Papanikolaou Vassilis, Iliopoulos Dimitrios, Dimou Ioannis, Dubos Stephanie, Tsougos Ioannis, Theodorou Kyriaki, Kitsiou-Tzeli Sofia, Tsezou Aspasia
Laboratory of Cytogenetics and Molecular Genetics, Medical School, University of Thessaly, 41110 Larissa, Greece.
Int J Oncol. 2009 Nov;35(5):1141-9. doi: 10.3892/ijo_00000430.
Cancer cell characteristics may play a pivotal role in the response to therapy by activating or deactivating different molecular pathways. In the present study, we investigated the implication of breast cancer cell features, such as HER2 and p53 in the activation of telomerase upon exposure to ionizing radiation. Telomerase is among the most important cancer biomarkers, conferring to tumor cells unlimited proliferative capacity, increased survival potential and resistance to several types of cellular stress. We investigated possible mechanisms regulating telomerase in six irradiated breast cancer cell lines (MCF-7, MCF-7/HER2, MDA-MB-231, SK-BR-3, BT-474 and HBL-100) differing in their HER2, p53 and ERalpha status. hTERT mRNA expression was evaluated by real-time PCR and telomerase activity by the TRAP assay. HER2, c-myc, p53 and p21 protein levels were evaluated by Western blotting. Silencing of hTERT and HER2 was achieved by small interfering RNA technology. Chromatin immunoprecipitation was used to evaluate H3 histone acetylation status, as well as myc/mad/max and p53 transcription factors interaction with the hTERT promoter. Our results showed for the first time, that only HER2-positive cells, independently of their p53 status, upregulated hTERT/telomerase, while knockdown of hTERT increased radio-sensitivity. Knockdown of HER2 also led to increased radio-sensitivity and downregulation of hTERT/telomerase. We also demonstrated that c-myc and mad1 regulate hTERT expression in all irradiated breast cancer cells. We conclude, for the first time, that HER2 phenotype upregulates hTERT through c-myc activation and confers radio-resistance to breast cancer cells.
癌细胞特征可能通过激活或失活不同分子途径在治疗反应中起关键作用。在本研究中，我们调查了乳腺癌细胞特征（如HER2和p53）在暴露于电离辐射后对端粒酶激活的影响。端粒酶是最重要的癌症生物标志物之一，赋予肿瘤细胞无限增殖能力、增强的生存潜力以及对多种细胞应激的抗性。我们研究了六种受辐射的乳腺癌细胞系（MCF-7、MCF-7/HER2、MDA-MB-231、SK-BR-3、BT-474和HBL-100）中端粒酶的可能调控机制，这些细胞系的HER2、p53和ERalpha状态各不相同。通过实时PCR评估hTERT mRNA表达，通过TRAP分析评估端粒酶活性。通过蛋白质印迹法评估HER2、c-myc、p53和p21蛋白水平。通过小干扰RNA技术实现hTERT和HER2的沉默。采用染色质免疫沉淀法评估H3组蛋白乙酰化状态，以及myc/mad/max和p53转录因子与hTERT启动子的相互作用。我们的结果首次表明，仅HER2阳性细胞，无论其p53状态如何，都会上调hTERT/端粒酶，而hTERT的敲低会增加放射敏感性。HER2的敲低也导致放射敏感性增加和hTERT/端粒酶的下调。我们还证明，c-myc和mad1在所有受辐射的乳腺癌细胞中调节hTERT表达。我们首次得出结论，HER2表型通过c-myc激活上调hTERT，并赋予乳腺癌细胞放射抗性。
