肿瘤细胞治疗研究
英文原题：PD-L1-armored CD19/CD22 dual-targeted CAR-T cell co-infusion bridging to allogeneic hematopoietic stem cell transplantation achieves 7-year sustained remission in an adult patient with early relapsed, chemorefractory B-cell acute lymphoblastic leukemia: a case report.
患者在第14天输注后达到微小残留病（MRD）阴性完全缓解（CR），经8色流式细胞术（MFC）（检测灵敏度：0.01%）和免疫球蛋白重链（IGH）基因重排的二代测序（NGS）（检测限[LOD]：10⁻⁶）确认。
背景：抗原逃逸以及肿瘤微环境（TME）中 PD-L1/PD-1 轴介导的免疫抑制，是成人复发/难治性（R/R）B 急性淋巴细胞白血病（B-ALL）治疗失败的主要原因。单独使用 CD19 靶向 CAR-T 难以克服这两种核心耐药机制；对高危、化疗难治性成人 B-ALL 患者而言，实现长期持久缓解仍是关键未满足需求。 方法：这是一项单中心回顾性病例报告，患者为早期复发、化疗难治性成人 B-ALL。研究者使用患者自体外周血单个核细胞（PBMC）制备两种独立 CAR-T 产品：（1）CD19 CAR-T，含抗 CD19 单链可变片段（scFv）、CD28 跨膜结构域、4-1BB 共刺激结构域和 CD3 信号结构域；（2）PD-L1 装甲型 CD22 CAR-T，含抗 CD22 scFv、CD8 跨膜结构域、4-1BB 共刺激结构域、CD3 信号结构域，以及膜锚定抗 PD-L1 scFv，以实现空间局限的免疫检查点调节。两种产品共同输注，剂量分别为 CD19 CAR-T 5.0×10⁵ 个细胞/kg、PD-L1 装甲型 CD22 CAR-T 3.1×10⁵ 个细胞/kg。研究评估了该方案作为异基因造血干细胞移植（allo-HSCT）桥接治疗的可行性、抗白血病疗效和长期安全性。治疗在河北医科大学第二医院伦理委员会批准的机构同情用药项目下实施（批准号：2017-R207），所有治疗前均取得书面知情同意。通过八色多参数流式细胞术（MFC）、常规 G 显带细胞遗传学分析和多重白血病融合基因筛查完成 B-ALL 全面诊断。整个治疗过程中定期腰椎穿刺并进行三联鞘内化疗（地塞米松 5 mg、甲氨蝶呤 10 mg、阿糖胞苷 30 mg）；脑脊液常规、生化和流式检测均未见异常，任何时间点均无中枢神经系统白血病受累证据。本研究是单例回顾性观察分析，并非前瞻性干预临床试验，因此按机构及国家回顾性研究监管指南，无需进行临床试验注册。 结果：输注后第 14 天患者达到微小残留病（MRD）阴性完全缓解（CR），经八色 MFC（检测灵敏度 0.01%）和免疫球蛋白重链（IGH）基因重排二代测序（NGS；检测下限 10⁻⁶）证实。输注后第 10 天 CAR-T 细胞体内扩增达到峰值；CD19 CAR-T 和 CD22 CAR-T 分别占循环 CD3⁺ T 淋巴细胞的 43.25% 和 14.58%。患者仅出现 1 级细胞因子释放综合征（CRS），按美国移植与细胞治疗学会（ASTCT）共识标准分级，经支持治疗后完全缓解；未出现免疫效应细胞相关神经毒性综合征（ICANS）。巩固性 allo-HSCT 后造血迅速重建，中性粒细胞于移植后第 +12 天植入，血小板于第 +14 天植入，与该机构单倍体相合移植的中位植入时间一致。移植后第 +30 天，短串联重复序列（STR）分析证实供者完全嵌合（99.86%）。值得注意的是，患者连续 7 年维持 MRD 阴性持续缓解，未发生急性或慢性移植物抗宿主病（GVHD）。摘要原文在“未发生迟发治疗相关不良事件”一处截断，后续内容缺失。
BACKGROUND: Antigen escape and PD-L1/PD-1 axis-mediated immunosuppression in the tumor microenvironment (TME) are the predominant drivers of treatment failure in adult patients with relapsed/refractory (R/R) B-cell acute lymphoblastic leukemia (B-ALL). CD19-targeted CAR-T monotherapy has limited ability to overcome these two core resistance mechanisms, and achieving long-term durable remission remains a critical unmet clinical need in adults with high-risk, chemorefractory B-ALL. METHODS: This is a single-center, retrospective case report of an adult patient with early-relapsed, chemorefractory B-ALL. Two independent CAR-T products were manufactured from the patient's autologous peripheral blood mononuclear cells (PBMCs): (1) CD19 CAR-T cells containing an anti-CD19 single-chain variable fragment (scFv), a CD28 transmembrane domain, a 4-1BB co-stimulatory domain, and a CD3 signaling domain; (2) PD-L1-armored CD22 CAR-T cells containing an anti-CD22 scFv, a CD8 transmembrane domain, a 4-1BB co-stimulatory domain, a CD3 signaling domain, and a membrane-tethered anti-PD-L1 scFv for spatially restricted immune checkpoint modulation. The two CAR-T products were co-infused at doses of 5.0 10 5 cells/kg (CD19 CAR-T) and 3.1 10 5 cells/kg (PD-L1-armored CD22 CAR-T), respectively. We evaluated the feasibility, anti-leukemic efficacy, and long-term safety profile of this regimen as a bridging strategy to allogeneic hematopoietic stem cell transplantation (allo-HSCT). This treatment was administered under an institutional compassionate use program approved by the Ethics Committee of the Second Hospital of Hebei Medical University (approval number: 2017-R207), with written informed consent obtained from the patient prior to all treatment procedures. Comprehensive diagnostic workup for B-ALL was performed using 8-color multiparameter flow cytometry (MFC), conventional G-banding cytogenetic analysis, and multiplex leukemia fusion gene screening. Serial lumbar punctures with triple intrathecal chemotherapy (dexamethasone 5 mg + methotrexate 10 mg + cytarabine 30 mg) were performed throughout the treatment course; no abnormalities were detected in cerebrospinal fluid (CSF) routine, biochemistry, or flow cytometry assays, and no evidence of central nervous system (CNS) leukemia involvement was observed at any time point. This study is a retrospective observational analysis of a single clinical case, not a prospective interventional clinical trial, and thus was exempt from clinical trial registration requirements in accordance with institutional and national regulatory guidelines for retrospective observational studies. RESULTS: The patient achieved minimal residual disease (MRD)-negative complete remission (CR) on day 14 post-infusion, as confirmed by 8-color MFC (detection sensitivity: 0.01%) and next-generation sequencing (NGS) of immunoglobulin heavy chain (IGH) gene rearrangements (limit of detection [LOD]: 10 -6 ). Peak in vivo expansion of CAR-T cells was observed on day 10 post-infusion, with CD19 CAR-T cells accounting for 43.25% and CD22 CAR-T cells accounting for 14.58% of circulating CD3 + T lymphocytes. Only grade 1 cytokine release syndrome (CRS), per the American Society for Transplantation and Cellular Therapy (ASTCT) consensus criteria, occurred and resolved completely with supportive care; no immune effector cell-associated neurotoxicity syndrome (ICANS) was observed. Following consolidative allo-HSCT, rapid hematopoietic reconstitution was achieved, with neutrophil engraftment on day +12 and platelet engraftment on day +14, consistent with the median engraftment timeline for haploidentical HSCT at our institution. Complete donor chimerism (99.86%) was confirmed by short tandem repeat (STR) analysis on day +30 post-transplantation. Notably, the patient maintained MRD-negative sustained remission for 7 consecutive years, with no occurrence of acute or chronic graft-versus-host disease (GVHD) or late treatment-related adverse ev
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