抗能量药物对胃癌细胞生长和转移的影响及其作用机制

Tumor cells mainly rely on glycolysis to gain energy, inhibition of glycolysis can induce cell apoptosis. Our group was first discovered on anti-energetic drug in vitro induced apoptosis of gastric cancer cells, and in the preliminary experiment, anti-energetic drug in vivo can significantly inhibit the growth of human gastric cancer in nude mice subcutaneous implanted tumor, but the action mechanism of anti-energetic drug has not yet been clearly. As a result of the rate-limiting enzyme plays a key role in glycolysis, thus we hypothesize that anti-energetic drug is by inhibiting the rate-limiting enzyme activity, promoting apoptosis gene expression and inhibiting tumor lymphatic vessels, thereby inhibiting growth and metastasis of gastric cancer cells. According to this theory, the project selects two kinds of anti-energetic drugs ( 3 - bromine pyruvic acid, sodium citrate) to study respectively. Application of gastric cancer cell lines, human gastric cancer in nude mice subcutaneous implanted tumor model and human gastric cancer in nude mice orthotopically metastatic model, observating the function of anti-energetic drug in vivo and in vitro to inhibit gastric cancer cells growth, and in vivo to inhibit metastasis, to determine its effect on glycolysis rate-limiting enzyme influence. Through the analysis of the relations, rate-limiting enzyme activity and several apoptotic genes expression and expression of VEGF-C, elucidating the action mechanism of anti-energetic drug. The results of this study will provide the application of anti-energetic drug in the treatment of gastric cancer provide new experimental basis and research thinking, to establish a new effective tumor therapy method to lay the foundation.

肿瘤细胞主要依赖糖酵解获取能量，抑制糖酵解可诱导细胞凋亡。本课题组首次发现抗能量药物在体外可诱导胃癌细胞凋亡，并在预实验中观察到抗能量药物在体内能显著抑制人胃癌裸鼠皮下移植瘤生长，但抗能量药物的作用机制迄今尚未清楚。由于糖酵解限速酶在糖酵解中起关键作用，因此我们推测抗能量药物是通过抑制限速酶活性、促进凋亡基因表达及抑制肿瘤淋巴管生成，从而抑制胃癌细胞生长和转移。依据这一推测，本项目选用两种抗能量药物（3-溴丙酮酸、柠檬酸钠）分别进行研究。通过应用胃癌细胞株、人胃癌裸鼠皮下移植瘤模型、人胃癌裸鼠原位移植瘤模型，观察抗能量药物在体内、体外抑制胃癌细胞生长和体内抑制转移的作用，确定其对糖酵解限速酶的影响；通过分析限速酶活性与几种凋亡基因及VEGF-C表达的关系，阐明抗能量药物的作用机制。本研究结果将为应用抗能量药物治疗胃癌提供新的实验依据和研究思路，为建立一种新的有效的肿瘤治疗方法奠定基础。

胃癌是最常见的恶性肿瘤之一，虽然手术技术，化放疗等综合治疗取得了巨大的进步，但 5年生存率仍不理想。因此，寻找新的治疗靶点，提高患者生存率迫在眉睫。80年前，Warburg发现恶性肿瘤细胞在有氧条件仍然主要依赖糖酵解来获取能量，而这一特点，为抗肿瘤药物研究提供了一个新靶点。申请人前期研究结果表明，抗能量药物在可抑制恶性胸膜间皮瘤增殖，但对胃癌细胞，特别是体内应用能否发挥抗肿瘤的作用未见报道。本项目通过应用胃癌细胞株SGC-7901及MGC-803，进行3-溴丙酮酸及柠檬酸钠的体外实验，检测细胞凋亡、周期，糖酵解关键酶，糖酵解产物，凋亡蛋白等指标，初步阐述药物抑制胃癌细胞增殖机制。然后，通过建立SGC-7901细胞系裸鼠皮下移植瘤及胃原位移植瘤模型，计算肿瘤生长抑制率，应用PET/CT扫描及相关凋亡基因检测，进一步阐明3-溴丙酮酸及柠檬酸钠抑制肿瘤生长的机制。结果显示：体外应用3-溴丙酮酸及柠檬酸钠能有效抑制SGC-7901及MGC-803人胃癌细胞的增殖，引起细胞周期阻滞，降低细胞糖酵解酶HK及PFK-1活性，减少ATP及乳酸产量，导致细胞能量枯竭，酸性微环境破坏，并可上调Bax表达、下调Bcl-2，Survivin表达，引起Cyt-C释放，激活caspase-3，促进细胞凋亡。体内实验成功通过构建裸鼠皮下移植瘤及胃原位移植瘤模型, 分别于皮下及腹腔内给药，结果表明3-溴丙酮酸及柠檬酸钠具有较高的生物安全性，体内应用可以抑制移植瘤的生长，降低胃癌细胞糖酵解水平，并能激活caspase-9、caspase-3级联反应，最终诱导胃癌肿瘤细胞的凋亡。本研究已按计划按时完成，证明了3-溴丙酮酸及柠檬酸钠在体内外对胃癌细胞均有明显抑制作用，阐明了3-溴丙酮酸及柠檬酸钠体内外抑制胃癌细胞的机制，可为胃癌治疗的探索提供一些新的研究思路。