MCF-7 serum withdrawal (Agilent mouse 60K microarray). MCF-7 serum withdrawal (Agilent mouse 60K microarray)

3’deoxyadenosine, also known as cordycepin, has been widely researched as a potential treatment for cancer, yielding promising results in tissue culture as well as in pre-clinical models. A wide variety of mechanisms of action have been proposed, with little agreement between different studies. Here, we confirm that cordycepin triphosphate is likely to be the active metabolite of cordycepin. Data from single and high throughput experiments showed that cordycepin represses growth factor induced gene expression. Bioinformatic analysis, quantitative PCR and western blotting confirmed that cordycepin blocks the PI3K/AKT/mTOR and/or MEK/ERK pathways in 6 cell lines. Effects of cordycepin on translation through mTOR pathway repression were detectable within 30 minutes, indicating a rapid process. Our data show that cordycepin has a broadly similar mechanism of action in all cell lines studied and indicate that its therapeutic target is a cordycepin triphosphate sensitive molecule that is required for growth factor signal transduction. Overall design: MCF-7 cells were incubated for four hours in serum free medium (serum withdrawal) or kept in the medium in which they were seeded (control), total RNA was isolated and microarray analysis performed.

3'-脱氧腺苷（3’deoxyadenosine）又称虫草素（cordycepin），作为潜在抗癌治疗手段被广泛研究，在组织培养与临床前模型中均已取得颇具潜力的研究结果。目前学界已提出多种作用机制假说，但不同研究间鲜有共识。本研究证实，三磷酸虫草素（cordycepin triphosphate）大概率为虫草素的活性代谢产物。单样本及高通量实验数据显示，虫草素可抑制生长因子诱导的基因表达。通过生物信息学分析、定量PCR（quantitative PCR）与蛋白质印迹（western blotting）实验，我们证实虫草素可在6种细胞系中阻断PI3K/AKT/mTOR及/或MEK/ERK信号通路。虫草素通过抑制mTOR通路对翻译过程产生的调控效应可在30分钟内被检测到，提示该过程十分迅速。本研究数据表明，虫草素在所有受试细胞系中具有大体一致的作用机制，同时提示其治疗靶点为一类对生长因子信号转导必需、且对三磷酸虫草素敏感的分子。总体实验设计：将MCF-7细胞分为两组，一组置于无血清培养基中孵育4小时（血清剥夺组），另一组保留原接种培养基作为对照组，随后提取总RNA并开展微阵列（microarray）分析。