TPM2通过调节Actin/Microtubule交互作用增强乳腺癌细胞对紫杉醇的敏感性

Breast cancer is one of the most common malignant tumors in women worldwide. Anti-microtubule paclitaxel is widely used in the treatment of breast cancer. The mechanism of paclitaxel resistance and coping strategies are urgent problems to be solved. Our previous study found that actin-binding protein TPM2 was low expression in breast cancer; Hypoxia increased the methylation status of this gene and down-regulated the expression of TPM2; The expression of TPM2 affected the invasion ability of breast cancer, the efficacy of paclitaxel and the prognosis of patients. Thus we hypothesize that: TPM2 binds actin, which regulates the interaction of actin with microtubule and enhances the sensitivity of paclitaxel to breast cancer cells. In order to prove this hypothesis, based on the previous studies, we first demonstrate the relationship between the expression of TPM2 and the efficacy of paclitaxel by using clinical data of breast cancer and TCGA public data, and then reveal the influence of TPM2-mediated paclitaxel sensibility to breast cancer from cell lines and animal levels. Finally, the molecular mechanism of TPM2 enhancing the sensitivity of paclitaxel to breast cancer cells through the regulation of the interaction between actin and microtubule was investigated. Our research will provide a personalized basis for the effective use of paclitaxel treatment of breast cancer.

乳腺癌是全球女性最常见的恶性肿瘤之一，抗微管药物紫杉醇广泛应用于乳腺癌的治疗，紫杉醇的耐药机制和应对策略是急需解决的问题。我们前期研究发现：Actin绑定蛋白TPM2在乳腺癌中呈低表达状态，低氧增强该基因启动子甲基化状态使TPM2表达下调，其表达影响乳腺癌的侵袭能力、紫杉醇的疗效及患者的预后。由此我们假设：TPM2绑定Actin，调节Actin与Microtubule的交互作用,增强紫杉醇对乳腺癌细胞的敏感性。为证明该假说，本课题在前期研究基础上，首先应用乳腺癌临床样本和TCGA公共数据证明TPM2表达与紫杉醇疗效的关系；继而从细胞株和动物水平揭示TPM2介导的紫杉醇对乳腺癌药物敏感性的影响；最后在分子水平揭示TPM2通过调节Actin与Microtubule的交互作用增强紫杉醇对乳腺癌细胞敏感性的机制。为有效应用紫杉醇治疗乳腺癌提供个性化依据。

乳腺癌是全球女性最常见的恶性肿瘤之一，抗微管药物紫杉醇广泛应用于乳腺癌的治疗，其耐药机制和应对策略是急需解决的问题。.应用公共数据、细胞学实验以及蛋白组学实验多学科交叉进行分析研究。应用乳腺癌新辅助化疗公共数数据I-SPY芯片数据进行分析，发现应用紫杉醇化疗获得病理完全缓解的患者，TPM2的表达相对较高。我们又分析了多种乳腺癌的细胞系及相对应的IC50的关系，发现TPM2的表达与紫杉醇的IC50程负相关。为进一步证明TPM2的表达与紫杉醇治疗敏感性的关系，进行了细胞学实验。应用BT-549和UACC-812构建TPM2下调的稳转细胞株，检测其对紫杉醇IC50的变化，发现下调TPM2后IC50明显上升。应用相同剂量的紫杉醇治疗对照组和敲降组，下调TPM2组细胞克隆形成率升高。由于TPM2是Actin绑定蛋白，因此我们从TPM2绑定Actin, 增强紫杉醇对Microtubule的作用从而增强乳腺癌细胞的敏感性的方向进行研究。应用稳转下调TPM2细胞株，相同剂量、时间紫杉醇（15nM，24h）作用于实验组和对照组，应用免疫荧光染色，发现TPM2下调后紫杉醇对微管的聚合能力减弱，表明TPM2下调后乳腺癌细胞对紫杉醇敏感性下降。紫杉醇作用乳腺癌细胞后，分离提取游离微管和结合微管，应用蛋白组学实验检测乙酰化tubulin的表达，进一步证明TPM2高表达，微管的稳定性更强，紫杉醇的作用更敏感。为探索TPM2与actin 的关系，我们下载并分析了TCGA数据，TPM2的表达与α-actin（ACTA2）、γ-actin（ACTG2）表达呈正相关。过表达TPM2后，ACTG1/2相应表达也有所升高。.以上研究初步证实了TPM2通过Actin调节紫杉醇对Microtubule的作用从而增强对乳腺癌细胞敏感性。为有效应用紫杉醇治疗乳腺癌提供个性化依据。