Escuin Daniel, Kline Erik R, Giannakakou Paraskevi
Department of Hematology and Oncology, Winship Cancer Institute, Robert Woodruff Health Sciences Center, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Cancer Res. 2005 Oct 1;65(19):9021-8. doi: 10.1158/0008-5472.CAN-04-4095.
We have recently identified a mechanistic link between disruption of the microtubule cytoskeleton and inhibition of tumor angiogenesis via the hypoxia-inducible factor-1 (HIF-1) pathway. Based on this model, we hypothesized that other microtubule-targeting drugs may have a similar effect on HIF-1alpha. To test that hypothesis, we studied the effects of different clinically relevant microtubule-disrupting agents, including taxotere, epothilone B, discodermolide, vincristine, 2-methoxyestradiol, and colchicine. In all cases, HIF-1alpha protein, but not mRNA, was down-regulated in a drug dose-dependent manner. In addition, HIF-1alpha transcriptional activity was also inhibited by all drugs tested. To further examine whether these effects were dependent on microtubule network disruption, we tested the ability of epothilone B to inhibit HIF-1alpha protein in the human ovarian cancer cell line 1A9 and its beta-tubulin mutant epothilone-resistant subclone 1A9/A8. Our data showed that epothilone B treatment down-regulated HIF-1alpha protein in the parental 1A9 cells but had no effect in the resistant 1A9/A8 cells. These observations were confirmed by confocal microscopy, which showed impaired nuclear accumulation of HIF-1alpha in parental 1A9 cells at epothilone B concentrations that induced extensive microtubule stabilization. In contrast, epothilone B treatment had no effect on either microtubules or HIF-1alpha nuclear accumulation in the resistant 1A9/A8 cells. Furthermore, epothilone B inhibited HIF-1 transcriptional activity in 1A9 cells, as evidenced by a hypoxia response element-luciferase reporter assay, but had no effect on HIF-1 activity in the resistant 1A9/A8 cells. These data directly link beta-tubulin drug binding with HIF-1alpha protein inhibition. Our results further provide a strong rationale for testing taxanes and epothilones in clinical trials targeting HIF-1 in cancer patients.
我们最近发现微管细胞骨架的破坏与通过缺氧诱导因子-1（HIF-1）途径抑制肿瘤血管生成之间存在机制联系。基于此模型，我们推测其他靶向微管的药物可能对HIF-1α有类似作用。为验证该假设，我们研究了不同临床相关的微管破坏剂的作用，包括多西他赛、埃坡霉素B、软海绵素、长春新碱、2-甲氧基雌二醇和秋水仙碱。在所有情况下，HIF-1α蛋白而非mRNA以药物剂量依赖性方式下调。此外，所有测试药物也抑制了HIF-1α转录活性。为进一步研究这些作用是否依赖于微管网络破坏，我们测试了埃坡霉素B在人卵巢癌细胞系1A9及其β-微管蛋白突变体埃坡霉素抗性亚克隆1A9/A8中抑制HIF-1α蛋白的能力。我们的数据显示，埃坡霉素B处理使亲本1A9细胞中的HIF-1α蛋白下调，但对抗性1A9/A8细胞无作用。共聚焦显微镜证实了这些观察结果，其显示在诱导广泛微管稳定的埃坡霉素B浓度下，亲本1A9细胞中HIF-1α的核积累受损。相比之下，埃坡霉素B处理对抗性1A9/A8细胞中的微管或HIF-1α核积累均无影响。此外，缺氧反应元件-荧光素酶报告基因检测证明，埃坡霉素B抑制1A9细胞中的HIF-1转录活性，但对抗性1A9/A8细胞中的HIF-1活性无作用。这些数据直接将β-微管蛋白药物结合与HIF-1α蛋白抑制联系起来。我们的结果进一步为在针对癌症患者HIF-1的临床试验中测试紫杉烷类和埃坡霉素类药物提供了有力依据。
