Molecular Heparan Sulfate Delivery in Guided Tissue Regeneration

描述(申请人提供)：蛋白多糖和碳水化合物聚合物，如硫酸乙酰肝素在生物过程中的作用正在得到更好的理解，但这一新的前沿才刚刚开始探索。组织再生也是一个新兴的前沿领域。这项拟议的研究完成后，将对蛋白多糖和组织再生领域做出重要贡献，并对牙周再生产生特定的临床影响。Perlecan最初被命名为硫酸乙酰肝素蛋白多糖-2，现在已知是所有基底膜(以及IV型胶原和层粘连蛋白)的重要组成部分。硫酸肝素修饰的Perlecan在血管生成或新生血管中的作用有强有力的科学支持和理论基础。据报道，Perlecan的硫酸乙酰肝素可以促进细胞黏附，促进增殖和/或分化，并结合和传递生长因子。自那以后，硫酸乙酰肝素修饰的重组Perlecan D1(Rd1)在各种细胞类型中的表达已经得到验证。体外实验表明，Rd1能与成纤维细胞生长因子结合，促进细胞黏附。在牙周再生中，屏障膜是旨在恢复口腔形态和功能的程序的重要附件。在这些引导组织再生(GTR)手术中，屏障膜被放置在手术瓣和下面的再生部位之间。然而，GTR手术的失败率不必要地高，原因是皮瓣未能愈合，膜和再生部位一期闭合，通常导致再生部位暴露在口腔环境中。这项第一阶段应用的目的是1)确定在临床可接受的参数范围内负载Perlecan DOMAIN 1表达载体的常用屏障膜的有效方式，2)比较由Perlecan D1转基因递送系统负载的膜产生的Perlecan DOMAIN 1的体外细胞表达和细胞反应，以及3)比较在体内通过放置加载的和未加载的外科手术膜而产生的反应，以便通过2意图愈合的动物模型。膜将通过在体外建立的参数进行预处理，然后固定在具有限定的膜暴露区域的牙龈瓣下。愈合将在临床和组织学上进行测量，其中裸露的上皮膜覆盖率将是主要结果。硫酸乙酰肝素和Perlecan D1提供支持组织再生的良好理论基础，再加上这些初步数据和Agenta的相关专利，使Agenta具有独特的地位，可以实现这一拟议项目的目标。

DESCRIPTION (provided by applicant): The role of proteoglycans and carbohydrate polymers such as heparan sulfate in biological processes are becoming better understood, but this new frontier has only begun to be explored. Tissue regeneration is also an emerging frontier. This proposed research, when completed, will make an important contribution to both the proteoglycan and tissue regenerative fields, with a specific, clinical impact on periodontal regeneration. Perlecan, originally named heparan sulfate proteoglycan-2, is now known to be an important component of all basement membranes (along with collagen type IV and laminin). There is strong scientific support and rationale for a role of heparan sulfate-decorated Perlecan in angiogenesis or neovascularization. The heparan sulfate of Perlecan is reported to promote cell adhesion, to promote proliferation and/or differentiation, and to bind and deliver growth factors. Expression of a heparan sulfate-decorated recombinant Perlecan D1 (rD1) in a variety of cell types has since been validated. The rD1 was shown to bind fibroblast growth factor and promoted cell adherence in vitro. In periodontal regeneration, barrier membranes are an important adjunct to procedures aimed at restoring the form and function of the mouth. During these guided tissue regenerative (GTR) procedures, barrier membranes are positioned between the surgical flap and the underlying regenerative site. Nevertheless, an unnecessarily high failure rate exists in GTR procedures due to failure of the flaps to heal with primary closure over the membranes and regenerative site, typically leading to exposure of the regenerative site to the oral environment. The objectives of this Phase I application are 1) to determine an effective manner of loading commonly used barrier membranes with Perlecan domain 1 expression constructs within clinically acceptable parameters, 2) to compare in vitro cellular expression of, and cellular responses to, the Perlecan domain 1 construct generated by membranes loaded with the Perlecan D1 transgene delivery system vs. the sham-loaded membranes, and 3) to compare in vivo responses generated by placement of the loaded vs. the sham-loaded surgical membranes in an animal model for healing by 2¿ intention. Membranes will be pre-treated by parameters established in vitro then secured beneath a gingival flap with a defined area of membrane exposure. Healing will be measured clinically and histologically where rate of exposed membrane coverage by epithelium will be the primary outcome. The well-supported rationale for heparan sulfate and Perlecan D1 delivery to support tissue regeneration, coupled with these preliminary data and Agenta's relevant patent, uniquely positions Agenta to fulfill the objectives of this proposed project.