Department of Biomedical Sciences, Cornell University, Ithaca, New York 14853-6401, USA.
Clin Cancer Res. 2010 Feb 15;16(4):1119-28. doi: 10.1158/1078-0432.CCR-09-2642. Epub 2010 Feb 9.
The miR-34 family is directly transactivated by tumor suppressor p53, which is frequently mutated in human epithelial ovarian cancer (EOC). We hypothesized that miR-34 expression would be decreased in EOC and that reconstituted miR-34 expression might reduce cell proliferation and invasion of EOC cells.
miR-34 expression was determined by quantitative reverse transcription-PCR and in situ hybridization in a panel of 83 human EOC samples. Functional characterization of miR-34 was accomplished by reconstitution of miR-34 expression in EOC cells with synthetic pre-miR molecules followed by determining changes in proliferation, apoptosis, and invasion.
miR-34a expression is decreased in 100%, and miR-34b*/c in 72%, of EOC with p53 mutation, whereas miR-34a is also downregulated in 93% of tumors with wild-type p53. Furthermore, expression of miR-34b*/c is significantly reduced in stage IV tumors compared with stage III (P = 0.0171 and P = 0.0029, respectively). Additionally, we observed promoter methylation and copy number variations at mir-34. In situ hybridization showed that miR-34a expression is inversely correlated with MET immunohistochemical staining, consistent with translational inhibition by miR-34a. Finally, miR-34 reconstitution experiments in p53 mutant EOC cells resulted in reduced proliferation, motility, and invasion, the latter of which was dependent on MET expression.
Our work suggests that miR-34 family plays an important role in EOC pathogenesis and reduced expression of miR-34b*/c may be particularly important for progression to the most advanced stages. Part of miR-34 effects on motility and invasion may be explained by regulation of MET, which is frequently overexpressed in EOC.
miR-34 家族直接被肿瘤抑制因子 p53 转录激活，而 p53 在人类上皮性卵巢癌（EOC）中经常发生突变。我们假设 EOC 中 miR-34 的表达会降低，并且恢复 miR-34 的表达可能会降低 EOC 细胞的增殖和侵袭能力。
通过定量逆转录-PCR 和原位杂交技术，在 83 个人类 EOC 样本中检测 miR-34 的表达。通过合成 pre-miR 分子在 EOC 细胞中重建 miR-34 的表达，然后确定增殖、凋亡和侵袭的变化，来实现对 miR-34 的功能进行特征描述。
miR-34a 在有 p53 突变的 EOC 中表达下调 100%，miR-34b*/c 在有 p53 突变的 EOC 中下调 72%，而 miR-34a 在野生型 p53 的肿瘤中也下调 93%。此外，与 III 期相比，IV 期肿瘤中 miR-34b*/c 的表达显著降低（分别为 P=0.0171 和 P=0.0029）。此外，我们观察到 mir-34 的启动子甲基化和拷贝数变异。原位杂交显示 miR-34a 的表达与 MET 免疫组织化学染色呈负相关，这与 miR-34a 的翻译抑制一致。最后，在 p53 突变的 EOC 细胞中进行 miR-34 重建实验导致增殖、运动性和侵袭性降低，后者依赖于 MET 的表达。
我们的工作表明 miR-34 家族在 EOC 的发病机制中发挥着重要作用，miR-34b*/c 的表达降低可能对进展到最晚期阶段尤为重要。miR-34 对运动性和侵袭性的部分影响可能是通过调节 MET 来解释的，MET 在 EOC 中经常过表达。
