Mechanisms of therapy resistance in colorectal carcinomas: regulation of apoptosis by microRNA-mediated oxidative-metabolic effects

凋亡信号级联的缺陷本质上导致恶性肿瘤的治疗抗性。许多结直肠癌患者的治疗反应差是由于肿瘤组织的原发性或获得性放射和化学抗性。然而，可靠的分子标记物治疗耐药的癌症尚未确定到目前为止，有效的治疗方法，以克服放射化学耐药的结直肠癌仍然缺失。因此，更好地理解治疗抗性的细胞和分子基础是开发改进的诊断和治疗工具的核心要求。肿瘤细胞对细胞凋亡的抗性增加通常以Bcl-2蛋白家族的促细胞凋亡成员的表达减少为特征。在我们的初步实验中，我们发现microRNA-210（miR-210）受到电离辐射的调节，并且miR-210的表达增加在结肠癌细胞凋亡的启动中起重要作用。我们的研究结果表明，miR-210的细胞毒性作用是由促凋亡Bim蛋白发挥的，Bim表达是由miR-210介导的ROS增加诱导的，随后CHOP和FOXO转录因子被激活。在所提出的项目中，我们旨在证明Bim表达的miR-210依赖性诱导是通过抑制NDUFA 4和SDHD（呼吸链复合物的两个亚基）以及连续抑制相应的呼吸链复合物和增加ROS产生来介导的。此外，我们将调查（无论是在细胞培养和动物实验），在何种程度上，这种特殊的机制，以及一般的氧化代谢活性调节细胞凋亡的阻力。此外，我们的初步实验证明了miR-210的辐射依赖性调节。在这种情况下，我们将分析哪些转录因子参与这种调制。此外，将研究辐射对非致瘤组织中miR-210表达的影响。此外，我们将分析miR-210和Bim在大量人类结直肠癌样本（n = 1250）中表达的预后和预测价值。

Defects in the apoptotic signaling cascades essentially contribute to the therapy resistance of malignant tumors. The poor treatment response in many patients with colorectal cancer results from a primary or acquired radio- and chemoresistance of the tumor tissue. However, reliable molecular markers for therapy-resistant carcinomas have not been identified so far, and effective therapeutic approaches to overcome radiochemoresistance in colorectal carcinomas are still missing. Thus, a better understanding of the cellular and molecular basis of therapy resistance is a central requirement for the development of improved diagnostic and therapeutic tools. Increased resistance to apoptosis of tumor cells is frequently characterized by a diminished expression of the pro-apoptotic members of the Bcl-2 protein family. In our preliminary experiments we found that the microRNA-210 (miR-210) is regulated by ionizing irradiation and that an increased expression of miR-210 plays an important role in the initiation of apoptosis in colon cancer cells. Our findings indicate that the cytotoxic effect of miR-210 is exerted by the pro-apoptotic Bim protein, and that Bim expression is induced by a miR-210-mediated increase in ROS with subsequent activation of the CHOP and FOXO transcription factors. In the proposed project we aim at demonstrating that the miR-210 dependent induction of Bim expression is mediated by suppression of NDUFA4 and SDHD (both subunits of the complexes of the respiratory chain) with consecutive inhibition of the corresponding respiratory chain complex and by increased ROS generation. Furthermore, we will investigate (both in cell culture and in animal experiments) to which extent this particular mechanism as well as a general modulation of the oxidative metabolic activity regulates the cellular resistance to apoptosis. Besides, our preliminary experiments demonstrate a radiation dependent regulation of miR-210. In this context, we will analyze which transcription factors are involved in this modulation. Additionally, the effects of radiation on miR-210 expression in non-tumorigenic tissue will be investigated. Moreover, we will analyze the prognostic and predictive value of the expression of miR-210 and Bim in a large collection of human colorectal cancer samples (n = 1250).