Development of novel anti-biofilm compounds for treating chronic wounds

描述(申请人提供)：对传统治疗干预措施无效的慢性伤口每年折磨数百万人，仅在美国，与治疗这些伤口相关的直接成本估计就达到100-250亿美元。根除已在慢性伤口上定居的病原菌因这些细菌形成生物膜的倾向而变得复杂。生物膜由一个被细胞外基质包围的细菌群落组成，它有效地抵抗抗生素的作用和宿主的免疫反应。处于生物被膜状态的细菌对抗生素的抗药性大约是抗生素的1000倍，目前还没有可靠的治疗策略来分散预形成的生物膜。这项SBIR项目的范围是评估一类新的分子，称为2-氨基咪唑(2-Ais)，用于治疗慢性伤口的生物膜感染。2-AI分子是第一类非杀菌小分子，已被证明可以分散革兰氏阳性和革兰氏阴性细菌的生物膜。鉴于2-AI分子的非杀微生物性，它们不会创造一个可能导致耐药性发展的选择性环境。在第一阶段合成了100多个2-AI分子，并使用Agile Sciences的高通量筛选和蒙大拿州立大学生物膜工程中心(CBE)的高级体外模型评估了它们的活性。通过这一努力，我们确定了一种名为H10的铅分子，它有效地分散了由滴流反应器形成的金黄色葡萄球菌和铜绿假单胞菌的强大生物膜。此外，在人类角质形成细胞划痕愈合模型中，当金黄色葡萄球菌生物膜的条件培养液用H10处理时，伤口完全闭合。这些结果为2-AI化合物治疗慢性伤口生物膜感染的潜力提供了强有力的体外证据，并推动了我们的第二阶段项目。在第二阶段，我们将：1)评估 H10在两个标准体内伤口模型(兔耳和猪)中的有效性：2)进行安全性评估，以便为后续的IND毒性研究提供信息。使用兔耳伤口模型和猪伤口模型对H10进行局部治疗的评估将由西北大学的Robert Galiano博士(兔模型)和迈阿密大学的Stephen Davis博士(猪模型)进行，CBE的Garth James博士将提供生物膜成像支持。由于H10没有杀灭微生物的作用，因此它将与抗生素联合使用，以协同去除生物膜，此外还能杀灭细菌。熟悉药物安全的合同研究机构将进行遗传毒性、皮肤刺激性、细胞色素P450抑制和急性毒性评估。该第二阶段项目的成功衡量标准是确定一种H10-抗菌剂组合，该组合可促进体内伤口愈合，并具有良好的毒性特征。这项工作中确定的联合疗法将是 第三阶段将在GLP条件下进行高级到IND-Enabling毒性研究。

DESCRIPTION (provided by applicant): Chronic wounds that fail to respond to traditional therapeutic interventions afflict millions of people each year, and direct costs associated with treating these wounds are estimated at $10-25 billion annually in the U.S. alone. Eradication of pathogenic bacteria that have colonized chronic wounds is complicated by the propensity of these bacteria to form biofilms. A biofilm consists of a community of bacteria encompassed by an extracellular matrix which efficiently resists the action of antibiotics and the host immune response. Bacteria in the biofilm state are approximately one-thousand times more resistant to antibiotics, and there are currently no reliable therapeutic strategies available for dispersing pr-formed biofilms. The scope of this SBIR project is to evaluate a new class of molecules, called the 2-aminoimidazoles (2-AIs), for treating biofilm-based infections in chronic wounds. The 2-AI molecules are the first class of non- microbicidal small molecules that have been shown to disperse biofilms of both Gram-positive and Gram- negative bacteria. Given the non-microbicidal nature of the 2-AI molecules, they do not create a selective environment that could lead to the development of resistance. Over one-hundred 2-AI molecules were synthesized in Phase I and evaluated for their activity using high-throughput screening at Agile Sciences and advanced in vitro models at the Center for Biofilm Engineering (CBE) at Montana State University. Through this effort, we identified a lead molecule, named H10, which effectively disperses robust biofilms of S. aureus and P. aeruginosa formed with a drip flow reactor. Furthermore, full closure of a wound in a human keratinocyte cell scratch closure model was achieved when conditioned media from S. aureus biofilms was treated with H10. These results provide strong in vitro evidence of the potential of the 2-AI compounds to treat biofilm-based infections in chronic wounds, and motivate our Phase II project. In Phase II, we will: 1) evaluate the effectiveness of H10 in two standard in vivo wound models (rabbit ear and porcine) and 2) conduct safety evaluations in order to inform subsequent IND-enabling toxicity studies. Evaluations of H10 as a topical therapeutic using the rabbit ear wound model and the pig wound model will be performed Dr. Robert Galiano of Northwestern (rabbit model) and Dr. Stephen Davis of the University of Miami (pig model) with Dr. Garth James of the CBE providing biofilm imaging support. Since H10 is non-microbicidal, it will be co-dosed with an antibiotic to provide synergistic removal of the biofilm in addition to killing of the bacteria. Contract Research Organizations that are well-versed in drug safety will perform genotoxicity, skin irritation, cytochrome P450 inhibition, and acute toxicity evaluations. The metric of success for this Phase II project is to identify an H10-antimicrobial combination that enhances wound healing in vivo and possesses a favorable toxicity profile. The combination therapy identified in this work will be advanced to IND-enabling toxicity studies to be conducted under GLP conditions in Phase III.