DRUG RESISTANCE & MECHANISMS OF RESISTANCE IN VITRO & IN CANCER PATIENTS

耐药性

• 批准号: 2464460
• 负责人: ANTONIO TITO FOJO
• 金额: --
• 依托单位: DIVISION OF CLINICAL SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2464460
• 关键词: DNA topoisomerases; P glycoprotein; acute lymphocytic leukemia; cell death; chromosome translocation; clinical research; gene expression; gene frequency; gene mutation; gene rearrangement; genetic translation; human subject; human therapy evaluation; multidrug resistance; neoplasm /cancer chemotherapy; oncogenes; phenotype; phosphorylation; protein isoforms; tissue /cell culture; tubulin; verapamil

In the field of multidrug resistance mediated by P-glycoprotein, our efforts continue to have a major focus on translational research, while trying to pursue basic investigations that have the potential for future clinical correlations. We have identified gene rearrangements as the mechanism responsible for the activation of MDR-1 in an increasing number of cell lines. These rearrangements occur randomly and are characterized by the juxtaposition of a transcriptionally active gene 5' to MDR-1, thus avoiding significant disruption of MDR-1 structure. While the occurrence of this phenomenon in clinical samples remains to be expanded, its demonstration in two samples from patients with refractory ALL, indicates this may be important in a defined group of patients. Our current investigations with MDR-1 arose out of studies which revealed a low frequency of acquired mutations in MDR-1. This prompted us to look at a second mechanism of drug resistance in which mutations had been described as a model for comparison. Similar studies with topoisomerase II-alpha have succeeded in isolating a larger number of acquired mutations, and characterization of these is underway. In one cell line, the identification of impaired nuclear translocation as the mechanism of resistance, led to the identification of a basic pentamer in the C terminal region of the gene as the motiff responsible for nuclear localization. In addition to the above studies, we have been able to show in the majority of single step isolates moderate to marked reductions in the level of topoisomerase II-alpha occurs as a mechanism of resistance, and are currently pursuing the molecular changes responsible for this decreased expression. A third active field of investigation began with our attempts to identify non-Pgp mechanisms of paclitaxel resistance. Selections performed with paclitaxel in the presence of verapamil succeeded in isolating a large number of cell lines with acquired resistance to paclitaxel that did not overexpress MDR-1. While the characterization of these cells is still underway, high resolution isoelectric focusing using purified tubulin has demonstrated the existence of new alpha and beta tubulin isoforms in the resistant cells, consistent with the occurrence of acquired mutations as the mechanism responsible for the resistant phenotype. Identification of the latter is ongoing. We have also used these models to confirm a possible pathway for cell death shared by tubulin active agents: activation of Raf-1 and phosphorylation of bcl-2.

在由p-糖蛋白介导的多药抗性领域中 努力继续关注转化研究，而 试图追求具有未来潜力的基本调查 临床相关性。我们已经确定基因重排是 导致MDR-1激活数量增加的机制 细胞系。 这些重排随机发生，并被表征 通过转录活性基因5'与MDR-1的并置，因此 避免了MDR-1结构的严重破坏。 而发生 在临床样本中的这种现象仍有待扩展， 来自难治性患者的两个样本中的示范表明 这在定义的患者组中可能很重要。 我们的目前 对MDR-1的研究是由研究揭示出低的研究 MDR-1中获得突变的频率。 这促使我们看了一个 描述突变的耐药性的第二种机制 作为比较的模型。 对拓扑异构酶II-α的类似研究 成功地隔离了大量获得的突变，并且 这些表征正在进行中。 在一个单元线中， 识别受损的核易位作为机制 电阻，导致C中的基本五聚物的鉴定 基因的末端区域作为负责核的Motiff 本土化。 除上述研究外，我们还能够证明 在大多数单步中，隔离物中等到明显的减少 拓扑异构酶II-α的水平是一种抗性机制， 并目前正在追求负责这一点的分子变化 表达下降。 第三个活跃的调查领域始于 我们试图识别紫杉醇抗性的非PGP机制。 在维拉帕米存在下用紫杉醇进行的选择 成功地隔离了大量的细胞系 对未表达MDR-1的紫杉醇的抗性。 而 这些细胞的表征仍在进行中，高分辨率 使用纯化的小管蛋白的等电聚焦已经证明了存在 抗性细胞中的新α和β微管蛋白同工型，一致 以获得的突变作为负责的机制 抗性表型。 后者的识别正在进行中。 我们 还使用这些模型确认了细胞死亡的可能途径 小管蛋白活性剂共享：RAF-1和磷酸化的激活 bcl-2。