lncRNA参与miR-29b-3p调控乳腺癌骨转移的机制研究

Clinical treatment for bone metastasis of breast cancer remains a major challenge. Many studies demonstrated that miRNAs play an important role in the pathogenesis of bone metastasis of breast cancer through regulating osteoclast differentiation and activity. Our previous studies found that miR-29b-3p could depress the differentiation and activity of osteoclast by inhibiting the expression of NFATc-1, thereby relieving the occurrence of bone metastases in breast cancer. Now, we have brought the concept of ceRNA in this study, and proved that LINC00511 competitively binds miR-29b-3p to regulate osteoclastic differentiation and bone resorption through lncRNA chip, bioinformatics prediction, dual-Luciferase reporter gene system, RIP technology, Q-PCR, TRAP staining method, etc. Furthermore, we will employ a mouse model of breast cancer bone metastasis as the research project, and will elucidate the mechanism of LINC00511, as the ceRNA of miR-29b-3p, in the occurrence of breast cancer bone metastasis via the targeted therapy mediated by osteoclast targeting delivery system, site-specific gene knockout technique, etc. These will provide theoretical and experimental basis for verifying the mechanism of breast cancer bone metastasis and the clinic treatment in breast cancer bone metastasis.

乳腺癌骨转移的临床治疗仍面临着严峻挑战。研究结果表明miRNAs和lncRNAs调控的破骨细胞分化和活性在肿瘤骨转移发病中起着重要的作用。我们前期研究发现miR-29b-3p可通过抑制NFATc-1的表达，降低了破骨细胞的分化和活性，从而减缓了乳腺癌骨转移的发生。本项目现将lncRNAs作为ceRNA的概念引入，通过lncRNA芯片、生物信息学预测、双荧光素酶报告基因系统、RIP技术、Q-PCR和TRAP染色法等实验证实了LINC00511通过竞争性结合miR-29b-3p，调控了破骨细胞的分化和骨吸收功能。进一步我们拟以乳腺癌骨转移模型小鼠为研究对象，利用破骨细胞靶向递送系统介导的靶向治疗、靶基因的特异性敲除技术等，阐明LINC00511作为miR-29b-3p的ceRNA参与乳腺癌骨转移发生的分子机制。这将为乳腺癌骨转移的发生发展机制及其临床治疗提供了理论基础和实验依据。

乳腺癌骨转移的临床治疗一直以来面临很大的挑战。研究表明非编码RNA（如：miRNAs和lncRNAs等）调控的破骨细胞分化和活性在肿瘤骨转移发病中起着重要的作用。我们的研究发现：在乳腺癌溶骨性骨转移患者的骨组织中和移植性人乳腺癌骨转移裸鼠的破骨细胞中，miR-29b-3p的表达水平是显著性降低的；并且利用瞬时转染技术提高或降低miR-29b-3p后，可以显著性抑制破骨细胞分化和骨吸收功能。通过生物信息学软件预测和分子实验验证，发现miR-29b-3p可通过靶向抑制NFATc-1调控了破骨细胞的分化；还发现miR-29b-3p可通过靶向抑制MMP-2来抑制乳腺癌MDA-MB-231细胞迁移和侵袭。进一步，我们通过Arraystar第三代LncRNA芯片技术筛选出在骨转移样本中差异表达的lncRNAs，并通过Q-PCR、生物信息学软件预测以及双萤光素酶报告基因系统等分子实验验证，发现实验LINC00511可与miR-29b-3p直接结合，促进了NFATc-1的表达和破骨细胞的分化。最后，我们以移植性人乳腺癌骨转移裸鼠为研究对象，利用实验室构建的骨吸收表面靶向递送系统(D-Asp8)-liposome将agomir-29b-3p（D-Ago-29b-3p）或LINC00511-siRNA递送到破骨细胞，发现agomir-29b-3p或LINC00511-siRNA可显著抑制移植性人乳腺癌骨转移裸鼠骨组织的骨破坏和肿瘤骨转移。这一研究为乳腺癌骨转移的发生发展机制、诊断分子标志物的筛选以及临床治疗提供了理论基础和实验依据。.通过本项目的实施，申请人以通讯作者身份，现已在Cell Death & Disease、European Journal of Medicinal Chemistry、Oxidative Medicine and Cellular Longevity、Frontiers in Cell and Developmental Biology等期刊杂志发表学术论文14篇。