Department of Obstetrics and Gynecology, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi, People's Republic of China.
Biochem Biophys Res Commun. 2010 Feb 26;393(1):144-9. doi: 10.1016/j.bbrc.2010.01.103. Epub 2010 Feb 1.
The release of Notch intracellular domain (NICD) is mediated by gamma-secretase. gamma-Secretase inhibitors have been shown to be potent inhibitors of NICD. We hypothesized that Notch1 is acting as an oncogene in ovarian cancer and that inhibition of Notch1 would lead to inhibition of cell growth and apoptotic cell death in ovarian cancer cells. In this study, expressions of Notch1 and hes1 in four human ovarian cancer (A2780, SKOV3, HO-8910, and HO-8910PM), and one ovarian surface (IOSE 144) cell lines were detected by Western blot and quantitative real-time RT-PCR. The effects of gamma-secretase inhibition (N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester, DAPT) were measured by MTT assay, flow cytometry, ELISA and colony-forming assay. Our results showed that Notch1 and hes1 were found in all the four human ovarian cancer and IOSE 144 cell lines, and they were significantly higher in ovarian cancer cells A2780 compared to another four ovarian cells. Down-regulation of Notch1 expression by DAPT was able to substantially inhibit cell growth, induce G1 cell cycle arrest and induce cell apoptosis in A2780 in dose- and time-dependent manner. In addition, hes1 was found to be down-regulated in dose- and time-dependent manner by DAPT in A2780. These results demonstrate that treatment with DAPT leads to growth inhibition and apoptosis of A2780 cells in dose- and time-dependent manner. These findings also support the conclusion that blocking of the Notch1 activity by gamma-secretase inhibitors represents a potentially attractive strategy of targeted therapy for ovarian cancer.
Notch 细胞内结构域（NICD）的释放是由 γ-分泌酶介导的。γ-分泌酶抑制剂已被证明是 NICD 的有效抑制剂。我们假设 Notch1 在卵巢癌中起癌基因的作用，抑制 Notch1 将导致卵巢癌细胞生长抑制和细胞凋亡。在这项研究中，通过 Western blot 和定量实时 RT-PCR 检测了四种人卵巢癌细胞（A2780、SKOV3、HO-8910 和 HO-8910PM）和一种卵巢表面细胞系（IOSE 144）中 Notch1 和 hes1 的表达。通过 MTT 测定、流式细胞术、ELISA 和集落形成测定来测量 γ-分泌酶抑制（N-[N-（3,5-二氟苯乙酰基）-L-丙氨酰]-S-苯甘氨酸叔丁酯，DAPT）的作用。我们的结果表明，Notch1 和 hes1 均存在于四种人卵巢癌细胞和 IOSE 144 细胞系中，并且在卵巢癌细胞 A2780 中明显高于其他四种卵巢细胞。DAPT 下调 Notch1 表达能够显著抑制 A2780 细胞的生长，诱导 G1 细胞周期阻滞并诱导细胞凋亡，呈剂量和时间依赖性。此外，在 A2780 中，DAPT 还能呈剂量和时间依赖性地下调 hes1 的表达。这些结果表明，DAPT 处理以剂量和时间依赖的方式导致 A2780 细胞的生长抑制和凋亡。这些发现还支持阻断 Notch1 活性的γ-分泌酶抑制剂代表了一种有吸引力的卵巢癌靶向治疗策略的结论。
