Anti-angiogenesis of HC-HA Covalent Complex and PTX3 Purified from Fetal Membrane

描述(申请人提供)：血管生成是成人伤口愈合的一部分，它始于炎症，可能最终形成疤痕。当炎症、血管生成和疤痕形成得到很好的控制时，成人伤口愈合会导致组织功能的恢复。相反，经常与失控炎症相关的病理性血管生成可能会导致病理性瘢痕。在视力是关键功能的眼睛中，任何这种容易留下疤痕的伤口愈合都可能导致失明。事实上，美国国立卫生研究院已经认识到，慢性伤口无法愈合是美国的一个主要健康问题，而且随着人口老龄化，这种问题的规模将会增加。我们一直认为，解决上述问题的一种新方法是更好地了解由羊膜(AM)和绒毛膜(CH)组成的胎儿膜的生物学功能。我们的推测部分源于AM移植在向眼表提供抗炎、抗瘢痕和抗血管生成效果方面的累积临床成功，以及胎儿伤口愈合的神秘，它缺乏炎症和血管生成，并且是无疤痕的。我们成功地从AM中提纯了透明质酸(HA)与间质抑制物(I I)重链(HC)形成的共价HC“HA复合体，并鉴定其为一种具有抗炎、抗瘢痕和抗血管生成作用的活性成分。我们进一步收集了强有力的证据支持从CH中提纯的HC“HA的效力是从AM中提纯的HA的25倍，并且从CH中提纯的PTX3可能增强HC”HA对血管内皮细胞增殖的抑制作用。因此，在这一阶段的应用中，我们建议通过验证PTX3确实是来自CH的关键蛋白来描述HC“HA的化学结构，以增强HC”HA的抗血管生成作用(目标1)；并表征和比较从CH提纯的HC“HA和从AM提纯的HC”HA的体外抗血管生成作用(目标2)。这项研究将由第二阶段(R44 EY017497)拨款支持，重点是测试从AM中提纯的HC“HA在体内的抗炎和抗瘢痕效果，我们相信，上述两个目标的成功完成将使我们首次提出一种新的治疗模式，其中单个HC”HA复合体可以同时抑制成人伤口愈合的三个关键步骤，即炎症、血管生成和瘢痕形成。因此，我们设想HC“HA不仅可能是一种胚胎基质，负责确保胎儿伤口无疤痕的愈合，而且还可以充分利用胎儿膜作为一种新的生物材料，从中我们可能推出新的治疗方法，使成人伤口愈合向再生方向发展。通过将PTX3引入HC”HA，我们可能开发一类新的血管生成抑制剂，用于治疗眼科内外的许多疾病，这些疾病的病理性血管生成威胁我们的视力和健康。 公共卫生相关性：本申请第一阶段介绍的研究旨在开发一种新型的血管生成抑制剂，它基于透明质酸(HA)和中间抑制剂重链(HC)之间形成的共价HC“HA复合体，并从胎儿膜，即羊膜和绒毛膜中提纯。我们的研究旨在证明PTX3，一个已知具有抗血管生成和抗炎作用的五星蛋白家族成员，第一，与从绒毛膜中提纯的HC“HA密切相关，第二，负责增强HC”HA的抗血管生成能力。我们认为，PTX3可以被添加到HC“HA中，产生一类新的血管生成抑制剂，不仅可以抑制bFGF和VEGF介导的血管生成，而且还可以通过减少额外的炎症和瘢痕形成而更有效地发挥作用。因此，我们可能会开发基于与HC“HA结合的PTX3的新疗法，以治疗眼科内外的许多疾病，在这些疾病中，不必要的血管生成损害我们的视力和健康。

DESCRIPTION (provided by applicant): Angiogenesis is a part of adult wound healing, which starts from inflammation and may end in scarring. When inflammation, angiogenesis, and scarring are well controlled, adult wound healing results in the restoration of tissue function. In contrast, pathologic angiogenesis that is frequently associated with uncontrolled inflammation may lead to pathologic scarring. In the eye, where vision is the key function, any such scar-prone wound healing can lead to blindness. In fact, the NIH has recognized that the inability of chronic wounds to heal is a major health problem in the United States and that such a problem will increase in magnitude as the population ages. We have long speculated that one novel solution to the aforementioned problem lies in a better understanding of the biological function of the fetal membrane, which consists of the amniotic membrane (AM) and the chorion (CH). Our speculation is derived partly from cumulative clinical successes of AM transplantation in delivering anti-inflammatory, anti- scarring, and anti-angiogenic efficacies to the ocular surface, as well as from the mystery of fetal wound healing that lacks inflammation and angiogenesis, and is "scarless". We have successfully purified the covalent HC"HA complex formed between hyaluronan (HA) and the heavy chain (HC) of inter- -inhibitor (I I) from the AM and has identified it as one active component that exerts anti-inflammatory, anti-scarring and anti-angiogenic actions. We have further gathered strong evidence supporting that HC"HA purified from the CH is 25 fold more potent than that purified from the AM and that PTX3 from the CH may enhance the HC"HA's suppression of vascular endothelial proliferation. In this Phase I application, we thus propose to delineate the chemical structure of HC"HA by verifying that PTX3 is indeed the key protein from CH to enhance HC"HA's anti-angiogenic action (Aim 1); and to characterize and compare the in vitro anti-angiogenic action of HC"HA purified from the CH to that from AM (Aim 2). Together with the study to be supported by a Phase II (R44 EY017497) grant, which focuses on testing in vivo anti-inflammatory and anti-scarring efficacies by HC"HA purified from the AM, we believe that successful completion of the above two aims will let us put forth a novel therapeutic paradigm, for the first time, where a single HC"HA complex can simultaneously curtail three key steps, i.e., inflammation, angiogenesis, and scarring of adult wound healing. As a result, we envision that HC"HA may not only be an embryonic matrix responsible for ensuring scarless fetal wound healing, but it can also be sufficiently harnessed from the fetal membrane as a novel biomaterial, from which we may launch new therapeutics to gear adult wound healing towards regeneration. By including PTX3 to HC"HA, we may develop a new class of angiogenesis inhibitors for treating many diseases in and beyond ophthalmology where pathologic angiogenesis threatens our vision and health. PUBLIC HEALTH RELEVANCE: The research described in this Phase I application is aimed at developing a novel class of angiogenesis inhibitors that are based on the covalent HC"HA complex formed between hyaluronan (HA) and the heavy chain (HC) of inter- -inhibitor and purified from the fetal membrane, i.e., the amniotic membrane and the chorion. Our proposed research intends to prove that PTX3, a pentraxin family member known to exert both anti-angiogenic and anti- inflammatory actions, first, is strongly associated with HC"HA purified from the chorion and, second, is responsible for enhancing the anti-angiogenic potency of HC"HA. We believe that PTX3 can be added to HC"HA to produce a new class of angiogenesis inhibitors, which will not only suppress angiogenesis mediated by bFGF and VEGF but will also be more useful and effective by curtailing additional inflammation and scarring. Consequently, we may develop new therapeutics based on PTX3 bound to HC"HA to treat many diseases in and beyond ophthalmology where unwanted angiogenesis is detrimental to our vision and health.