miRNA-143靶向调控己糖激酶-2肿瘤治疗的分子影像可视化评估

In cancer cells, glucose is preferentially metabolized by aerobic glycolysis, which differs from mitochondrial oxidative phosphorylation in normal, non-tumorigenic cells. This phenomenon, termed as the Warburg effect, is characterized by increased glycolysis and lactate production regardless of oxygen availability. Based on aerobic glycolysis accompanied by increased glucose uptake, a method named as [18F]Fluorodeoxyglucose Positron Emission Tomography (18F-FDG PET) imaging, combined with computer tomography (PET/CT), has been used worldwide as a diagnostic tool to detect malignant tumors, determine tumor stage, and evaluate clinical therapeutic effect. Moreover, targeting cancer metabolism is being extensively investigated as anti-neoplasia strategies. Hexokinases catalyze the first and irreversible step of glucose metabolism (ATP-dependent phosphorylation of glucose to yield glucose-6-phosphate). Hexokinase-2 (HK-2) is the major isozyme that is overexpressed in tumors and contributes to accelerated glucose uptake and accumulation of glucose-6-phosphate in cancer cells, and thus it is documented as a pivotal player in the Warburg effect and has been used as a metabolic target for cancer therapeutic development. miRNAs are small, non-coding RNAs that negatively regulate protein-coding genes at the post-transcriptional level. The causal roles of miRNAs in cancer have been well documented and miRNA-based anticancer therapies are in development. Our previous results indicate that HK-2 is a target of miR-143. And our 18F-FDG PET/CT imaging assay showed that miR-143 represses 18F-FDG uptake in xenograft breast tumors through targeting HK-2. In this research plan, we will use miR-143 and HK-2 as cancer therapeutic targets, and explore the potential application of miR-143-mediated inhibition of HK-2 in cancer therapeutics. We will assess the early response to treatment dynamically through non-invasive molecular imaging method, pave a way for optimizing the therapeutic and assessment window. Our obtained results may provide theoretical information and useful methods for both of miRNA-based and metabolic target-based anticancer therapies as well as specific real-time dynamic and visible treatment assessment.

肿瘤细胞即使在有氧条件下也以糖酵解为主要产能方式，即Warburg效应。基于该现象建立的18F-FDG PET/CT显像技术已广泛用于肿瘤诊断、分期及疗效评估，而靶向肿瘤糖代谢通路的肿瘤治疗研究正在兴起。己糖激酶2（HK-2）是糖酵解限速酶，在肿瘤细胞中异常高表达，加速肿瘤细胞葡萄糖摄取及G-6-P积聚，可作为肿瘤治疗靶点。miRNA在转录后水平调控靶基因表达，对肿瘤发生发展具有重要调控作用。我们的前期研究显示，miR-143能靶向抑制HK-2表达，18F-FDG PET/CT显像显示，miR-143可通过负调控HK-2抑制葡萄糖摄取。本申请项目拟探索miR-143靶向抑制HK-2在肿瘤治疗中的应用，将通过分子影像方法，早期特异动态评估治疗反应，为筛选最佳治疗及评估时间窗提供依据。项目研究获得结果将为基于miRNA和能量代谢途径的肿瘤治疗及其特异性可视化动态疗效评估提供理论基础和技术方法。

靶向肿瘤糖代谢通路是重要的抗肿瘤治疗策略。因此，本研究进行了miR-143靶向抑制己糖激酶-2(HK2)改变葡萄糖代谢治疗乳腺癌与失分化甲状腺癌的分子影像实验研究。本课题通过qRT-PCR及Western blot验证了肿瘤细胞中miR-143靶向抑制HK2 mRNA及蛋白的表达。通过MTT增殖实验、Transwell迁移实验及Annexin V-FITC/PI双染流式细胞术验证了miR-143抑制肿瘤细胞增殖、迁移并促进其凋亡的作用。通过葡萄糖代谢、乳酸生成检测及体外18F-FDG摄取实验验证了miR-143对肿瘤细胞糖代谢的抑制作用。随后，我们通过尾静脉注射脂质体包裹的miR-143 agomir治疗肿瘤荷瘤裸鼠模型，肿瘤生长曲线分析表明miR-143治疗组肿瘤生长较对照组明显减低。18F-FDG microPET/CT显像可见，miR-143治疗组肿瘤的标准摄取值(SUVmax及SUVmean)在第10天、15天较对照组明显下降。此外，通过监测裸鼠体重变化及血生化指标验证了miR-143体内治疗的安全性。最后免疫组化证实了miR-143治疗组肿瘤HK2、PCNA/Ki-67表达降低而Caspase-3表达上调。Western Blot进一步验证了miR-143治疗组肿瘤HK2表达降低。本研究以miR-143靶向抑制HK2改变葡萄糖代谢治疗肿瘤的策略取得了初步疗效，为基于miRNA和能量代谢途径的肿瘤治疗及其特异性可视化疗效评估提供了新途径。