Chimeric antigen receptor (CAR) T cells hold promise as a therapy for B-cell–derived malignancies, and despite their impressive initial response rates, a significant proportion of patients ultimately experience relapse. Although recent studies have explored the mechanisms of in vivo CAR T-cell function, little is understood about the activation of surrounding CAR bystander T cells and their potential to enhance tumor responses. We performed single-cell RNA sequencing on nonhuman primate (NHP) and patient-derived T cells to identify the phenotypic and transcriptomic hallmarks of bystander activation of CAR T cells following B-cell–targeted CAR T-cell therapy. Using a highly translatable CD20 CAR NHP model, we observed a distinct population of activated CD8 CAR T cells emerging during CAR T-cell expansion. These bystander CD8 CAR T cells exhibited a unique transcriptional signature with upregulation of natural killer-cell markers (, , and ), chemokines, and chemokine receptors (, and ), and downregulation of naïve T-cell-associated genes ( and ). A transcriptionally similar population was identified in patients after a tisagenlecleucel infusion. Mechanistic studies revealed that interleukin-2 (IL-2) and IL-15 exposure induced bystander-like CD8 T cells in a dose-dependent manner. In vitro activated and patient-derived T cells with a bystander phenotype efficiently killed leukemic cells through a T-cell receptor–independent mechanism. Collectively, to our knowledge, these data provide the first comprehensive identification and profiling of CAR bystander CD8 T cells following B-cell–targeting CAR T-cell therapy and suggest a novel mechanism through which CAR T-cell infusion might trigger enhanced antileukemic responses. Patient samples were obtained from the trial #NCT03369353, registered at .

中文翻译：

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B 细胞定向 CAR T 细胞疗法可激活患者和非人灵长类动物中的 CD8+ 细胞毒性 CARneg 旁观 T 细胞

嵌合抗原受体 (CAR) T 细胞有望成为 B 细胞衍生恶性肿瘤的治疗方法，尽管其初始缓解率令人印象深刻，但仍有相当一部分患者最终出现复发。尽管最近的研究已经探索了体内 CAR T 细胞功能的机制，但人们对周围 CAR 旁观者 T 细胞的激活及其增强肿瘤反应的潜力知之甚少。我们对非人灵长类动物 (NHP) 和患者来源的 T 细胞进行了单细胞 RNA 测序，以确定 B 细胞靶向 CAR T 细胞治疗后旁观者激活 CAR T 细胞的表型和转录组特征。使用高度可翻译的 CD20 CAR NHP 模型，我们观察到在 CAR T 细胞扩增过程中出现了独特的活化 CD8 CAR T 细胞群。这些旁观者 CD8 CAR T 细胞表现出独特的转录特征，其中自然杀伤细胞标记（、、和）、趋化因子和趋化因子受体（、、）的上调，以及幼稚 T 细胞相关基因（、）的下调。在输注 tisagenlecleucel 后，在患者中发现了转录相似的群体。机制研究表明，白介素 2 (IL-2) 和 IL-15 暴露以剂量依赖性方式诱导旁观者样 CD8 T 细胞。体外激活的患者来源的具有旁观者表型的 T 细胞通过 T 细胞受体独立机制有效杀死白血病细胞。总的来说，据我们所知，这些数据首次对 B 细胞靶向 CAR T 细胞治疗后的 CAR 旁观者 CD8 T 细胞进行全面鉴定和分析，并提出了一种新机制，通过该机制 CAR T 细胞输注可能会触发增强的抗白血病反应。患者样本从试验#NCT03369353 中获得，注册于 。