Aptamer chimeras for the in vivo modulation of beta cell mass and immunogenicity

1型（T1 D）糖尿病是由免疫介导的β细胞质量和功能丧失引起的。具有减小的 增殖和免疫介导的β细胞凋亡增加，胰岛素产生细胞变得不足以 充分调节身体的平衡。在T1 D诊断后不久，β细胞被完全 最近的研究表明，即使是长期患有癌症的患者， 如胰腺中胰岛素阳性胰岛的存在所证明的， 增加混合餐耐量试验后内源性胰岛素的产生。因此，T1糖尿病可能是 被视为一种慢性疾病，其中β细胞自身免疫破坏，再生， 和抗凋亡。临床上这种平衡明显倾向于β细胞的进行性丢失 然而，理论上可能通过减少损失来使β细胞稳态正常化， 恢复功能，促进残余β细胞再生。 在理解β细胞生物学和RNA治疗领域的最新进展提供了 这是一个前所未有的机会，可以在β细胞中选择性地靶向操纵那些控制细胞增殖的基因。 增殖、凋亡和对自身免疫的抗性。 我们认为，胰岛细胞特异性RNA适体可能是特异性递送胰岛素的工具。 治疗性RNA能够正向和负向调节内源性和移植的基因表达， β细胞。 RNA适体是一种小的、具有RNA酶抗性的、非免疫原性的寡核苷酸，可以深入细胞内， 并凭借其具有高亲和力的三维结构识别其靶标， 的特异性这些合成抗体是化学合成的，可以很容易地与小分子结合， 干扰或小的激活RNA（siRNA或saRNA），并且可以选择渗透到细胞质中 在目标结合时。 我们最近通过一种无监督的选择方法，生物信息学分析和实证研究， 验证，两种单克隆RNA适体能够在体内优先识别人β细胞。 长期的目标是有益地调节β细胞稳态和防御自身免疫， 在体内，我们决定将这些适体与能够上调PDL 1和Xip的小RNA缀合（以允许β细胞 分别对抗免疫攻击和抑制细胞凋亡）和下调p57 kip 2（以 促进β细胞增殖）。使用非解离的人胰岛的初步体外数据表明， 所得的治疗性RNA/胰岛特异性适体嵌合体有效地调节靶的表达 基因. 通过这个提议，我们将检验适体嵌合体可以介导体内基因调节的假设 在人类胰岛细胞中。这应该允许通过防止它们的损失，通过以下方式来改变β细胞功能和稳态： 增加它们的增殖，并提供自身免疫保护。这种创新的方法可以 允许开发基于体内靶向调节β的新型T1 D治疗策略 细胞基因

Type 1 (T1D) diabetes results from the immune mediated loss of β cell mass and function. With a reduced proliferation, and an increase immune mediated β cell apoptosis, insulin producing cells become insufficient to adequately regulate body glycemia. The historical dogma for which soon after T1D diagnosis β cell were fully and permanently lost has been challenged by recent studies revealing that even patients with long-standing diabetes residual β cells are retained as demonstrated by the presence of insulin positive islets in the pancreas and by increase production of endogenous insulin after mixed-meal tolerance test. T1 diabetes can thus been seen as a chronic disease in which the balance exist between β cells autoimmune destruction, regeneration, and resistance to apoptosis. Clinically this balance is obviously tilted toward the progressive loss of β cell mass, however, it might be theoretically possible to normalize β cell homeostasis by reducing loss, recovering function, and enhancing regeneration of remnant β cells. Recent advancements in the understanding β cell biology and in the field of RNA therapeutics offer an unprecedented opportunity for a targeted manipulation selectively in β cells of those genes that control cell proliferation, apoptosis and resistance to autoimmunity. We propose that RNA aptamers specific for islet cells might be the tool for the specific delivery of therapeutic RNA able to positively and negatively modulate gene expression in endogenous and transplanted β cells. RNA aptamers are small, RNAse resistant, non-immunogenic oligonucleotide that can penetrate deeply into the tissue and recognize their target in virtue of their three dimensional structure with high affinity and specificity. These synthetic antibodies are chemically synthetized, can be easily conjugated with small interference or small activating RNAs (siRNA or saRNA) and can be selected to penetrate into the cytoplasm upon target binding. We have recently identified by an unsupervised selection method, bioinformatics analysis and empirical validation, two monoclonal RNA aptamers able to recognized preferentially human β cells in vivo. With the long term goal to beneficially modulate β cell homeostasis and defense from auto-immunity in vivo, we decided to conjugate these aptamer with small RNA able to upregulate PDL1 and Xiap (to allow β cell to fight back the immunologic attack and inhibit apoptosis respectively) and to down-regulate p57kip2 (to promote β cell proliferation). Preliminary in vitro data using non-dissociated human islets demonstrated that the resulting therapeutic-RNA/islet-specific aptamer chimera effectively modulate the expression of the target genes. With this proposal we will test the hypothesis that aptamer-chimeras can be mediated in vivo gene modulation in human islet cells. This should allow to modify β cell function and homeostasis by preventing their loss, by increasing their proliferation, and by providing protection from autoimmunity. This innovative approach may allow the development of novel therapeutic strategies for T1D based on the in vivo targeted modulation of β cell genes.