Exploring miR-29 in melanoma progression and prevention

项目总结 MiR-29家族由miR-29a、miR-29b1、miR-29b2和miR-29c组成。 由两个miRNA簇(miR-29a~b1和miR-29b2~c)编码，在 各种人类癌症。我们发现，矛盾的是，致癌基因BRAFV600E诱导了这两种基因的转录 MAPK途径激活(miR-29a~b1)或致癌应激介导的黑素细胞簇 P53活化(miR-29b2~c)。这会产生一种屏障来防止黑色素瘤的发展，我们在一个 海绵结构使miR-29家族失活的基因工程小鼠模型 黑色素瘤进展。有趣的是，工程化的人类细胞系模型和来自nevi的人类表达数据 黑色素瘤表明，黑色素瘤的进展与miR-29b2~c的表达降低有关。 基于这些发现，我们假设关键的miR-29靶点的去抑制和过度表达 促进黑色素瘤的发展，这些靶点是黑色素瘤的弱点，可以被利用 接受治疗。我们已经建立了一个高通量的基因工程小鼠建模平台来评估 MiR-29靶点在黑色素瘤中的作用并评估其作为治疗靶点的潜力。在这项提案中，我们 将广泛利用这一平台首先测试miR-29的表达是否恢复，从而抑制所有miR-29 29个靶点损害黑色素瘤的发展。我们将进一步评估单个miR-29目标的沉默是否会导致 对黑色素瘤的发生有显著影响。我们已经鉴定了致癌转录因子MYBL2是一个博纳基因 真核miR-29靶点，其沉默可降低体外黑色素瘤细胞系的侵袭性。我们将测试是否去- 抑制的MYBL2通过沉默MYBL2在黑色素瘤小鼠模型中介导miR-29缺失的影响 通过miR-29灭活。此外，我们将在具有正常miR-29活性的模型中使MYBL2沉默，以确定是否 MYBL2是黑色素瘤的一种常见易感性。为了确保与人类黑色素瘤的相关性，我们将补充 这些基因工程小鼠模型在人类黑色素瘤PDX上进行了实验。我们建议的研究 将使用我们最新开发的鼠标建模平台作为管道，系统地评估miR-29目标 总的来说，特别是MYBL2，作为黑色素瘤的治疗靶点具有进一步探索的潜力。

PROJECT SUMMARY The miR-29 family of microRNAs (miRNAs), consisting of miR-29a, miR-29b1, miR-29b2, and miR-29c, is encoded by two miRNA clusters (miR-29a~b1 and miR-29b2~c) and has tumor suppressive properties in a variety of human cancers. We found that, paradoxically, oncogenic BRAFV600E induces the transcription of both clusters in melanocytes through either MAPK pathway activation (miR-29a~b1) or oncogenic stress-mediated p53 activation (miR-29b2~c). This creates a barrier that prevents melanoma development, and we showed in a genetically engineered mouse model that inactivation of the miR-29 family with a sponge construct promotes melanoma progression. Interestingly, engineered human cell line models and human expression data from nevi and melanomas show that melanoma progression is associated with decreased expression of miR-29b2~c. Based on these findings we hypothesize that de-repression and overexpression of critical miR-29 targets promotes melanoma development and that such targets are vulnerabilities of melanoma that can be exploited for therapy. We have established a high-throughput genetically engineered mouse modeling platform to evaluate the role of miR-29 targets in melanoma and to assess their potential as therapeutic targets. In this proposal, we will extensively use this platform to first test if restoration of miR-29 expression and, thus, repression of all miR- 29 targets impairs melanoma development. We will further evaluate if silencing of a single miR-29 target elicits significant effects on melanomagenesis. We have identified the oncogenic transcription factor MYBL2 as a bona fide miR-29 target whose silencing reduces the aggressiveness of melanoma cell lines in vitro. We will test if de- repressed MYBL2 mediates the effects of miR-29 loss by silencing MYBL2 in a melanoma mouse model driven by miR-29 inactivation. Moreover, we will silence MYBL2 in a model with normal miR-29 activity to determine if MYBL2 is a general vulnerability of melanoma. To ensure relevance to human melanoma, we will complement these genetically engineered mouse models with experiments in human melanoma PDXs. Our proposed studies will use our newly developed mouse modeling platform as a pipeline to systematically evaluate if miR-29 targets in general, and MYBL2 specifically, have the potential to be further explored as therapeutic targets in melanoma.