Amelioration of Vesicant-Induced Skin Injury by High Dose 25-Hydroxyvitamin D

描述（由申请人提供）：直接皮肤接触氮芥或氮芥相关亚硝基脲（BCNU（卡莫司汀））等起泡剂会产生剂量依赖性症状，这些症状的发作、严重程度和伤口愈合持续时间各不相同。虽然暴露于起泡剂本身会引起原发性损伤（第一次打击），但对皮肤的直接损伤会导致活化的高炎症单核细胞/巨噬细胞（M）延迟流入，从而构成对皮肤的额外损伤（第二次打击），从而促进灾难性的皮肤损伤。然而，第一次和第二次组织击中之间的间隔代表了对策机会。据报道，维生素 D 可以减弱 M 的激活，并且在临床轶事中证实了这种效果，我们最近治疗了一位患者，该患者在使用口服维生素 D 进行局部 BCNU 治疗后表现出持续的毒性皮肤反应。这一成功使我们推测，用 25(OH) D3 减弱过度激活的 M 将是一种有效的对策，可以阻止皮肤破坏的进展，而皮肤破坏是暴露于出疱剂后的一个关键发病因素。这里介绍的多学科转化方法利用了凯斯西储大学和大学医院凯斯医疗中心皮肤科和医学系强大的基础设施。人类临床试验样本以及小鼠体内和体外样本将用于评估维生素 D 作为对抗亚硝脲和氮芥暴露后 M 激活引起的皮肤损伤的对策的潜力。我们组织了一个国家专家团队来评估我们的进展并帮助确定拟议工作的补充方法。使用从正在进行的烷化剂试验中获得的人类临床样本以及用氮芥治疗后获得的小鼠皮肤样本，我们建议评估：1.) 在 BCNU 暴露之前和之后的组织中，通过诱导可溶性信号分子 TNFa 和 iNOS 来评估 M 渗透和激活的水平。我们将进一步 2.) 确定补充维生素 D 是否会调节组织破坏，以及这是否是由 M¿ 激活的变化介导的。使用巨噬细胞限制性 VDR 敲除小鼠系统将确认 M 介导的反应的特异性。最后，我们将 3.) 开始一项维生素 D 介入试验，利用暴露于局部 BCNU 和氮芥的患者来评估患者炎症生物标志物的变化、细胞浸润和过度激活的 M 的免疫组织化学染色，同时监测血清维生素 D 水平从基线到治疗后的变化。将在补充前和补充后收集的血清进行体外自体巨噬细胞活化直接抑制的测定。由于维生素 D 是安全且广泛可用的，因此成功实施这一策略可能会在大规模暴露的情况下挽救生命。证明维生素 D 相关化合物可以防止出疱剂介导的组织破坏，这将导致对这种潜在治疗干预措施的进一步评估。 公共健康相关性：用作武器的氮芥或氮芥相关亚硝脲 (BCNU) 等化学发泡剂可能对人类健康产生毁灭性影响。芥末是强效烷化剂，上皮细胞（例如角质形成细胞）接触这些试剂会导致氧化和亚硝化应激的诱导，最终导致细胞死亡。在皮肤中，这种细胞死亡表现为严重的皮肤起泡和脱屑。可以阻止这些化学物质作用的新疗法对于治疗这些皮肤损伤将具有无价的价值。该提案使用小鼠和人体组织来解决维生素 D 是否可以用作对抗剂，以对抗因暴露于发泡剂而引发的过度活化巨噬细胞的激活和随后的组织破坏。

DESCRIPTION (provided by applicant): Direct cutaneous exposure to vesicants such as nitrogen mustard or nitrogen mustard-related nitrosoureas (BCNU (carmustine)) produce dose-dependent symptoms that are varied in onset, severity and duration of wound healing. Although exposure to the vesicant itself evokes a primary injury (1st hit), the direct insult to the skin results in a delayed influx of activated hyper-inflammatory monocyte/macrophages (M) that constitute an additional insult to the skin (2nd hit) that promotes catastrophic skin injury. However, the interval between the first and second tissue hits represents a countermeasure opportunity. Attenuation of M activation by vitamin D has been reported, and in a clinical anecdotal confirmation of this effect, we have recently treated a patient who exhibited a persistent toxic skin response following topical BCNU treatment using oral vitamin D. This success leads us to hypothesize that attenuating hyper-activated M with 25(OH) D3 will be an effective countermeasure that halts progression of skin destruction, a key morbidity factor following exposure to vesicants. The multidisciplinary translational approach presented here that takes advantage of a strong infrastructure in the Departments of Dermatology and Medicine at Case Western Reserve University and University Hospitals Case Medical Center. Human clinical trial samples as well as murine in vivo and in vitro samples will be used assess the potential for vitamin D to act as a countermeasure to skin damage elicited by M activation following nitrosurea and nitrogen mustard exposure. We have organized a team of national experts to evaluate our progress and help identify complementary approaches to the proposed work. Using human clinical samples obtained from ongoing trials of alkylating agents as well as murine skin samples obtained post-treatment with mechlorethamine we propose to assess: 1.) The level of M infiltration and activation using induction of the soluble signaling molecules TNFa and iNOS in pre- and post-BCNU exposed tissue. We will further 2.) Determine whether vitamin D supplementation modulates tissue destruction and if this is mediated by changes in M¿ activation. Use of a macrophage-restricted VDR knockout mouse system will confirm the specificity of M mediated response. Finally, we will 3.) Begin a vitamin D interventional trial utilizing patients exposed to topical BCNU and nitrogen mustard to assess alteration of inflammatory biomarkers, cellular infiltration and immunohistochemical staining of hyper-activated M in patients while monitoring changes in serum vitamin D levels from baseline to post-therapy. Serum collected pre- and post- supplementation will be assayed for direct suppression of autologous macrophage activation in vitro. Successful implementation of this strategy is likely to save lives in the event of mass exposure as vitamin D is safe and widely available. Demonstration that vitamin D-related compounds protect against tissue destruction mediated by vesicants will lead to advanced evaluation of this potential therapeutic intervention. PUBLIC HEALTH RELEVANCE: Chemicals vesicants such as nitrogen mustard or nitrogen mustard-related nitrosureas (BCNU) used as weapons can have a devastating impact on human health. The mustards are powerful alkylating agents and exposure to these agents by epithelial cells, such as keratinocytes, result in induction of oxidative and nitrosative stress an eventual cell death. In the skin, this cell death manifests as severe skin blistering and desquamation. New therapeutics which can halt the effects of these chemicals will be invaluable to treating these skin injuries. Using both mouse and human tissue this proposal addresses whether vitamin D can be used as a counteragent to the activation and subsequent tissue destruction caused hyper-activated macrophages initiated through vesicant exposure.