Chen Jianfang, Ding Zhenyu, Peng Yonghai, Pan Feng, Li Jianjun, Zou Lan, Zhang Yanling, Liang Houjie
Department of Oncology and Southwest Cancer Center, Southwest Hospital, Third Military Medical University, Chongqing, China.
Department of Oncology and Southwest Cancer Center, Southwest Hospital, Third Military Medical University, Chongqing, China; Department of Oncology, General Hospital of Shenyang Military Region, Shenyang, Liaoning, China.
PLoS One. 2014 Jun 5;9(6):e98882. doi: 10.1371/journal.pone.0098882. eCollection 2014.
Multidrug resistance (MDR) is one of the major reasons chemotherapy-based treatments fail. Hypoxia is generally associated with tumor chemoresistance. However, the correlation between the heterodimeric hypoxia-inducible factor-1 (HIF-1) and the multidrug resistance (MDR1) gene/transporter P-glycoprotein (P-gp) remains unclear. This study aims to explore the molecular mechanisms of reversing colon cancer MDR by focusing on the target gene HIF-1α.
A chemotherapeutic sensitivity assay was used to observe the efficiency of MDR reversal in LoVo multicellular spheroids (MCS). The apoptotic level induced by different drugs was examined by flow cytometry (FCM). Binding of HIF-1α to the MDR1 gene promoter was evaluated by Chromatin immunoprecipitation (ChIP). The relationship between HIF-1α/P-gp expression and sensitivity to chemotherapy was analyzed.
The sensitivity of LoVo MCS to all four chemotherapy drugs was decreased to varying degrees under hypoxic conditions. After silencing the HIF-1α gene, the sensitivities of LoVo MCS to all four chemotherapy drugs were restored. The apoptotic levels that all the drugs induced were all decreased to various extents in the hypoxic group. After silencing HIF-1α, the apoptosis level induced by all four chemotherapy drugs increased. The expression of HIF-1α and P-gp was significantly enhanced in LoVo MCS after treatment with hypoxia. Inhibiting HIF-1α significantly decreased the expression of MDR1/P-gp mRNA or protein in both the LoVo monolayers and LoVo MCS. The ChIP assay showed that HIF-1α was bound to the MDR1 gene promoter. Advanced colon carcinoma patients with expression of both HIF-1α and P-gp were more resistant to chemotherapy than that with non expression.
HIF-1α inhibition reverses multidrug resistance in colon cancer cells via downregulation of MDR1/P-gp. The expression of HIF-1α and MDR1/P-gp can be used as a predictive marker for chemotherapy resistance in colon cancer.
多药耐药（MDR）是基于化疗的治疗失败的主要原因之一。缺氧通常与肿瘤化疗耐药相关。然而，异二聚体缺氧诱导因子-1（HIF-1）与多药耐药（MDR1）基因/转运蛋白P-糖蛋白（P-gp）之间的相关性仍不清楚。本研究旨在聚焦靶基因HIF-1α，探索逆转结肠癌多药耐药的分子机制。
采用化疗敏感性试验观察洛伏多细胞球体（MCS）中多药耐药逆转的效率。通过流式细胞术（FCM）检测不同药物诱导的凋亡水平。采用染色质免疫沉淀法（ChIP）评估HIF-1α与MDR1基因启动子的结合情况。分析HIF-1α/P-gp表达与化疗敏感性之间的关系。
在缺氧条件下，洛伏MCS对所有四种化疗药物的敏感性均有不同程度降低。沉默HIF-1α基因后，洛伏MCS对所有四种化疗药物的敏感性得以恢复。缺氧组中所有药物诱导的凋亡水平均有不同程度降低。沉默HIF-1α后，所有四种化疗药物诱导的凋亡水平升高。缺氧处理后，洛伏MCS中HIF-1α和P-gp的表达显著增强。抑制HIF-1α可显著降低洛伏单层细胞和洛伏MCS中MDR1/P-gp mRNA或蛋白的表达。ChIP试验表明，HIF-1α与MDR1基因启动子结合。同时表达HIF-1α和P-gp的晚期结肠癌患者比不表达的患者对化疗更耐药。
抑制HIF-1α可通过下调MDR1/P-gp逆转结肠癌细胞的多药耐药。HIF-1α和MDR1/P-gp的表达可作为结肠癌化疗耐药的预测标志物。
