Identification of AMD3100 (Plerixafor) as a potential lead compound for chlorine toxicity

项目总结/摘要 问题陈述：氯气（Cl 2）是美国最常见的吸入性刺激物， 导致严重的副作用，包括肺损伤和死亡。PI和其他人之前的研究表明， 表明白细胞（中性粒细胞和巨噬细胞）迁移和归巢到肺中起着关键作用。 Cl 2气体暴露后肺发病率和死亡率的作用。趋化因子配体，基质- 衍生因子-1（SDF-1）与C-X-C趋化因子受体4（CXCR 4）在肺免疫、上皮和 内皮细胞促进白细胞从循环到肺的迁移。SDF-1/CXCR 4轴还 传播肺中白细胞的活化和存活。我们的初步数据显示， 和CXCR 4水平在暴露于Cl 2的动物的肺中升高。因此，我们的假设是，SDF- 1/CXCR 4轴参与暴露于Cl 2后肺中白细胞的迁移、归巢和存活， 因此，通过FDA批准的化合物AMD 3100（Plerixafor）抑制该轴将减弱Cl 2-。 导致肺部发病率和死亡率。具体目的：1）确定SDF-1/CXCR 4轴在Cl 2诱导的细胞凋亡中的作用。 肺白细胞迁移、活化和存活。2)针对Cl 2毒性优化AMD 3100的给药方案。 3)阐明Cl 2暴露后SDF-1/CXCR 4调节的机制。实验方法： 将C57 BL/6小鼠暴露于Cl 2气体（500 ppm，30分钟），然后将C57 BL/6小鼠中的SDF-1浓度与C57 BL/6小鼠中的SDF-1浓度进行比较。 支气管肺泡灌洗液和肺泡白细胞及全肺CXCR 4表面表达 将随时间测量组织。还将分析SDF-1/CXCR 4轴是否介导Cl 2 肺白细胞的依赖性迁移、活化和存活。接下来，暴露于Cl 2的小鼠将被 肌内注射临床安全剂量（0.01-0.16 mg/kg）的AMD 3100， 将测量慢性肺损伤和死亡率。最后，研究了Cl 2诱导的溶血和缺氧在细胞凋亡中的作用。 将在体外和体内研究暴露于Cl 2后肺SDF-1和CXCR 4水平的调节。预期 结果：我们预期AMD 3100将减弱白细胞向肺部的移位， 肺损伤和改善暴露于Cl 2的小鼠的存活率。创新：该研究首次提出使用 一种新的，FDA批准的，小分子化合物，AMD 3100，以减轻肺损伤和死亡后， 暴露在氯气中。AMD 3100可能有助于减轻暴露于其他有毒气体后的肺损伤 例如溴和光气，它们也与溶血和增加的 白细胞进入肺。

PROJECT SUMMMARY/ABSTRACT Statement of problem: Chlorine (Cl2) gas is the most common inhalational irritant in the United States, which results in serious adverse effects including lung injury and death. Previous studies by the PI and others have demonstrated that migration and homing of leukocytes (neutrophils and macrophages) into lungs play a critical role in lung morbidity and mortality post Cl2 gas exposure. The binding of the chemokine ligand, stromal- derived-factor-1 (SDF-1), to the C-X-C chemokine receptor type 4 (CXCR4) on lung immune, epithelial, and endothelial cells promote the migration of leukocytes from the circulation to lungs. The SDF-1/CXCR4 axis also propagates the activation and survival of leukocytes in the lungs. Our preliminary data shows that both SDF-1 and CXCR4 levels are elevated in the lungs of Cl2 exposed animals. Therefore, our hypothesis is that SDF- 1/CXCR4 axis is involved in the migration, homing, and survival of leukocytes in lung post exposure to Cl2 and therefore inhibiting this axis by an FDA approved compound, AMD3100 (Plerixafor), would attenuate Cl2- induced lung morbidity and mortality. Specific aims: 1) Establish the role of SDF-1/CXCR4 axis in Cl2-induced lung leukocyte migration, activation, and survival. 2) Optimize the dosage regimen of AMD3100 for Cl2 toxicity. 3) Delineate the mechanisms of SDF-1/CXCR4 regulation post-Cl2 exposure. Experimental approach: C57BL/6 mice will be exposed to Cl2 gas (500ppm, 30min) and then the SDF-1 concentration in broncholaveolar lavage fluid and the CXCR4 surface expression on the alveolar leukocytes and whole lung tissue will be measured over time. It will also be analyzed whether, the SDF-1/CXCR4 axis mediates Cl2 dependent migration, activation, and survival of lung leukocytes. Next the Cl2 exposed mice will be administered a clinically safe dose (0.01-0.16 mg/kg) of AMD3100 intramuscularly and the indices of acute and chronic lung injury and mortality will be measured. Finally, the role of Cl2-induced hemolysis and hypoxia in the regulation of lung SDF-1 and CXCR4 levels post Cl2 exposure will be studied in vitro and in vivo. Anticipated results: We anticipate that AMD3100 will attenuate leukocyte translocation to lungs and subsequently mitigate lung injury and improve survival in mice exposed to Cl2. Innovation: The study is the first to propose the use of a novel, FDA-approved, small molecule compound, AMD3100, to mitigate lung injury and mortality post exposure to Cl2 gas. AMD3100 may be beneficial in mitigating lung injury post exposure to other toxic gases such as bromine and phosgene, which are also associated with hemolysis and increased migration of leukocytes into lung.