靶向沉默HK-Ⅱ和VEGF基因抵抗肝转移瘤TACE术后残瘤存活的分子机制及活体成像研究

Under the hypoxic conditions, the enhancement of glycolysis and tumor angiogenesis plays a key role in tumor residual of liver metastases after TACE. Anti-angiogenesis by inhibition of VEGF alone does not achieve the desired long-term effect and has potential risks in tumor metastases. The reason may be related to the enhancement of glycolysis in hypoxia conditions in tumor. Hexokinase (HK)-Ⅱ is a key rate-limiting glycolytic enzyme. The overexpression of HK-Ⅱ in tumor tissue is able to supply sufficient energy even in hypoxic conditions. Preliminary studies have shown that tumor cell growth can be inhabited more effectively by targeted silencing HK-Ⅱ in hypoxic conditions than that in normoxic conditions. Then, whether can we obtain synergistic therapeutic effect through targeted silencing HK-Ⅱ and VEGF genes in hypoxic conditions? This project is planned to construct siRNA adenovirus vector of HK-Ⅱ and VEGF and to transfect colon cancer cells and animal model of liver metastases. Then TACE treatment will be available to animal model, the effect of targeted silencing HK-Ⅱ and VEGF genes to tumor's residual and metastases will be observed dynamically by bioluminescence imaging, and the molecular mechanism will be explored. This project might be expected to solve the important problem of recurrence and metastases in residual tumor after TACE and to improve the long-term survival rate in patients with liver metastases.

缺氧条件下，糖酵解增强及肿瘤微血管生成在肝转移瘤TACE术后残瘤存活中起关键的作用。单纯抑制VEGF的抗血管生成并未取得理想的中远期疗效，且有诱发肿瘤转移的风险，原因可能与肿瘤缺氧导致糖酵解增强有关。HK-Ⅱ是糖酵解途径中的限速酶，它在肿瘤组织中过表达，使得肿瘤组织在缺氧的情况下，仍能保证足够的能源。前期研究显示抑制HK-Ⅱ在缺氧条件下较在常氧条件下能更有效的抑制肿瘤细胞的生长。那么，在靶向抑制VEGF的同时，能否通过抑制HK-Ⅱ对糖酵解进行干预以获得协同作用？本研究拟构建HK-Ⅱ和VEGF的siRNA腺病毒载体并转染结肠癌细胞及其肝转移瘤模型，并对模型采用TACE治疗，通过生物发光成像动态观察缺氧状态下靶向抑制HK-Ⅱ和VEGF对TACE术后残瘤存活及转移的影响，并初步探讨其作用机制，以解决TACE术后缺氧导致残瘤复发转移的重要问题，以期提高肝转移瘤患者的长期生存率。

缺氧诱导HK-Ⅱ和VEGF高表达在结直肠癌肝转移瘤介入治疗后肿瘤残留复发中起着关键作用。单纯抑制VEGF的抗血管生成并未取得理想的中远期疗效，且有诱发肿瘤转移的风险，原因可能与肿瘤缺氧导致糖酵解增强有关。HK-Ⅱ是糖酵解途径中的限速酶，它在肿瘤组织中过表达，使得肿瘤组织在缺氧的情况下，仍能保证足够的能源。本课题组研究发现，利用siRNA干扰技术，同时敲低HK-Ⅱ和VEGF的表达能够显著的促进结直肠癌细胞的凋亡。然而腺病毒载体存在着较明显的缺陷，包括靶向性、安全性和多效性等。我们将进一步的利用核酸纳米颗粒的制备技术，以3WJ模体为骨架，共价连接HK-Ⅱ和VEGF siRNA、叶酸和FITC，构建兼具靶向与治疗功效的核酸纳米颗粒并转染结直肠癌细胞及其肝转移瘤动物模型，动态观察核酸纳米颗粒对结直肠癌肝转移瘤的疗效。在结直肠癌侵袭转移中NPTX2、HSP27、HKⅡ和VEGF基因可能存在着网络式协同的调控作用是本项目的另一发现。以上研究有望为肝转移瘤提供新的治疗思路，以期提高肝转移瘤患者的长期生存率。