探討合併B細胞淋巴瘤蛋白2抑制劑ABT-737和放射線治療對子宮頸癌細胞的標靶策略

本篇研究目的為評估B細胞淋巴瘤蛋白2 (Bcl-2) 抑制劑ABT-737和放射線療法對子宮頸癌的增強效果。根據美國癌症基因圖譜 (TCGA)的統計數據，得知子宮頸癌病患的TNM (原位腫瘤、淋巴轉移、遠端轉移) 分期與Bcl-2 mRNA的表現量具有相關性，我們推測抑制Bcl-2可能有助於減少子宮頸癌的增生，因此將ABT-737加入放射線療法中，選用子宮頸癌細胞株SiHa和Caski 來進行體外模式的實驗，評估此合併療法對腫瘤細胞的增生作用之影響。在本研究中，我們針對TCGA的數據進行統計，儘管未達統計學上的顯著差異，但發現子宮頸癌晚期病患 (Stage III) 的Bcl-2 mRNA表現量有略高於子宮頸癌初期病患 (Stage I) 的現象；接著，我們進一步進行體外模式的實驗，研究結果顯示：使用MTT分析法，發現合併處理ABT-737和放射線療法出現協同性的毒殺作用；使用JC-1單染色法和Annexin V/PI雙染色法並以流式細胞儀分析，發現ABT-737結合放射線療法能誘導細胞凋亡的發生；子宮頸癌細胞株Caski 和SiHa在單獨處理ABT-737或放射線療法會造成粒線體膜電位降低，而合併療法能夠增強膜電位下降的幅度；經共同處理ABT-737和放射線療後，凋亡細胞的比例有顯著性的提升；使用西方墨點法分析，發現合併療法誘導切割型PARP和caspase-7的蛋白表現量增加。綜合上述結果，子宮頸癌細胞株Caski 和SiHa在合併處理ABT-737和放射線療法後，可能透過粒線體膜電位的降低和啟動ROS依賴型切割PARP或caspase-7的路徑，誘導細胞凋亡的發生。本研究認為ABT-737具有潛力作為放射線治療子宮頸癌時合併使用之佐劑。

The aim of this study is to examine the role of ABT-737, an inhibitor of B-cell lymphoma protein 2 (Bcl-2), in enhancing the effect of irradiation on uterine cervical cancer. Based on The Cancer Genomic Atlas (TCGA), we associated the mRNA expression of Bcl-2 with TNM (tumor, node and metastasis) stage of cervical cancer. We infer that Bcl-2 inhibition may reduce the progression of cervical cancer. ABT-737 was added to irradiation to evaluate their effect on cancer cell progression. The SiHa and Caski cancer cells were selected for in vitro assays. In our study, although it didn’t reach a statistical significance, patients with advanced stage III seemed to have higher mRNA expression of Bcl-2 than those with stage I according to TCGA data. ABT-737 and radiation administration induced a synergistic cytotoxic effect according to MTT assay. That ABT-737 and irradiation induced apoptosis was determined by JC-1 staining and annexin V/propidium iodide flow cytometry analyses. In addition to the individual effect of ABT-737 and irradiation on the decrease of mitochondrial membrane potential, their cotreatment augmented this effect in Caski and SiHa cells. The apoptotic percentages were significantly increased in cells treated with ABT-737 and irradiation. In Western blotting, expressions of cleaved forms of PARP and caspase-7 were induced by combined treatment. In conclusions, ABT-737 and irradiation may induce apoptosis via mitochondria membrane potential loss and ROS-dependent cleaved PARP/caspase-7 pathway in Caski and SiHa cells. This study indicates that ABT-737 may be a potential irradiation adjuvant against cervical cancer.