Development of an acute myeloid leukemia murine model of invasive pulmonary aspergillosis to gain insights into the role of leukemia and its treatments in the pathobiology of aspergillosis

项目总结 侵袭性肺曲霉菌病(IPA)是血液系统恶性肿瘤患者常见且令人恐惧的并发症。 尤其是接受缓解诱导化疗(RIC)的急性髓系白血病(AML)患者。作为教宗- 在严重免疫功能障碍的背景下，传统抗真菌药物疗效有限，长期目标是发展 简便的免疫治疗策略，以改善机会性侵袭性霉菌感染的结果。关键之路-- 阻碍了这一领域的临床前研究，这导致了相当大的抗真菌注射的工作台/床边脱节。 免疫疗法，是依赖于药物免疫抑制但在其他方面健康的啮齿动物模型，不能 概述由活跃的血液系统恶性肿瘤和化疗引起的复杂的免疫变化。此外， 白血病背景下缺乏与病理生理学相关的IPA感染模型阻碍了更好的理解 肿瘤免疫疗法在抗霉菌免疫方面的潜在“非靶点益处”。因此，主要目标是 R03项目的目的是在急性髓细胞白血病背景下建立小鼠IPA感染模型，并研究作为原理的证明， 先前提出的“双重打击假说”认为，当ICIS作为白血病RIC的一部分给予时，可能伴随而来 增强宿主对霉菌的防御，从而改善IPA并发RICs的结果。在目标1a中，简单和 成本效益高的同基因C1498 AML小鼠模型将被改编和优化，用于RIC期间IPA的研究。 不同烟曲霉分生孢子感染白血病小鼠的发病率、死亡率和真菌负荷 在常规高强度化疗期间接种阿糖胞苷，或在低甲基化化疗期间接种 5-氮胞苷。在目标1b中，该模型将被验证为治疗平台，通过展示 伏立康唑，一线曲霉活性抗真菌药。在目标2中，新开发的模型将被用于 比较IPA的发病率/死亡率和对烟曲霉菌感染的关键免疫反应(例如， 血清和肺组织，适应性免疫极化，免疫耗竭标记物，以及肺白细胞募集- 在伴有或不伴有PD-1/PD-L1阻断的RIC去甲基化过程中，发生IPA的小鼠。支持- Proposed研究具有创新性，因为它引入了使用白血病模型来研究肿瘤- IPA免疫病理的逻辑治疗。一旦建立，这个临床前平台将提供无价的 未来研究抗真菌免疫疗法、新的抗真菌药物以及新的抗真菌药物的影响的资源 白血病药物在急性髓系白血病背景下的抗霉菌免疫。这个项目的意义源于它的三个主要方面 可交付成果：(I)急性髓细胞白血病小鼠的病理生理相关的IPA感染模型，该模型定位于设置新的 抗白血病治疗中机会性霉菌感染的临床前研究的“金标准”，(Ii)比较 PD-1途径抑制剂作为抗白血病化疗的一部分对IPA的保护作用的测定 在患有AML的小鼠中，以及(Iii)与IPA的良好结果相关的免疫特征的评估 该技术可用于治疗急性髓系白血病小鼠，并可为未来的人类生物标记物和免疫治疗研究提供信息。

PROJECT SUMAMRY Invasive pulmonary aspergillosis (IPA) is a common and feared complication in patients with hematological malignancies, especially patients with acute myeloid leukemia (AML) undergoing remission-induction chemotherapy (RIC). As conven- tional antifungals have limited efficacy in the background of severe immune dysfunction, the long-term goal is to develop facile immunotherapeutic strategies to improve the outcomes of opportunistic invasive mold infections. The critical road- block for preclinical studies in this area, which contributes to the considerable bench/bedside disconnect of antifungal im- munotherapy, is the reliance on pharmacologically immunosuppressed but otherwise healthy rodent models that cannot recapitulate the complex immune alterations caused by an active hematological malignancy and chemotherapy. Moreover, the lack of pathophysiologically relevant IPA infection models in a leukemia background prevents a better understanding of potential “off-target benefits” of oncological immunotherapies on anti-mold immunity. Therefore, the primary objectives of this R03 project are to develop a murine IPA infection model in an AML background and to study, as a proof of principle, the previously proposed “double-hit hypothesis” that ICIs, when given as part of RIC in leukemia, might concomitantly enhance host defense against molds and thereby improve the outcomes of IPA complicating RIC. In Aim 1a, the simple and cost-efficient, syngeneic C1498 AML mouse model will be adapted and optimized for studies of IPA during RIC by com- paring morbidity, mortality, and fungal burden of leukemic mice infected with different Aspergillus fumigatus conidial inoculums during either conventional high-intensity chemotherapy with cytarabine or hypomethylating chemotherapy with 5-azacytidine. In Aim 1b, the model will be validated as a therapeutic platform by demonstrating protective activity of voriconazole, the first-line Aspergillus-active antifungal agent. In Aim 2, the newly developed model will be leveraged to compare morbidity/mortality of IPA and key immunological responses to A. fumigatus infection (e.g., cytokine profiles in serum and lung tissue, polarization of adaptive immunity, immune exhaustion markers, and pulmonary leukocyte recruit- ment) in mice developing IPA during hypomethylating RIC with or without concomitant PD-1/PD-L1 blockade. The pro- posed research is innovative as it introduces the conceptual novelty of using a leukemia model to study the impact of onco- logical therapies on the immunopathology of IPA. Once established, this preclinical platform will provide an invaluable resource for future studies of antifungal immunotherapy, new antifungal agents, and the impact of new classes of anti- leukemia agents on anti-mold immunity in an AML background. The significance of this project stems from its three major deliverables, (i) a pathophysiologically relevant IPA infection model in mice with AML that is positioned to set the new “gold standard” for preclinical studies of opportunistic mold infections during anti-leukemia therapy, (ii) a comparative determination of protective effects of PD-1 pathway inhibitors, given as part of anti-leukemia chemotherapy, against IPA in mice with underlying AML, and (iii) an assessment of immune features that correlate with favorable outcomes of IPA in mice with AML and could inform future human biomarker and immunotherapy studies.