SGI-1776 钝化Pim-1 对卵巢癌细胞增殖和侵袭的抑制作用及其机制

目的：检测SGI-1776 对卵巢癌HO-8910 细胞增殖、迁移、侵袭的抑制作用及其可能机制。方法：体外培 养卵巢癌HO-8910 细胞；M法和克隆形成法检测SGI-1776 对HO-8910 细胞的增殖抑制作用；PI 染色流式细胞术(ow cytometry，FCM) 检测SGI-1776 对HO-8910 细胞周期分布的影响；Transwell 法检测SGI-1776 对HO-8910 细胞迁移 及侵袭的抑制作用；ELISA，Western 印迹，siRNA/cDNA 转染检测SGI-1776 对HO-8910 细胞增殖、迁移、侵袭抑制 作用的可能分子机制。结果：SGI-1776 在体外对HO-8910 细胞增殖、迁移、侵袭呈现出浓度依赖性的抑制作用，阻 滞细胞周期于G1 期。随Pim-1 激酶活性降低，Pim-1 蛋白表达下调，同时Pim-1 下游蛋白CDK6，pCDK6，CDK4， pCDK4，CDK2 和pCDK2 表达下调，而P21 和P27 蛋白表达上调。用siRNA 干扰沉默Pim-1 基因，则SGI-1776 对 HO-8910 细胞增殖、迁移、侵袭的抑制作用明显加强；用Pim-1-cDNA 转染HO-8910 细胞，则能部分抵消SGI-1776 对细胞增殖、迁移、侵袭的抑制作用。结论：SGI-1776 通过钝化Pim-1 而发挥其抑制卵巢癌HO-8910 细胞增殖、迁 移和侵袭的作用，提示Pim-1 是卵巢癌治疗的新的分子靶。

SGI-1776, an imidazo pyridazine compound,inhibits the proliferation of ovarian cancer cells by inactivating Pim-1

Objective: To investigate the antitumor effect of SGI-1776 on human ovarian cancer HO-8910 cells and its molecular mechanism. Methods: HO-8910 cells were cultured in vitro, and the proliferation inhibitory effects of SGI- 1776 were determined by MTT assay and colony formation assay. The effect of SGI-1776 on the distribution of cell cycle phase was observed by flow cytometry with propidium iodide (PI) staining. The inhibition rate of migration and invasion were valued by transwell cell assay. Multiple molecular techniques, such as ELISA, Western blot, siRNA and cDNA transfection were used to explore the molecular mechanism. Results: SGI-1776 presented dramatic anti-tumor activity against HO-8910 cells in vitro, inhibited the cells proliferation and colony formation, and attenuated the migration and invasion in a dosedependent manner, accompanied by cell cycle arrest in G1 phase. SGI-1776 caused the proliferation inhibition with concomitant decrease in Pim-1 kinase activity, down-regulated the expression of Pim-1 protein and and its downstream genes, such as CDK6, pCDK6, CDK4, pCDK4, CDK2 and pCDK2, and increased the expression of P21 and P27. Down-regulation expression of Pim-1 by siRNA followed SGI-1776 treatment resulted in enhanced cell proliferation inhibition rate and attenuated migration/invasion. Up-regulation of Pim-1 by cDNA transfection attenuated SGI- 1776-induced cell proliferation inhibition and its migration/invasion. Conclusion: Pim-1 mediates the biological effect of SGI-1776 in human ovarian cancer HO-8910 cells, suggesting Pim-1 might be a novel target for human ovarian cancer.