CACNA2D3对食管鳞癌化疗敏感性的影响及其机制研究

Esophageal squamous cell carcinoma (ESCC) is one of the most common solid tumors in the world with poor prognosis. Deletion of chromosome 3p is one of the most frequent chromosomal alterations in ESCC, suggesting the existence of one or more tumor suppressor genes (TSGs) at this region. We found that deletion of 3p21 was frequently detected and one candidate tumor suppressor gene CACNA2D3(calcium channel, voltage-dependent, alpha 2/delta subunit 3) was identified within the region. Functional studies showed that CACNA2D3 was able to suppress tumorigenicity of ESCC cells both in vitro and in vivo , including foci formation, colony formation in soft agar and tumor formation in nude mice. In addition, CACNA2D3 could up-regulate intracellular free cytosolic Ca2+ and subsequently promote the apoptosis induced by Staurosporine (STS). However, the effect of CACNA2D3 gene on chemosensitivity of esophageal squamous cell carcinoma is still not clear. We will use immunohistochemistry, Western blot, stable transfection, RNA interference and other methods to verify the following three contents: 1) In vivo and in vitro experiments to discover the relationship between CACNA2D3 and the chemosensitivity of esophageal squamous cell carcinoma; 2) Investigate whether CACNA2D3 through its effect on cell cycle and apoptosis affect the chemosensitivity of esophageal squamous cell carcinoma; 3) Discover the relationship between cisplatin-induced apoptosis promoted by CACNA2D3 and the mitochondrial pathway. Further researches on the effect of CACNA2D3 on chemosensitivity of esophageal squamous cell carcinoma will indicate the potential value of CACNA2D3 as a sensitizer in esophageal squamous cell carcinoma treatment.

食管癌是世界上最常见的恶性肿瘤之一。染色体3p区缺失是食管癌最常见的染色体变异，我们前期从3p21区域分离出候选抑癌基因CACNA2D3，发现CACNA2D3能显著抑制细胞的增殖能力和裸鼠体内成瘤能力，并能上调细胞内钙离子浓度促进星形孢菌素诱导的细胞凋亡。然而，CACNA2D3基因对食管癌化疗敏感性的影响目前还不清楚。我们将采用免疫组化、Western blot、稳定转染、RNA干扰等手段，从分子、细胞、组织以及动物水平等多层次验证以下三个内容：1）体内、体外实验确定CACNA2D3与食管癌化疗敏感性之间的关系；2）探讨CACNA2D3是否通过作用于细胞周期和细胞凋亡影响食管癌化疗敏感性；3）明确CACNA2D3促进顺铂诱导的细胞凋亡与线粒体途径的关系。通过研究CACNA2D3对食管癌细胞化疗敏感性的影响，为 CACNA2D3有可能作为化疗增敏剂应用于食管癌的临床治疗提供理论依据。

钙离子通道蛋白（CACNA2D3）的异常表达与食管癌的生长转移、复发和预后密切相关。低表达CACNA2D3的食管癌患者预后更差，目前CACNA2D3基因与食管癌化疗敏感性之间的关系没有报道。我们研究发现CACNA2D3异常表达影响化疗药物的敏感性，进而导致肿瘤耐药的发生。过表达CACNA2D3可以增强食管癌细胞对铂类化疗药物的化疗敏感性；沉默表达CACNA2D3后则可诱导食管癌细胞对顺铂的耐药；Annexin V-FITC和PI双染色发现过表达CACNA2D3增加顺铂诱导的细胞凋亡率并且加剧线粒体膜电位的下降。裸鼠体内实验证明相对于顺铂单独用药组，CACNA2D3和顺铂联合治疗可以更好地抑制食管癌细胞在动物体内的生长；分子机制研究发现PI3K/AKT/mTOR信号通路与CACNA2D3引起的化疗敏感性增加相关。本项目阐明了抑癌基因CACNA2D3在食管癌细胞生长及化疗敏感性的作用，为临床应用CACNA2D3联合化疗药物治疗食管癌提供理论和实验基础。