The reparative immunologic consequences of stem cell transplantation as a cellular therapy for refractory Crohn’s disease

Background Treatment strategies for Crohn’s disease (CD) suppress diverse inflammatory pathways but many patients remain refractory to treatment. Autologous hematopoietic stem cell transplantation (SCT) is an emerging therapy for medically-refractory CD though the mechanisms through which it circumvents refractory pathophysiology are unknown. Objective The objective of this study is to understand how the immune system reconstitutes post-SCT and whether SCT may function as a cellular therapy restoring appropriately responsive immune cell populations from hematopoietic stem cells (HSCs). Design Adults with CD with active clinical and endoscopic disease who failed available medical therapies were enrolled in a Phase II study of SCT for refractory CD (n=19). Blood and intestinal samples were collected longitudinally and analyzed using CyTOF, scRNA-seq and TCRβ-seq. Stem cell autografts were functionally assayed in mouse xenograft models. Results scRNA-seq and CyTOF analyses reveal that SCT predominantly affected the intestinal myeloid lineage with loss of inflammatory populations and return of macrophages capable of supporting mucosal healing. Xenograft models using patient HSCs suggested HSCs support the early reconstitution of the myeloid lineage and reveal an impairment of short and long-term HSC engraftment that may determine SCT outcomes. Conclusions This study suggests SCT functions as a myeloid-directed cellular therapy reinforcing the critical role of macrophages in refractory CD pathophysiology and as a target for cellular therapies. Furthermore, we report an unrecognized functional heterogeneity among HSC subpopulations in CD that may be relevant to our understanding of CD treatment and pathophysiology. Samples of tissue from endoscopic biopsies, peripheral blood and peripheral mobilized stem cells were collected from subjects with CD enrolled for SCT. Blood and tissue samples were processed for 10x Genomics single cell RNA sequencing (scRNA-seq) and bulk T cell receptor beta (TCRβ) sequencing through Adaptive technologies.

背景：克罗恩病（Crohn’s disease, CD）的治疗策略可抑制多条炎症通路，但仍有大量患者对治疗产生耐药性。自体造血干细胞移植（autologous hematopoietic stem cell transplantation, SCT）是针对药物难治性CD的新兴疗法，但其规避耐药病理生理过程的具体机制尚不明确。
目标：本研究旨在阐明造血干细胞移植后免疫系统的重建过程，并探讨SCT是否可作为一种细胞疗法，从造血干细胞（hematopoietic stem cells, HSCs）中恢复功能正常且应答适宜的免疫细胞群。
研究设计：本研究纳入经现有药物治疗失败的活动性临床及内镜下CD成人患者，开展针对难治性CD的SCT II期临床试验（n=19）。研究人员纵向采集血液与肠道样本，采用质谱流式细胞术（CyTOF）、单细胞RNA测序（scRNA-seq）及T细胞受体β链测序（TCRβ-seq）进行分析；同时在小鼠异种移植模型中对自体干细胞移植物开展功能学检测。
研究结果：单细胞RNA测序与质谱流式细胞术分析结果显示，SCT主要作用于肠道髓系细胞谱系，可清除炎症性细胞亚群，并恢复能够促进黏膜愈合的巨噬细胞。采用患者造血干细胞构建的异种移植模型表明，HSCs可促进髓系细胞谱系的早期重建，同时还发现短期与长期造血干细胞的植入功能存在损伤，这或可决定SCT的治疗结局。
研究结论：本研究表明，SCT可作为一种以髓系细胞为靶向的细胞疗法，印证了巨噬细胞在难治性CD病理生理过程中的关键作用，同时也为细胞疗法提供了潜在靶点。此外，本研究首次揭示了CD患者HSCs亚群间存在未被认知的功能异质性，这或有助于加深我们对CD治疗与病理生理机制的理解。研究人员从入组SCT临床试验的CD患者中采集内镜活检组织、外周血及外周动员干细胞样本，并通过Adaptive Technologies平台完成样本处理，用于10x Genomics单细胞RNA测序及批量T细胞受体β链测序。