CD19-specific chimeric antigen receptor (CD19-CAR)-T cell therapies mediate durable responses in late-stage B-cell malignancies, but can be complicated by a potentially severe immune effector cell-associated neurotoxicity syndrome (ICANS). Despite broad efforts, the precise mechanisms of ICANS are not entirely known, and resistance to current ICANSdirected therapies (especially corticosteroids) has been observed. Recent data suggest that inflammatory cytokines and/or targeting of cerebral CD19-expressing pericytes can disrupt the blood-brain barrier and facilitate influx of immune cells, including CAR-T cells. However, specific tools for CD19-CAR-T cell analysis within often minute samples of cerebrospinal fluid (CSF) are not broadly available. Here, we applied our recently developed digital-PCR assays to monitor CD19-CAR-T cell kinetics in CSF and blood in real-world patients with neurotoxicity. Consistently, we observed a CAR-T cell enrichment within CSF in ICANS patients with further progressive accumulation despite intense corticosteroid-containing immuno-chemotherapies in a subset of patients with prolonged and therapy-resistant grade 3-4 neurotoxicity. We used next-generation T-cell receptor-β sequencing to assess the repertoire of treatment-refractory cells. Longitudinal analysis revealed a profound skewing of the T-cell receptor repertoire, which at least partly reflected selective expansion of infused Tcell clones. Interestingly, a major fraction of eventually dominating hyperexpanded T-cell clones were of non-CAR-T cell derivation. These findings hint to a role of therapy-refractory T-cell clones in severe ICANS development and prompt future systematic research to determine if CAR-T cells may serve as 'door openers' and to further characterize both CARpositive and non-CAR-T cells to interrogate the transcriptional signature of these possibly pathologic T cells.

中文翻译：

---

真实世界 CD19 特异性嵌合抗原受体 T 细胞治疗后严重神经毒性中 T 细胞动力学和迁移的分子监测。

CD19 特异性嵌合抗原受体 (CD19-CAR)-T 细胞疗法可介导晚期 B 细胞恶性肿瘤的持久反应，但可能因潜在的严重免疫效应细胞相关神经毒性综合征 (ICANS) 而变得复杂。尽管做出了广泛的努力，但 ICANS 的确切机制尚不完全清楚，并且已经观察到对当前 ICANS 指导疗法（尤其是皮质类固醇）的耐药性。最近的数据表明，炎症细胞因子和/或靶向表达大脑 CD19 的周细胞可以破坏血脑屏障并促进免疫细胞（包括 CAR-T 细胞）的流入。然而，在脑脊液 (CSF) 的微小样本中进行 CD19-CAR-T 细胞分析的特定工具并不广泛可用。这里，我们应用了我们最近开发的数字 PCR 检测方法来监测现实世界中具有神经毒性的患者脑脊液和血液中的 CD19-CAR-T 细胞动力学。一致地，我们观察到 ICANS 患者脑脊液中的 CAR-T 细胞富集，尽管在长期和治疗耐药的 3-4 级神经毒性患者亚群中进行了强烈的含皮质类固醇的免疫化疗，但 CAR-T 细胞进一步进行性积累。我们使用下一代 T 细胞受体-β 测序来评估难治性细胞的所有组成部分。纵向分析揭示了 T 细胞受体库的严重偏差，这至少部分反映了输注 T 细胞克隆的选择性扩增。有趣的是，最终占主导地位的超扩增 T 细胞克隆的主要部分是非 CAR-T 细胞衍生的。