新生血管在肿瘤的生长、侵袭和转移中发挥着关键作用。研究表明，靶向新生血管的治疗策略对多种癌症都具有显著疗效。血管生成素2是新生血管过程中的重要因子。靶向血管生成素2 的方法是一种极具潜力的癌症治疗策略。虽然miRNA逐渐成为新生血管的重要调控因子，但是尚未有关于调控血管生成素2的miRNA的报道。在本研究中，我们首次报道了miR-542-3p可以靶向血管生成素2，通过抑制血管生成素2的促新生血管活性从而抑制肿瘤进展。在体外培养的血管内皮细胞中，miR-542-3p能够结合血管生成素2 mRNA的3’非翻译区，从而抑制血管生成素2 mRNA的翻译。在血管内皮细胞中过表达miR-542-3p可以抑制其新生血管活性。对荷瘤鼠施加miR-542-3p能够抑制肿瘤生长、新生血管和转移。此外，在人的乳腺癌临床样本中，miR-542-3p的表达水平与癌症的恶性程度负相关。为了进一步了解miR-542-3p抑制新生血管活性的机理，我们筛选了肿瘤细胞分泌的、可能参与调控miR-542-3p表达的胞外因子。研究发现，肿瘤细胞分泌的血管生长素可以下调血管内皮细胞的miR-542-3p。无论在体外还是体内模型中，对肿瘤细胞过表达血管生长素都可以通过抑制miR-542-3p激活内皮细胞的新生血管活性。深入研究发现，血管生长素可以抑制CCAAT/enhancer binding protein beta (CEBPB)和POU class 2 homeobox 1 (POU2F1)，后两者都是miR-542-3p的转录因子，从而揭示了一个新的肿瘤细胞和内皮细胞之间的新生血管调控机制。此外，乳腺肿瘤中的血管生长素水平与疾病进展具有显著的相关性。乳腺癌患者血浆内的血管生长素含量在转移的患者中高于非转移患者。综上，我们的研究揭示了一个新的肿瘤新生血管调控机制：肿瘤细胞分泌的血管生长素作用于血管内皮细胞，抑制转录因子CEBPB和POU2F1的表达，进而下调miR-542-3p，最终激活促新生血管蛋白血管生成素2的表达。因此，血管生长素和miR-542-3p是潜在的抗新生血管治疗靶点和监测乳腺癌疾病进展的生物标志物。
Angiogenesis is essential for tumor growth, invasion, and metastasis. Therapeutic strategies for targeting angiogenesis have been proven as successful treatments for divergent cancers. Angiopoietin2 (Angpt2) plays a critical role in angiogenesis. Whereas miRNAs are emerging as important modulators of angiogenesis, regulation of Angpt2 by miRNAs has not been established. Here we report that Angpt2 is targeted by a microRNA, miRNA-542-3p, which inhibits tumor progression by impairing Angpt2’s pro-angiogenic activity. In cultured endothelial cells, miR-542-3p inhibited translation of Angpt2 mRNA by binding to its 3'UTR, and addition of miR-542-3p to cultured endothelial cells attenuated angiogenesis. Administration of miR-542-3p to tumor-bearing mice reduced tumor growth, angiogenesis, and metastasis. Moreover, the level of miR-542-3p in primary breast carcinomas inversely correlated with clinical progression in primary tumor samples from stage III and IV patients. To further explore the mechanism of miR-542-3p induced angiogenic inhibition, we screened for tumor-derived factors that were responsible for miR-542-3p alteration in endothelial cells and found that tumor cell secreted angiogenin (ANG) downregulated miR-542-3p in endothelial cells. Overexpression of ANG in tumor cells facilitated angiogenic activation in both in vitro and in vivo models via inhibition of miR-542-3p. Furthermore, our results showed that ANG could suppress CEBPB and POU2F1, which were transcription factors for miR-542-3p, suggesting a novel tumor-endothelial angiogenesis switch. In addition, the level of ANG in primary breast carcinomas correlated with clinical progression. Serum levels of ANG were associated with metastatic development of breast cancer patients. Together, these findings reveal a novel regulatory pathway whereby tumor-derived ANG directly activates angiogenesis through inhibition of miR-542-3p, which directly targets the key angiogenesis-promoting protein Angpt2. Thus, ANG and miR-542-3p may represent promising targets for anti-angiogenic therapy and potential markers for monitoring disease progression of breast cancer.
