脉滴注4小时多肽介导的基因诱导的II期研究。 3)不能切除的肺部恶性肿瘤逆行隔离肺灌注紫杉醇和中温热疗的I期研究。 4)阿霉素吸入治疗成人晚期肺部实体瘤的I期及临床药理学研究

Pulmonary, esophageal, and pleural malignancies arise via multistep mechanisms in cancerization fields. Although their etiologies may differ, these malignancies exhibit universal molecular themes including aberrant signal transduction via ras pathways, inactivation of the Rb and p53 tumor suppressor pathways, and expression of telomerase, all of which appear critical for malignant transformation. In addition to disrupting cell cycle regulation, oncogene and tumor suppressor gene mutations influence the metastatic phenotype of thoracic neoplasms by modulating expression of adhesion molecules, matrix metalloproteinases, as well as proangiogenic ligands such as vascular endothelial growth factor. Global derangements in chromatin structure and DNA methylation which silence a variety of tumor suppressor genes facilitate de novo expression of several cancer testis antigens in thoracic malignancies. Mutations involving erbB1/2, cyclin D, Rb, p16, p14/ARF, and p53 are attractive targets for intervention in thoracic malignancies and their precursor lesions; SV40 oncoproteins may be unique targets for molecular intervention in malignant pleural mesotheliomas. In published studies we have demonstrated that abrogation of erbB signal transduction by 17-AAG and related compounds markedly enhances sensitivity to chemotherapeutic agents and inhibits the metastatic phenotype of lung and esophageal cancer cells. Additional laboratory efforts have focused on the analysis of gene induction in thoracic malignancies mediated by DNA demethylating agents and histone deacetylase (HDAC) inhibitors. We have observed that under exposure conditions that facilitate expression of tumor suppressor genes such as p16, 5 aza 2' deoxycytidine (DAC) mediates induction of MAGE-3 and NY-ESO-1 cancer testis antigens. In addition we have demonstrated that DAC-mediated target gene induction can be augmented by the HDAC inhibitor Depsipeptide FR901228 (DP). Furthermore, we have shown that following exposure to DAC, or sequential DAC/DP, cancer cells can be recognized by cytolytic T lymphocytes specific for NY-ESO-1, and that sequential DAC/DP treatment mediates profound proapoptotic effects in cancer cells but not cultured normal human bronchial epithelial cells. These data have provided the preclinical rationale for two active clinical trials examining gene induction mediated by DAC and DP, as well as another protocol evaluating sequential DAC/DP infusion in thoracic oncology patients scheduled to commence in the near future. Having observed that DAC, DP, and sequential DAC/DP mediate growth arrest irrespective of histology or tumor suppressor gene status, we have utilized cDNA microarray techniques to examine the mechanisms by which these agents mediate apoptosis preferentially in cancer cells. Thus far, this analysis has revealed highly complex effects of these agents in cancer cells without obvious patterns of induction relative to genotype. Several novel mechanisms have been identified, and results pertaining to some of these experiments have been submitted for publication. In addition to our in vitro experiments, we have used cDNA microarray techniques to analyze gene expression in tissue biopsy specimens following Decitabine, or Depsipeptide treatment of lung cancer patients. We can reliably amplify mRNA from FNA and bronchoscopic biopsy specimens from patients on these trials. Because the evaluation of gene induction in cancer specimens may be adversely affected by stromal contamination, current efforts are underway to optimize laser capture microdissection and RNA amplification techniques in order to enhance the specificity of genetic analysis.Thus far, evidence of target gene induction has been observed in several patients on these trials. Nearly one-third of all patients dying from extrathoracic malignancies suffer from pulmonary metastases. In preclinical studies, we have evaluated the pharmacokinetics and acute toxicity of paclitaxel administered by retrograde isolated lung perfusion techniques, and have observed significant enhancement of paclitaxel cytotoxicity in cancer cells by moderate hyperthermia. These experiments have provided the preclinical rationale for an ongoing Phase I study. Furthermore, we have extended our efforts pertaining to the regional treatment of pulmonary malignancies by examining the toxicity and potential efficacy of cytotoxic agents administered via novel aerosolization techniques. The following clinical protocols submitted by the Thoracic Oncology Section are open for patient accrual: 1) Phase I Study of Decitabine Mediated Induction of Tumor Antigen and Tumor Suppressor Gene Expression in Patients with Cancers Involving the Lungs, Esophagus, or Pleura. 2) Phase II Study of Gene Induction Mediated by Four-Hour Depsipeptide Infusion in Lung Cancer Patients. 3) Phase I Study of Retrograde Isolated Lung Perfusion with Paclitaxel and Moderate Hyperthermia in Patients with Unresectable Pulmonary Malignancies. 4) Phase I and Clinical Pharmacologic Study of Inhaled Doxorubicin in Adults with Advanced Solid Tumors Affecting the Lungs.