To Treat Some Ocular Diseases with Id1/Id3 Inhibitors

ID1和ID3基因和蛋白是血管生成过程中的关键成分，参与了细胞分化和增殖的广泛过程。目前已知，ID蛋白有助于调节骨髓来源的和循环中的内皮前体细胞的释放和募集，这些细胞有助于形成肿瘤的新血管。最近，同样的观察已经扩展到眼睛中的新生血管。为此，我们认为，无论是阻止id1/id3基因的表达，还是抑制id1和id3蛋白在眼内的功能，都可能是预防或治疗的有效方法 眼部新生血管。在老年性黄斑变性(ARMD)、糖尿病视网膜病变(DR)和早产儿视网膜病变(ROP)中，异常的血管形成会导致严重的视力丧失，因此抑制病理性眼血管生成至关重要。我们和另一个团队已经成功地使用了ID1的反义分子来阻止与眼睛无关的动物模型中的血管生成。如果玻璃体内注射到眼睛上，这样的制剂可能会很有用。除了它的治疗潜力，反义分子还将被用作人类眼病动物模型的概念验证实验的重要工具。因此，拟议研究的总体目标是确认对ID1/ID3的抑制 基因表达是治疗以病理为特征的眼部疾病的有效方法 新生血管。为了实现这些目标，我们提出了以下三个特定的目标：(1)通过量化id1和id3缺乏的小鼠缺血诱导的视网膜新生血管模型中的视网膜新生血管，以及使用玻璃体内传递的具有阻断id1表达和血管生成的已知活性的反义dna寡核苷酸来复制id1/id3基因敲除小鼠的抗血管生成表型，来研究id1/id3基因敲除小鼠的视网膜血管生成中id1/id3的作用；(2)使用体外/体内Matrigel研究和小鼠高氧模型来测试siRNA寡核苷酸和另一个反义分子，以及(3)优化一个或多个基于DNA的或siRNA作用的寡核苷酸，通过评估各种载体来改善药物的持续时间和效果，并建立至少一种活性寡核苷酸的剂量-反应关系，从而显示出有希望的活性。 这些目的的实现将证明使用特定的寡核苷酸通过阻断Id1的表达来抑制眼血管生成的可行性。第二阶段的活动将扩大该方法，包括阻断ID3的表达，探索同时阻断ID1和ID3表达的可能性，并从活性、药代动力学和生物利用度方面进一步优化寡核苷酸和可能的寡核苷酸组合。此外，候选寡核苷酸将在一个或多个额外的人类眼病动物模型中进行评估，例如小鼠的Ar激光光凝模型。

The Id1 and Id3 genes and proteins are critical components of the angiogenic process and participate in a wide range of processes involved in cellular differentiation and proliferation. It is now known that Id proteins help regulate the release and recruitment of bone marrow derived and circulating endothelial precursor cells which help form the new blood vessels of tumors. Recently this same observation has been extended to neovascularization in the eye. For this reason, we propose that either preventing expression of Id1/Id3 genes or inhibiting the functions of the Id1 and Id3 proteins in the eye could be an effective way to prevent or treat ocular neovascularization. Inhibiting pathologic ocular angiogenesis is of critical importance because aberrant blood vessel formation leads to severe visual loss in age-related macular degeneration (ARMD), diabetic retinopathy (DR) and retinopathy of prematurity (ROP). We, and one other group, have successfully used an antisense molecule to Id1 to block angiogenesis in an animal model unrelated to the eye. Such an agent might be useful if administered intravitreally to the eye. In addition to its therapeutic potential, an antisense molecule would also find significant use as a tool for proof-of-concept experiments in animal models of human ocular disease. Thus, the over-all goal of the proposed research is to confirm that inhibition of Id1/Id3 gene expression is a useful approach for treating ocular diseases characterized by pathologic neovascularization. To accomplish these goals, we propose the following three specific aims: (1) Examine the role of Id1/Id3 in retinal angiogenesis by quantifying retinal neovascularization in a mouse model of ischemia-induced retinal neovascularization in mice deficient in Id1 and Id3 and by using an intravitreally delivered anti-DNA antisense oligonucleotide with known activity for blocking Idl expression and angiogenesis to duplicate the anti-angiogenic phenotype of the Id1/Id3 knockout mouse, (2) Use in vitro/in vivo Matrigel studies and the mouse hyperoxia model to test the activity of siRNA oligonucleotides and one additional antisense molecule, and (3) Optimize one or more DNA-based or siRNA acting oligonucleotides that demonstrate promising activity by evaluating various vehicles to improve the duration and effect of the agent and establishing a dose-response relationship for at least one of the active oligonucleotides. Accomplishing these aims will demonstrate the feasibility of using specific oligonucleotides to inhibit ocular angiogenesis by blocking Id1 expression. Phase II activities will expand the approach to include blocking Id3 expression, exploring the possibility of blocking both Id1 and Id3 expression and further optimization of the oligonucleotides and possibly oligonucleotide combinations in terms of activity, pharmacokinetics and bioavailability. In addition, candidate oligonucleotides will be evaluated in one or more additional animal models of human ocular disease such as the argon laser photocoagulation model in mice.