Macrophage Elastase in Host Defense

描述(申请人提供)：基质金属蛋白酶(MMPs)是一种 一组基质降解酶，其异常或过度表达可 导致各种组织破坏性疾病。人们对此知之甚少 MMPs的正常生理功能。我们提供的数据表明，巨噬细胞弹性蛋白酶 (基质金属蛋白酶-12)是唯一具有直接抗菌活性的基质金属蛋白酶。基质金属蛋白酶-12的作用 肺内巨噬细胞作为抵御微生物的第一道防线 肺泡腔。基质金属蛋白酶因其促进肿瘤的作用而广为人知 进步。然而，随着血管生成抑制素的发现， 纤溶酶原的蛋白水解性片段，很明显，蛋白水解酶可以 参与限制肿瘤生长。我们提出的证据表明，基质金属蛋白酶-12在 在限制肺内肿瘤生长方面发挥主要作用。此属性可能具有 至少6项使用基质金属蛋白酶抑制剂的3期临床试验的临床意义 去年，与此有关的癌症治疗和两种关节炎治疗被停止。 低估某些MMPs的特性以限制肿瘤生长。为了进一步 明确巨噬细胞和基质金属蛋白酶-12在宿主防御细菌和 我们建议：1.检验基质金属蛋白酶-12的假说 代表了一种新型的巨噬细胞介导的细胞内抗菌剂。我们 提供初步数据表明，作为响应，MMP12-/-小鼠的预后较差 金黄色葡萄球菌肺炎时，巨噬细胞对细胞内的损伤 对金黄色葡萄球菌的杀灭实验表明，基质金属蛋白酶-12具有直接抑菌作用。 这种活性与催化能力无关，并且涉及 非催化C末端结构域。建议进行研究以定义…的光谱 受基质金属蛋白酶-12影响的细菌。我们还将定义的结构组件 基质金属蛋白酶-12负责这一活动。2.我们将扩展假设 基质金属蛋白酶-12通过抑制血管生成干扰肿瘤生长 定义潜在的行动机制。我们提供了基质金属蛋白酶-12的初步数据 是维持Lewis肺细胞癌(LLC)转移休眠所必需的。 这种活性似乎与抑制血管生成有关。这不仅仅是 由于血管抑素的产生。额外的抗血管生成蛋白片段 发挥作用，我们推测，基质金属蛋白酶-12也干扰基质金属蛋白酶-2介导 促进肿瘤生长。基质金属蛋白酶-12可能通过切割基质金属蛋白酶-2以及 通过与基质金属蛋白酶-2竞争内皮细胞和肿瘤细胞结合 它的C-末端结构域。3.我们将确定巨噬细胞在肺中的作用 发展、细菌感染和肿瘤进展。我们会利用这个优势 基质金属蛋白酶-12巨噬细胞特异性表达与白喉的完全发生 毒素(DT)“敲入”到基质金属蛋白酶-12基因座。我们假设这将导致 在肺(和腹膜)巨噬细胞缺乏的小鼠中，这些小鼠 会经历正常的肺部发育。如果这一假设是正确的，那么 小鼠将被用来研究巨噬细胞在宿主防御和 发炎。如果突变是致命的或不是完全缺乏肺脏 巨噬细胞，然后肺特异性转基因小鼠将被用于诱导 在成年小鼠肺中表达DT。

DESCRIPTION (provided by applicant): Matrix metalloproteinases (MMPs) are a group of matrix degrading enzymes whose aberrant or excessive expression can lead to a variety of tissue destructive diseases. Less is known about the normal physiologic functions of MMPs. We present data that macrophage elastase (MMP-12) is the only MMP that has direct antimicrobial activity. MMP-12 acts within the lung macrophage as the first line of defense against microbes within the alveolar space. MMPs are well known for their roles in promoting tumor progression. However, with the discovery of angiostatin, an antiangiogenic proteolytic fragment of plasminogen, it became clear that proteinases can be involved in limiting tumor growth. We present evidence that MMP-12 plays a major role in limiting tumor growth within the lung. This property might have clinical importance since at least 6 phase 3 trials using MMP inhibitors for cancer therapy and two for arthritis were stopped last year related to this under-appreciated property of certain MMPs to limit tumor growth. To further define the role of macrophages and MMP-12 in host defense against bacteria and tumors in the lung, we propose to: 1. Test the hypothesis that MMP-12 represents a novel macrophage-mediated intracellular antimicrobial agent. We provide preliminary data that MMP-12-/- mice have a poorer outcome in response to S. aureus pneumonia, MMP-12-/- macrophages have impaired intracellular killing of S. aureus, and show that MMP-12 has direct antimicrobial capacity. This activity is independent of catalytic capacity and involves the non-catalytic C-terminal domain. Studies are proposed to define the spectrum of bacteria influenced by MMP-12. We will also define the structural components of MMP-12 responsible for this activity. 2. We will extend the hypothesis that MMP-12 interferes with tumor growth via inhibition of angiogenesis and further define potential mechanisms of action. We provide preliminary data that MMP-12 is required to maintain dormancy of Lewis lung cell carcinoma (LLC) metastases. This activity appears related to inhibition of angiogenesis. This is not merely due to generation of angiostatin. Additional antiangiogenic protein fragments play a role, and we postulate that MMP-12 also interferes with MMP-2-mediated promotion of tumor growth. MMP-12 might do this by cleavage of MMP-2 as well as by competition with MMP-2 for endothelial cell and tumor cell binding through its C-terminal domain. 3. We will determine the role of macrophages in lung development, bacterial infection, and tumor progression. We will take advantage of MMP-12 macrophage specific expression and complete generation of diphtheria toxin (DT) "knock-in" to the MMP-12 locus. We hypothesize that this will result in mice deficient in lung (and peritoneal) macrophages, and that these mice will undergo normal lung development. If this hypothesis is correct, then the mice will be used to study the requirement of macrophages in host defense and inflammation. If the mutation is lethal or not fully deficient in pulmonary macrophages, then lung-specific transgenic mice will be used to inducibly express DT in lungs of mature mice.