X-连锁神经连粘蛋白4（NLGN4X）携带人类白细胞抗原（HLA）-A2限制性肿瘤相关抗原，在人类胶质瘤中过表达。发现该抗原能够诱导新诊断的胶质母细胞瘤患者接受多肽疫苗接种后产生特异性细胞毒性T细胞反应。通过对疫苗接种后的NLGN4X四聚体分选的T细胞进行基于液滴的单细胞T细胞受体（TCR）测序，发现了该TCR。将鉴定的TCR传递到Jurkat T细胞和人体原代T细胞（NLGN4X-TCR-T）。使用流式细胞术和细胞毒性试验对NLGN4X-TCR-T进行功能分析。在携带NLGN4X表达实验性胶质瘤的NOD scid gamma(NSG)主要组织相容性复合物（MHC）I/II敲除（NSG MHC I/II KO）小鼠中评估侧脑室内注射的NLGN4X-TCR-T的治疗效果。使用HLA-A * 02限制性疫苗诱导的结合于NLGN4X131-139的TCR用于治疗。通过多种细胞模型展示了该NLGN4X特异性TCR的反应性、细胞毒性和多功能性。与对照组相比，侧脑室内注射NLGN4X-TCR-T能够延长实验性胶质瘤携带NSG MHC I/II KO小鼠的生存时间，并呈现44.4%的客观反应率。NLGN4X-TCR-T在临床前胶质母细胞瘤模型中显示出疗效。从全球范围来看，我们首次提供了利用疫苗诱导的人类TCR进行脑胶质瘤患者常备治疗的治疗性证据。© The Author(s) 2023. Published by Oxford University Press on behalf of the Society for Neuro-Oncology.

Neuroligin 4 X-linked (NLGN4X) harbors a human leukocyte antigen (HLA)-A2-restricted tumor-associated antigen, overexpressed in human gliomas, that was found to induce specific cytotoxic T cell responses following multi-peptide vaccination in patients with newly- diagnosed glioblastoma.T cell receptor (TCR) discovery was performed using droplet-based single cell TCR sequencing of NLGN4X-tetramer-sorted T cells post vaccination. The identified TCR was delivered to Jurkat T cells and primary human T cells (NLGN4X-TCR-T). Functional profiling of NLGN4X-TCR-T was performed by flow cytometry and cytotoxicity assays. Therapeutic efficacy of intracerebroventricular NLGN4X-TCR-T was assessed in NOD scid gamma (NSG) major histocompatibility complex (MHC) I/II knockout (KO) (NSG MHC I/II KO) mice bearing NLGN4X-expressing experimental gliomas.An HLA-A *02-restricted vaccine-induced T cell receptor specifically binding NLGN4X131-139 was applied for therapeutic use. Reactivity, cytotoxicity, and polyfunctionality of this NLGN4X-specific TCR is demonstrated in various cellular models. Intracerebroventricular administration of NLGN4X-TCR-T prolongs survival and leads to an objective response rate (ORR) of 44.4 % in experimental gliomas-bearing NSG MHC I/II KO mice compared to 0.0 % in control groups, respectively.NLGN4X-TCR-T demonstrates efficacy in a preclinical glioblastoma model. On a global scale, we provide first evidence for the therapeutic retrieval of vaccine-induced human TCRs for the off-the-shelf treatment of glioblastoma patients.© The Author(s) 2023. Published by Oxford University Press on behalf of the Society for Neuro-Oncology.