Label-free imaging of CAR T cell metabolism

CAR T 细胞代谢的无标记成像

• 批准号: 10751581
• 负责人: Christian Capitini
• 金额: $ 66.2万
• 依托单位: MORGRIDGE INSTITUTE FOR RESEARCH, INC.
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2028-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10751581
• 关键词: Antitumor Response; Autologous; B-Cell Lymphomas; Benchmarking; Biological Assay; Biological Markers; CAR T cell therapy; CD19 gene; Cancer Patient; Cell physiology; Cells; Cellular Metabolic Process; Clinical; Clinical Trials; Combined Modality Therapy; Cytolysis; Dasatinib; Data; Disease remission; FDA approved; Flow Cytometry; Frequencies; Genetic Engineering; Goals; Human; Hypoxia; Image; Imaging technology; Immunofluorescence Immunologic; In Vitro; Infiltration; Infusion procedures; Intervention; Knowledge; Label; Malignant Neoplasms; Measurement; Measures; Memory; Metabolic; Methods; Modeling; Monitor; Neuroblastoma; Optics; Patient Selection; Patients; Phenotype; Process; Production; Resolution; Resources; Sampling; Solid Neoplasm; Source; Specificity; Spleen; T cell response; T cell therapy; T memory cell; T-Lymphocyte; Technology; Technology Assessment; Testing; Time; Xenograft procedure; alternative treatment; cancer cell; cancer therapy; cellular imaging; chimeric antigen receptor; chimeric antigen receptor T cells; cytokine; exhaust; exhaustion; fitness; fluorescence imaging; improved; in vivo; manufacture; metabolic imaging; mouse model; neoplastic cell; patient screening; predictive modeling; process optimization; response; response biomarker; single cell technology; stem cells; therapy development; tumor

PROJECT SUMMARY / ABSTRACT The goal of this proposal is to develop non-invasive single-cell technologies to improve the potency of T cell therapies against cancer. The first 6 chimeric antigen receptor (CAR) T cell therapies were recently approved and >800 CAR and T cell therapies are in clinical trials. However, barriers remain in achieving durable remissions (>1 year) for ~50% of patients who receive CAR T cell therapy. Due to the rapid development of these therapies and a great need for process optimization, we focus on improving three translational roadblocks to effective CAR T cell therapy: (1) screening patients whose T cells are unfit for CAR T cell manufacturing, (2) optimizing in vitro CAR T cell production for higher potency, and (3) identifying metabolic features of potent CAR T cells in vivo. CAR T cell therapy could be improved by enriching for naïve and stem cell memory (SCM) T cells in starting materials and final products. Deficiencies in naïve and SCM T cells occurs in ~50% of untreated cancer patients, and manufacturing autologous CAR T cell products from these sources has been unsuccessful. Even if SCM T cells can be isolated, after CAR incorporation, the expansion process typically diminishes potency through T cell exhaustion. After infusion, the presence of memory-like phenotypes in vivo correlate with better responses. To date, there are no robust, non-destructive technologies to monitor CAR T cell manufacturing to optimize production and assess potency in vivo at a single-cell level. These issues limit the impact of CAR T cell therapy. Current approaches to measure T cell function are labor-intensive, destructive, or lack single-cell resolution, which limits the frequency or specificity of these measurements. For CAR T cell therapy to realize its clinical potential, new methods are needed to monitor T cells for optimal potency throughout manufacturing and post- infusion. Changes in cell metabolism provide an attractive yet under-explored assay to track T cell potency. Previous studies, including our own, show that T cells undergo drastic metabolic changes with activation, and that naïve, exhausted, and memory-like T cells have distinct metabolic features. Our preliminary data shows that non-invasive single-cell imaging of the fluorescence intensity and lifetime of NAD(P)H and FAD (optical metabolic imaging, or OMI) can predict CAR T cell manufacturing conditions that produce a more vs. less potent anti-tumor response in vivo. Given these metabolic features of CAR T cell potency, we propose to determine whether label-free OMI of T cell autofluorescence and multivariate models can identify patient T cell fitness, optimal in vitro expansion conditions, and in vivo cell biomarkers of potent and persistent CAR T cell response. Overall, these technologies will streamline processes and interventions for consistently potent T cell therapy and increase our knowledge of CAR T cell metabolism in vitro and in vivo.

项目概要/摘要 该提案的目标是开发非侵入性单细胞技术以提高 T 细胞的效力 抗癌疗法。首批 6 种嵌合抗原受体 (CAR) T 细胞疗法最近获得批准 超过 800 种 CAR 和 T 细胞疗法正在进行临床试验。然而，实现持久缓解仍存在障碍 （>1 年）约 50% 接受 CAR T 细胞治疗的患者。由于这些疗法的快速发展 由于对流程优化的巨大需求，我们专注于改善有效 CAR 的三个转化障碍 T细胞疗法：(1)筛选T细胞不适合CAR T细胞制造的患者，(2)体外优化 CAR T 细胞生产以获得更高的效力，以及 (3) 识别有效 CAR T 细胞在体内的代谢特征。 CAR T 细胞疗法可以通过在起始阶段富集幼稚 T 细胞和干细胞记忆 (SCM) T 细胞来改善 材料和最终产品。约 50% 未经治疗的癌症患者存在幼稚 T 细胞和 SCM T 细胞缺陷， 从这些来源生产自体 CAR T 细胞产品并不成功。即使单片机T 细胞可以被分离，在 CAR 掺入后，扩增过程通常会降低 T 细胞的效力 精疲力尽。输注后，体内记忆样表型的存在与更好的反应相关。到 迄今为止，还没有强大的、非破坏性的技术来监控 CAR T 细胞的制造以优化 在单细胞水平上生产并评估体内效力。这些问题限制了 CAR T 细胞疗法的影响。 目前测量 T 细胞功能的方法是劳动密集型的、破坏性的或缺乏单细胞分辨率， 这限制了这些测量的频率或特异性。使CAR T细胞疗法实现临床 潜力，需要新的方法来监测 T 细胞在整个制造和后期的最佳效力 输液。细胞代谢的变化为追踪 T 细胞效力提供了一种有吸引力但尚未充分探索的测定方法。 之前的研究（包括我们自己的研究）表明，T 细胞在激活过程中会经历剧烈的代谢变化，并且 幼稚、疲惫和记忆样 T 细胞具有独特的代谢特征。我们的初步数据表明 NAD(P)H 和 FAD 的荧光强度和寿命的非侵入性单细胞成像（光学 代谢成像（OMI）可以预测 CAR T 细胞的制造条件，从而产生更多或更少的结果 体内有效的抗肿瘤反应。鉴于 CAR T 细胞效力的这些代谢特征，我们建议 确定 T 细胞自发荧光和多变量模型的无标记 OMI 是否可以识别患者 T 细胞 强效和持久 CAR T 细胞的适应性、最佳体外扩增条件和体内细胞生物标志物 回复。总体而言，这些技术将简化持续有效的 T 细胞的流程和干预措施 疗法并增加我们对 CAR T 细胞体外和体内代谢的了解。