Immune reconstitution following autologous and allogeneic stem cell transplant

自体和同种异体干细胞移植后的免疫重建

• 批准号: 8938439
• 负责人: Frances Hakim
• 金额: $ 97.45万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8938439
• 关键词: Activities of Daily Living; Acute; Acute leukemia; Adoptive Transfer; Affect; Aftercare; Allogenic; Autologous; Autologous Transplantation; B-Cell Activation; Biological Assay; Biopsy; Blood; Blood Cells; Blood Circulation; Bronchial Lavages; Bronchiolitis Obliterans; CXCR3 gene; Cells; Childhood; Chimerism; Chronic; Clinical; Clinical Protocols; Clinical Trials; Complication; Core Facility; Cutaneous; Data; Dendritic Cells; Dermatology; Development; Disease; Enzyme-Linked Immunosorbent Assay; Fibrosis; Flow Cytometry; Frequencies; Functional disorder; Gene Expression; Genes; Glucocorticoids; Goals; Hematopoietic Stem Cell Transplantation; Homologous Transplantation; Imatinib; Immune; Immunohistochemistry; Immunologic Monitoring; Immunosuppression; In Situ; Infection; Inflammation; Inflammatory; Inflammatory Infiltrate; Interferon Type I; Interferon-alpha; Interferons; K-Series Research Career Programs; Laboratory Finding; Leukocytes; Leukotriene Receptor; Link; Lung; Lymphocyte; Lymphocyte Depletion; Lymphocyte Subset; Lymphoma; Malignant lymphoid neoplasm; Marrow; Measures; Mediating; Microarray Analysis; Modeling; Molecular; Monitor; Morbidity - disease rate; Mouthwash; Multiple Myeloma; Natural History; Oral; Outcome; Pathogenesis; Pathway interactions; Patients; Pattern; Phagocytosis; Plasma; Population; Predisposition; Process; Production; Protocols documentation; RNA; Receptor Gene; Recovery; Regimen; Regulatory T-Lymphocyte; Relapse; Reporter; Reporting; Research; Resolution; Rest; Role; Salivary; Sampling; Scientist; Secure; Services; Siblings; Signal Transduction; Sorting - Cell Movement; Stem cell transplant; Steroids; Symptoms; T-Cell Receptor; T-Lymphocyte; Testing; Therapy Clinical Trials; Time; Tissues; Transforming Growth Factor beta; Transplant Recipients; Transplantation; Transplantation Immunology; United States National Institutes of Health; Up-Regulation; Vaccines; Work; arm; base; chemokine; chemokine receptor; chronic graft versus host disease; collaborative trial; conditioning; cytokine; graft vs host disease; in vivo; inhibitor/antagonist; leukemia; migration; monocyte; mortality; multidisciplinary; novel; patient population; peripheral blood; pre-clinical; receptor; receptor expression; reconstitution; research study; response; trafficking; tumor

The Preclinical Development and Clinical Monitoring Facility (PDCMF) projects have developed from transplantation protocols implemented by the clinical staff of ETIB. Using peripheral blood and marrow, and tumor and CGVHD tissue biopsies, we have evaluated lymphocyte subsets, cytokine content, T cell receptor repertoire diversity and thymopoietic activity. All data are incorporated into protocol-specific spreadsheets, linking samples to protocol arms and transplant time points, and are accessible by branch clinicians over secure NIH networks. In several ongoing ETIB clinical trials of autologous and allogeneic stem cell transplantation therapies (04-C-0055, 07-C-0195, 11-C-0016, 11-C-0136; PIs Daniel Fowler, Steven Pavletic, Claude Sportes, Ronald Gress and Kirsten Williams) we have assessed lymphodepletion during conditioning and immune reconstitution after transplant. We currently are using multiparameter flow cytometry to characterize lymphocyte repopulation, plasma ELISA to monitor cytokines and molecular assays of gene expression and T cell receptor repertoire diversity. We correlate the laboratory findings with clinical outcomes of relapse, infection, and chronic graft versus host disease (CGVHD). These parallel studies involve analysis of ongoing trials that cover the breadth of current transplant practices: non-myeloablative reduced intensity allogeneic transplantation for lymphoma using sibling and unrelated donors, autologous transplantation for myeloma, and myeloablative transplant for acute leukemias. We also support ongoing studies of lineage-specific immune reconstitution in patients transplanted for monogenic immune deficiencies involving GATA2 or DOCK8 (09-C-0096, 10-C-0174, PI: Dennis Hickstein) by utilizing the ETIB Flow Cytometry Facility (William Telford) to sort lymphocytes for subset-specific donor chimerism analysis. These immune monitoring studies have contributed to a report on the efficacy of EPOCH-F, a novel non-myeloablative induction regimen, in allogeneic stem cell transplantation in patients with multiple myeloma and lymphoid malignancies (Salit et al, 2012; Salit et al, 2013), as well as reports on allogeneic transplant supported by adoptive transfer of T2-RAPA cells (Fowler et al, Blood 2013). Chronic graft vs host disease (CGVHD), the principle cause of non-relapse morbidity and mortality after allogeneic transplantation, is a major focus for research in the PDCMF core. We have supported the efforts of the multidisciplinary clinical team in an ongoing natural history protocol studying patients who have developed CGVHD (P.I. Steven Pavletic: 04-C-0281) by storing patient blood, biofluids and tissue for research. We have contributed to the research study by analyzing mechanisms of CGVHD pathogenesis. Furthermore, we have supported four therapeutic trials for CGVHD patient populations: (1) We are assessing leukotriene receptor (LTR) expression in leukocytes and in bronchial lavage cells to define the role of LTR in progressive fibrosis of lung airways in patients developing bronchiolitis obliterans, a severe complication of CGVHD (08-C-0097: P. I. Ronald Gress and Kirsten Williams:). (2) We have assessed the effect of Imatinib, a TGFbeta signaling inhibitor, on Th17 and Treg populations, as part of a collaborative trial testing Imatinib therapy on sclerotic cutantaneous CGVHD (Dermatology and Pediatric Branch, protocol 08-C-0148, P.I. Edward Cowen and Kristin Baird). (3) We are analyzing changes in inflammatory gene expression in a trial of Pomalidomide (12-C-0197; P.I. Steven Pavletic) to assess efficacy against fibrosis in CGVHD patients. (4) Finally we support tissue immunohistochemistry analyses and salivary cytokine tests in a trial of a mouthwash containing a potent steroid, Clobetasol, as a topical therapy for severe oral CGVHD (12-C-0068; P.I. Steven Pavletic/Jacqueline Mays). As part of our studies of the pathogenesis of CGVHD, we have characterized regulatory T cells in CGVHDthrough coordinated studies of blood and tissue. These studies determined that FoxP3+ regulatory T cells (Treg) increased in proportion to T effectors in tissue infiltrates in oral and cutaneous lichenoid cGVHD. Both T effector and FoxP3+ Treg cells expressed Tbet and the chemokine receptor CXCR3, consistent with a common mechanism of chemokine-mediated migration into tissue. Furthermore, functional markers (ICOS and CD39) and chemokine receptors (CXCR3) were both present in a higher proportion of FoxP3+ cells in tissues than in peripheral blood, consistent with recruitment and activation of Treg in cGVHD target tissues. Finally, the 'activated' CD45RA-FoxP3hi subset of Treg cells, which highly expresses functional markers, was found in comparable frequencies in cGVHD patients and normal controls, despite a significant deficit in naive 'resting' Treg. These findings are consistent with Treg functional capacity in cGVHD, and support efforts to expand Treg cells in vivo as therapy. (Imanguli et al, Leukemia 2014) In studies of CGVHD pathogenesis, we profiled gene expression of circulating monocytes in CGVHD patients, to use these cells as in situ reporter cells for systemic cytokine patterns in order to test the hypothesis that Interferon (IFN)-induced inflammatory processes may underlie many of the systemic processes in CGVHD (Imanguli et al, 2009; Hakim, 2010). Based on microarray analysis and confirmed by multiplex RNA assessments (supported by an NCI Staff Scientist Career Development Award), we determined that pathways induced by IFN were consistently upregulated across a broad spectrum of CGVHD patients, both those with severe inflammatory infiltrates in tissue and those with widespread sclerotic involvement. IFN-inducible genes, including ones specifically induced by type I IFN, were upregulated in parallel at the time of onset of CGVHD, and were reduced during treatment and after resolution of CGVHD symptoms. In addition, we found a consistent upregulation of receptor genes from the innate immune TLR/NLR/CLR pathways; these pathways are triggered by debris from dead cells to stimulate phagocytosis, induce IFNa production, and form inflammasomes. Many of these receptors are also inducible by IFN, consistent with a self-reinforcing inflammatory process sustaining CGVHD. Furthermore, through assessments of glucocorticoid-inducible genes, we discriminated the confounding effects of immunosuppressive steroids from disease-associated gene patterns. Finally, we corroborated the gene expression data through analyses of IFN-inducible factors in plasma and tissue. These findings substantiated a role for IFN in lymphocyte and dendritic cell trafficking into tissue, and, through upregulation of BAFF, an involvement of IFN in B cell activation in CGVHD. These interlocking assessments, performed on a broad spectrum of patients severely affected with CGVHD, support a new model for the initiation and persistence of CGVHD. This model defines CGVHD as a disorder of inflammation-driven immune dysregulation and these results support a new range of options for therapy of CGVHD through interdiction of interferon pathways.

临床前开发和临床监测设施（PDCMF）项目是根据ETIB临床人员实施的移植方案开发的。使用外周血和骨髓以及肿瘤和CGVHD组织活检，我们评估了淋巴细胞亚群，细胞因子含量，T细胞受体库库多样性和胸腺异端活性。所有数据均纳入特定于协议的电子表格，将样品与协议臂和移植时间点联系起来，并可以通过安全的NIH网络访问分支机构。 In several ongoing ETIB clinical trials of autologous and allogeneic stem cell tr​​ansplantation therapies (04-C-0055, 07-C-0195, 11-C-0016, 11-C-0136; PIs Daniel Fowler, Steven Pavletic, Claude Sportes, Ronald Gress and Kirsten Williams) we have assessed lymphodepletion during conditioning and immune reconstitution after移植。目前，我们正在使用多参数流式细胞仪来表征淋巴细胞再生，血浆ELISA来监测基因表达和T细胞受体库库多样性的细胞因子和分子测定。我们将实验室发现与复发，感染和慢性移植与宿主疾病（CGVHD）的临床结果相关联。这些平行研究涉及涵盖当前移植实践广度的正在进行的试验：使用兄弟姐妹和不相关供体的淋巴瘤的非肌瘤性降低强度同种异体移植，骨髓瘤自体移植，以及肌力移植的急性白血病。 We also support ongoing studies of lineage-specific immune reconstitution in patients transplanted for monogenic immune deficiencies involving GATA2 or DOCK8 (09-C-0096, 10-C-0174, PI: Dennis Hickstein) by utilizing the ETIB Flow Cytometry Facility (William Telford) to sort lymphocytes for subset-specific donor chimerism analysis. These immune monitoring studies have contributed to a report on the efficacy of EPOCH-F, a novel non-myeloablative induction regimen, in allogeneic stem cell tr​​ansplantation in patients with multiple myeloma and lymphoid malignancies (Salit et al, 2012; Salit et al, 2013), as well as reports on allogeneic transplant supported by adoptive transfer of T2-RAPA cells (Fowler et al, Blood 2013).慢性移植与宿主疾病（CGVHD）是同种异体移植后非释放发病率和死亡率的主要原因，是PDCMF核心研究的主要重点。我们通过存储患者的血液，生物流体和组织进行研究来支持开发CGVHD的患者（P.I. Steven Pavletic：04-C-0281）的患者的跨学科临床团队的努力。我们通过分析CGVHD发病机理的机制为研究做出了贡献。此外，我们支持了CGVHD患者人群的四项治疗试验：（1）我们正在评估白细胞和支气管灌洗细胞中的白三烯受体（LTR）表达，以定义LTR在肺纤维化中的作用，以确定LTR在肺部进行性纤维化在患者中的进行性纤维化的作用。 Gress和Kirsten Williams :)。 (2) We have assessed the effect of Imatinib, a TGFbeta signaling inhibitor, on Th17 and Treg populations, as part of a collaborative trial testing Imatinib therapy on sclerotic cutantaneous CGVHD (Dermatology and Pediatric Branch, protocol 08-C-0148, P.I. Edward Cowen and Kristin Baird). （3）我们正在分析Pomalidomide（12-C-0197; P.I. Steven Pavletic）试验中炎症基因表达的变化，以评估CGVHD患者纤维化的功效。 （4）最后，我们支持组织免疫组织化学分析和唾液细胞因子在一项含有有效类固醇clobetasol的漱口水试验中，作为严重口服CGVHD的局部疗法（12-C-0068; P.I. Steven Pavletic/Jacqueline Mays）。作为我们对CGVHD发病机理的研究的一部分，我们表征了CGVHD通过协调的血液和组织研究中的调节性T细胞。这些研究确定FOXP3+调节性T细胞（TREG）与口腔和皮肤地衣CGVHD中组织浸润的T效应子成比例增加。 T效应和FOXP3+ Treg细胞表达了TBET和趋化因子受体CXCR3，与趋化因子介导的迁移到组织的共同机制一致。此外，功能标记物（ICOS和CD39）和趋化因子受体（CXCR3）均与周围血液中的FOXP3+细胞比例更高，与外周血相比，与CGVHD目标组织中Treg的募集和激活一致。最后，尽管在CGVHD患者和正常对照组中发现了Treg细胞的“激活” CD45RA-FOXP3HI子集，该子群在可比的频率中发现了高度表达功能标记。这些发现与CGVHD中的Treg功能能力一致，并支持在体内扩展Treg细胞作为治疗的努力。 （Imanguli等，白血病，2014）在CGVHD发病机理的研究中，我们介绍了CGVHD患者循环单核细胞的基因表达，将这些细胞用作原位报道细胞进行全身细胞因子模式的原位报道细胞，以测试在诱导的（IFN）诱导的许多过程中，许多人可能会诱导的许多方法，这些过程可能会被诱导的许多eTic eTip ersic ersic ersim castie ersim castip ot themie ersim castip ot。 Al，2009年； Hakim，2010年）。根据微阵列分析，并通过多重RNA评估（由NCI员工科学家职业发展奖支持）确认，我们确定IFN诱导的途径始终在各种CGVHD患者中始终上调，这些患者均在组织中具有严重的炎性浸润性渗透率，并且具有广泛的螺旋菌丝参与。 IFN诱导的基因，包括特异性IFN诱导的基因，在CGVHD发作时并行上调，在治疗期间和解决CGVHD症状后降低。此外，我们发现来自先天免疫TLR/NLR/CLR途径的受体基因的一致上调。这些途径是由死细胞的碎屑引起的，以刺激吞噬作用，诱导IFNA产生并形成炎症。这些受体中的许多也可以通过IFN诱导，这与维持CGVHD的自我增强炎症过程一致。此外，通过评估糖皮质激素诱导的基因，我们区分了免疫抑制类固醇与疾病相关基因模式的混杂作用。最后，我们通过分析血浆和组织中的IFN诱导因子来证实基因表达数据。这些发现证实了IFN在淋巴细胞和树突状细胞中运输到组织中的作用，并通过上调BAFF，在CGVHD中涉及IFN在B细胞激活中的参与。这些互锁评估对严重影响CGVHD影响的广泛患者进行，支持新的CGVHD启动和持久性模型。该模型将CGVHD定义为炎症驱动的免疫失调的疾病，这些结果支持通过拦截干扰素途径来治疗CGVHD的新选择。