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Endothelial-Leukocyte Adhesion in CAR T Cell Treatment Associated Neurotoxicity

CAR T 细胞治疗相关神经毒性中的内皮-白细胞粘附

基本信息

批准号：
10735681
负责人：
Juliane Gust
金额：
$ 60.25万
依托单位：
SEATTLE CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-22 至 2028-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10735681
关键词：
3-Dimensional Ablation Acute Address Adhesions Affect Aftercare Animal Model Antibodies Basic Science Behavior Behavioral Biology Biomedical Engineering Blood Vessels Blood capillaries Brain Brain Death Brain Edema CAR T cell therapy CD19 gene Capillary Endothelial Cell Cell Adhesion Molecules Cell-Cell Adhesion Cells Cellular immunotherapy Central Nervous System Cerebral Edema Child Childhood Clinical Clinical Trials Collaborations Collagen Coma Delirium Development Dose Endothelium Engineering Epitopes Equilibrium Flow Cytometry Gene Expression Profiling Genetic Hematologic Neoplasms Hemorrhage Human Human Engineering Hydrogels Hypoxia Image Immune system Immunocompetent Immunoprecipitation Impaired cognition Impairment In Vitro Inflammatory Infusion procedures Injury Integrin Binding Integrins Interdisciplinary Study International Investigation Leukocytes Link Malignant - descriptor Malignant Neoplasms Measures Mediating Medical Modality Modeling Molecular Motor Mus Neurologic Neuronal Dysfunction Neurosciences Oncology Pathway Analysis Patients Perfusion Pericytes Phenotype Plasma Positioning Attribute Property Proteins Protocols documentation Receptor Activation Risk Risk Factors Safety Sampling Seizures Signal Induction Signal Transduction Small Interfering RNA Stimulus T-Cell Proliferation T-Lymphocyte Techniques Testing Therapeutic Intervention Toxic effect Transgenic Organisms Translating Work brain dysfunction brain endothelial cell cancer cell cancer immunotherapy cell type cerebral capillary cerebral microvasculature chimeric antigen receptor chimeric antigen receptor T cells cohort cytokine cytokine release syndrome experience experimental study genetically modified cells high risk human model immunological synapse in vitro Model in vivo in vivo imaging in vivo two-photon imaging innovation knock-down leukemia/lymphoma manufacture mouse model natalizumab neurotoxicity neurovascular unit novel peripheral blood preclinical safety prevent preventive intervention protein protein interaction receptor binding side effect systemic inflammatory response tumor

项目摘要

PROJECT SUMMARY This project studies the mechanism of neurologic toxicity in chimeric antigen receptor (CAR) T cell therapy. CAR T cells are genetically modified, patient derived T cells that use the CAR to recognize and destroy malignant target cells. Although CAR T therapy has shown impressive results against leukemia and lymphoma, approximately 30-40% of patients experience neurologic side effects in the first month after receiving CD19- targeted CAR T cells. This includes cognitive disturbances, seizures, and in rare cases fatal cerebral edema. Systemic cytokine release syndrome after CAR T cell infusion is a well-established risk factor for neurotoxicity, but the connection between systemic inflammation and brain dysfunction is poorly understood. To study the mechanisms of neurotoxicity, we have developed an immunocompetent mouse model. After treatment with high dose CD19-directed murine CAR T cells, mice develop motor and balance difficulties, as well as brain microhemorrhages. Surprisingly, we found that >10% of cortical capillaries are obstructed by white blood cells during neurotoxicity. This was accompanied by capillary remodeling and decreased vessel coverage by pericytes. Based on these findings, we now propose the following experiments: Aim 1: What molecular mechanisms cause white blood cells to plug capillaries during neurotoxicity? We will use in vivo two-photon imaging in mice to determine which cell types cause the capillary plugging – the mouse’s own or the transferred CAR T cells? We will then measure how CAR T cell treatment changes the expression of adhesion molecules in brain capillary endothelial cells and in leukocytes, and test whether blockade of these adhesion interactions can prevent capillary plugging and neurotoxicity. Aim 2: Is neuroendothelial-leukocyte adhesion increase in human microvessels during neurotoxicity? In parallel to our work in mice, we will use a 3D in vitro model of human brain capillaries to measure how soluble factors in patient plasma affect adhesion molecule expression in the endothelium. We will then test whether white blood cells from CAR T cell patients with neurotoxicity have increased predilection for plugging synthetic microvessels that mimic capillaries, and whether we can prevent this plugging by blocking adhesion molecules. Aim 3: Can the strength of cell-cell adhesion signaling separate CAR T cell efficacy from toxicity? We will use quantitative multiplex immunoprecipitation to probe protein-protein interaction networks in CAR T cells that cause high or low neurotoxicity in mice to understand what activation states are conducive to neurotoxicity. We will then test whether knock down of adhesion molecule expression can direct CAR T cells away from a toxicity phenotype by impairing their ability to signal to other cells, and to adhere to the brain microvasculature. This work is innovative because it combines advanced imaging, in vitro modeling techniques, and protein network analysis in a unique collaboration between neuroscience, vascular biology, and oncology. The work is significant because it addresses key safety issues in emerging cancer immunotherapy modalities.

项目概要该项目研究嵌合抗原受体（CAR）T细胞的神经毒性机制治疗。 CAR T 细胞是经过基因改造、源自患者的 T 细胞，使用 CAR 来识别和破坏恶性靶细胞。尽管 CAR T 疗法在治疗白血病和淋巴瘤方面显示出令人印象深刻的效果，大约 30-40% 的患者在接受 CD19- 治疗后的第一个月内会出现神经系统副作用靶向 CAR T 细胞。这包括认知障碍、癫痫发作，以及在极少数情况下致命的脑水肿。 CAR T 细胞输注后的全身细胞因子释放综合征是神经毒性的一个公认的危险因素，但全身炎症和脑功能障碍之间的联系却知之甚少。为了研究神经毒性的机制，我们开发了免疫活性小鼠模型。后使用高剂量 CD19 定向鼠 CAR T 细胞治疗后，小鼠出现运动和平衡困难，如还有脑部微出血。令人惊讶的是，我们发现 >10% 的皮质毛细血管被白色阻塞神经毒性期间的血细胞。这伴随着毛细血管重塑和血管覆盖减少通过周细胞。基于这些发现，我们现在提出以下实验：目标 1：什么分子机制导致白细胞在神经毒性过程中堵塞毛细血管？我们将在小鼠体内使用双光子成像来确定哪些细胞类型导致毛细血管堵塞—— 小鼠自身的还是转移的 CAR T 细胞？然后我们将测量 CAR T 细胞治疗如何改变脑毛细血管内皮细胞和白细胞中粘附分子的表达，并测试是否阻断这些粘附相互作用可以防止毛细血管堵塞和神经毒性。目标 2：神经毒性期间人体微血管中的神经内皮-白细胞粘附是否会增加？在与我们在小鼠身上的工作并行，我们将使用人脑毛细血管的 3D 体外模型来测量溶解度患者血浆中的因素影响内皮细胞中粘附分子的表达。然后我们将测试是否具有神经毒性的 CAR T 细胞患者的白细胞更倾向于堵塞合成模仿毛细血管的微血管，以及我们是否可以通过阻断粘附分子来防止这种堵塞。目标 3：细胞间粘附信号传导的强度能否将 CAR T 细胞的功效与毒性区分开来？我们将使用定量多重免疫沉淀来探测 CAR T 细胞中的蛋白质-蛋白质相互作用网络引起小鼠高或低的神经毒性，以了解哪些激活状态有利于神经毒性。我们然后将测试粘附分子表达的敲低是否可以引导 CAR T 细胞远离毒性通过损害它们向其他细胞发出信号以及粘附到大脑微血管系统的能力来改变表型。这项工作具有创新性，因为它结合了先进的成像、体外建模技术和蛋白质神经科学、血管生物学和肿瘤学之间独特合作的网络分析。工作是意义重大，因为它解决了新兴癌症免疫治疗方式中的关键安全问题。