Novel Therapy for Chronic Non-healing Wounds

摘要：随着糖尿病和代谢综合征的日益流行，目前估计美国约有2000万人（约占人口的7%）受到糖尿病和代谢综合征相关并发症的影响，迫切需要解决这些并发症。伤口修复受损是血糖控制不良的主要并发症，每年造成的损失高达100亿美元。本实验室首先根据兔（Larrick et al., 1991）和人类阳离子抗微生物蛋白（hCAP18）的脂多糖结合和抑菌活性鉴定并克隆。在过去的10年里，hCAP18已被证明是先天免疫系统的一个重要组成部分，其c端片段LL-37具有广泛的抗微生物活性。hCAP18在白细胞中组成性产生，并在炎症和感染时在屏障器官（如肺、胃肠道、皮肤）中被诱导。最近的研究表明，hCAP18在皮肤伤口的血管化和再上皮化中起着关键作用。体内皮肤在受伤后产生高水平的hCAP18，在受伤后48小时达到峰值，在伤口愈合后下降到损伤前水平。hCAP18在炎症浸润和伤口床上迁移的上皮中被检测到。例如，Heilbron等人（2003）使用由器官培养的人皮肤组成的非炎性离体伤口愈合模型表明，使用抗ll -37抗体治疗可抑制再上皮化。在慢性未愈合伤口中，hCAP18水平较低，溃疡边缘上皮中缺乏hCAP18/LL-37的免疫反应性。我们假设LL-37在伤口愈合中起着关键作用，慢性伤口缺乏LL-37会损害再上皮化，促进细菌定植，并导致伤口愈合失败。因此，本提案的总体目标是开发一种蛋白酶抵抗形式的LL-37作为慢性非愈合伤口的新疗法。为此，在第一阶段，我们将制备prLL-37，一种由d -氨基酸组成的蛋白酶抗性形式，证明蛋白酶降解的抗性，并在糖尿病小鼠伤口愈合模型中评估prLL-37的活性。在II期，我们将开展临床前研究，以支持IND的提交。该疗法将解决烧伤和慢性不愈合、糖尿病足、卧疮和静脉淤滞性溃疡患者的主要未满足需求。摘要：随着糖尿病和代谢综合征的日益流行，目前美国约有1.2亿人（约占总人口的7%）受到糖尿病和代谢综合征相关并发症的影响。伤口修复受损是血糖控制不良的主要并发症，每年造成的损失高达100亿美元。本实验室首先根据兔（Larrick et al., 1991）和人类阳离子抗微生物蛋白（hCAP18）的脂多糖结合和抑菌活性鉴定并克隆。在过去的10年里，hCAP18已被证明是先天免疫系统的一个重要组成部分，其c端片段LL-37具有广泛的抗微生物活性。hCAP18在白细胞中组成性产生，并在炎症和感染时在屏障器官（如肺、胃肠道、皮肤）中被诱导。最近的研究表明，hCAP18在皮肤伤口的血管化和再上皮化中起着关键作用。体内皮肤在受伤后产生高水平的hCAP18，在受伤后48小时达到峰值，在伤口愈合后下降到损伤前水平。hCAP18在炎症浸润和伤口床上迁移的上皮中被检测到。例如，Heilbron等人（2003）使用由器官培养的人皮肤组成的非炎性离体伤口愈合模型表明，使用抗ll -37抗体治疗可抑制再上皮化。在慢性未愈合伤口中，hCAP18水平较低，溃疡边缘上皮中缺乏hCAP18/LL-37的免疫反应性。我们假设LL-37在伤口愈合中起着关键作用，慢性伤口缺乏LL-37会损害再上皮化，促进细菌定植，并导致伤口愈合失败。因此，本提案的总体目标是开发一种蛋白酶抵抗形式的LL-37作为慢性非愈合伤口的新疗法。为此，在第一阶段，我们将制备prLL-37，一种由d -氨基酸组成的蛋白酶抗性形式，证明蛋白酶降解的抗性，并在糖尿病小鼠伤口愈合模型中评估prLL-37的活性。在II期，我们将开展临床前研究，以支持IND的提交。该疗法将解决烧伤和慢性不愈合、糖尿病足、卧疮和静脉淤滞性溃疡患者的主要未满足需求。

DESCRIPTION (provided by applicant): ABSTRACT: A significant need exists to address complications associated with the growing pandemic of diabetes mellitus and metabolic syndrome now estimated to affect >20 million people in the United States (~7% of the population!). Impaired wound repair, a major complication of poor glycemic control, costs >$10 billion annually. Our laboratory first identified and cloned rabbit (Larrick et al., 1991) and then human Cationic Anti- microbial Protein (hCAP18) based on its LPS binding and anti-microbial activities. Over the past 10 years hCAP18 has been shown to be an important component of the innate immune system with broad anti-microbial activity conferred by its C-terminal fragment LL-37. hCAP18 is constitutively produced in leukocytes and is induced in barrier organs (e.g. lung, GI, skin) upon inflammation and infection. Recent work demonstrated a key role of hCAP18 in vascularization and re-epithelialization of skin wounds. High levels of hCAP18 are produced in skin in vivo upon wounding with a peak at 48 h post-injury, declining to pre-injury levels upon wound closure. hCAP18 is detected in the inflammatory infiltrate and in the epithelium migrating over the wound bed. For example, using a noninflammatory ex vivo wound healing model, composed of organ-cultured human skin, Heilbron et al. (2003) showed that treatment with anti-LL-37 antibodies inhibits re-epithelialization. In chronic non-healing wounds, hCAP18 levels are low and immunoreactivity for hCAP18/LL-37 is absent in ulcer edge epithelium. We hypothesize that LL-37 plays a critical role in wound closure and that its absence in chronic wounds impairs re-epithelialization, promotes bacterial colonization and contributes to failure of the wounds to heal. Hence, the overall goal of this proposal is to develop a protease resistance form of LL-37 as a novel therapy for chronic, non-healing wounds. To this end in phase I we will prepare prLL-37, a protease resistant form comprised of D-amino acids, demonstrate resistance to degradation by proteases, and evaluate activity of prLL-37 in a wound healing model using diabetic mice. In phase II we will carry out preclinical studies to support submission of an IND. This therapy will address a major unmet need among patients suffering from burn injuries and chronic non-healing, diabetic foot, decubitus and venous stasis ulcers. ABSTRACT: A significant need exists to address complications associated with the growing pandemic of diabetes mellitus and metabolic syndrome now estimated to affect >20 million people in the United States (~7% of the population!). Impaired wound repair, a major complication of poor glycemic control, costs >$10 billion annually. Our laboratory first identified and cloned rabbit (Larrick et al., 1991) and then human Cationic Anti- microbial Protein (hCAP18) based on its LPS binding and anti-microbial activities. Over the past 10 years hCAP18 has been shown to be an important component of the innate immune system with broad anti-microbial activity conferred by its C-terminal fragment LL-37. hCAP18 is constitutively produced in leukocytes and is induced in barrier organs (e.g. lung, GI, skin) upon inflammation and infection. Recent work demonstrated a key role of hCAP18 in vascularization and re-epithelialization of skin wounds. High levels of hCAP18 are produced in skin in vivo upon wounding with a peak at 48 h post-injury, declining to pre-injury levels upon wound closure. hCAP18 is detected in the inflammatory infiltrate and in the epithelium migrating over the wound bed. For example, using a noninflammatory ex vivo wound healing model, composed of organ-cultured human skin, Heilbron et al. (2003) showed that treatment with anti-LL-37 antibodies inhibits re-epithelialization. In chronic non-healing wounds, hCAP18 levels are low and immunoreactivity for hCAP18/LL-37 is absent in ulcer edge epithelium. We hypothesize that LL-37 plays a critical role in wound closure and that its absence in chronic wounds impairs re-epithelialization, promotes bacterial colonization and contributes to failure of the wounds to heal. Hence, the overall goal of this proposal is to develop a protease resistance form of LL-37 as a novel therapy for chronic, non-healing wounds. To this end in phase I we will prepare prLL-37, a protease resistant form comprised of D-amino acids, demonstrate resistance to degradation by proteases, and evaluate activity of prLL-37 in a wound healing model using diabetic mice. In phase II we will carry out preclinical studies to support submission of an IND. This therapy will address a major unmet need among patients suffering from burn injuries and chronic non-healing, diabetic foot, decubitus and venous stasis ulcers.