The RNA-activation platform to treat X-linked disease in a locus-specific manner.

描述（由申请人提供）：所提出的研究将通过应用于我的实验室开发的原始“RNA激活”技术（RNA-a;专利申请中，PCT-US 2011 -065939）的技术创新来开发上调X连锁基因的方法。如果成功，这种新一代RNA-a方法将有可能治愈数十种难治性X连锁疾病，如Rett综合征，脆性X综合征和Duchenne肌营养不良症。在制药行业，目前的治疗策略几乎完全集中在蛋白质和microRNA靶点上。然而，lncRNA赋予蛋白质和小RNA不可能的时间和空间特异性，并且疾病相关突变更经常发生在非编码空间而不是编码空间。治疗策略可能有效地靶向长非编码RNA（lncRNA）。事实上，设计针对位点特异性顺式作用lncRNA的药物将避免多效性效应，即通过小分子的表观遗传复合物的一般活性。第一代RNA-a策略通过破坏Polycomb抑制复合物2（PRC 2）与将其靶向目标基因座的lncRNA之间的结合来重新唤醒表观遗传学沉默的基因。RNA-a技术实现了与RNAi相反的效果，并已在临床前测试中证明对各种疾病基因有效。然而，X连锁疾病基因不适合于RNA-a，可能是因为X染色体失活（XCI）导致的许多额外的异染色质层。XCI是哺乳动物中的剂量补偿机制，其中雌性细胞中的一条X染色体被沉默以平衡与雄性细胞的基因表达。我的实验室在过去的16年里专门从事XCI的研究，最近发现了与XCI沿着不活跃的X染色体（Xi）传播有关的X连锁元件。从这些发现中，我们假设应该可以选择性地重新唤醒特定的Xi基因座，而不重新激活大部分或全部的Xi。事实上，重新激活整个Xi是不可取的，因为我们已经证明，整体X-重新激活会导致癌症。因此，治疗X连锁疾病需要基因座特异性的基因激活方法。随着对XCI传播方式的新发现，设想中的下一代RNA-a技术有可能做到这一点。由于我们在XCI和RNA-a方面的专业知识，我的实验室处于独特的地位，可以进一步开发RNA-a技术，以基因座特异性方式解锁Xi基因的表达。在此，我们提出通过利用基因特异性控制元件沿着Xi重新激活模型疾病基因MECP 2来实现这一点。

DESCRIPTION (provided by applicant): The proposed research will develop methods of upregulating X-linked genes through technical innovations applied to the original "RNA-activation" technology developed in my laboratory (RNA-a; patent pending, PCT- US2011-065939). If successful, this next-generation RNA-a methodology will have the potential to cure dozens of intractable X-linked diseases such as Rett syndrome, Fragile X syndrome, and Duchenne muscular dystrophy. In the pharmaceutical industry, current therapeutic strategies focus almost exclusively on protein and microRNA targets. Yet, lncRNAs confer a temporal and spatial specificity not possible with proteins and small RNAs, and disease-associated mutations occur more often in noncoding than coding space. Therapeutic strategies might be as effectively targeted at long noncoding RNAs (lncRNA). Indeed, designing drugs against site-specific cis-acting lncRNAs would circumvent the pleiotropic effects plaguing modalities that general activities of epigenetic complexes via small molecules. The first-generation RNA-a strategy reawakens epigenetically silenced genes by disrupting binding between Polycomb repressive complex 2 (PRC2) and the lncRNA that targets it to the locus of interest. The RNA-a technology achieves the opposite of RNAi and has proven efficacious for various disease genes in preclinical testing. However, X-linked disease genes have not been amenable to RNA-a, likely because of many additional layers of heterochromatin resulting from X- chromosome inactivation (XCI). XCI is the mechanism of dosage compensation in mammals in which one X- chromosome is silenced in female cells to balance gene expression with male cells. My laboratory has specialized in the study of XCI for the past 16 years and recently discovered X-linked elements involved in the spreading of XCI along the inactive X chromosome (Xi). From these discoveries, we hypothesize that it should be possible to selectively reawaken specific Xi loci, without reactivating most or all of the Xi. Indeed, reactivating the entire Xi would be undesirable, as we have shown that global X-reactivation results in cancer. Thus, treating X-linked diseases requires a locus-specific approach to gene activation. With newfound knowledge of how XCI spreads, the envisioned next-generation RNA-a technology has the potential to do this. Because of our expertise in XCI and RNA-a, my lab is in a unique position to further develop the RNA-a technology to unlock expression of Xi genes in a locus-specific manner. Herein we propose to achieve this by leveraging gene-specific control elements along the Xi to reactivate the model disease gene, MECP2.