Identification of Canine Minor Histocompatibility Antigens

犬次要组织相容性抗原的鉴定

基本信息

批准号：
8459332
负责人：
Jay Ashok Shendure
金额：
$ 30.64万
依托单位：
FRED HUTCHINSON CANCER RESEARCH CENTER
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至 2014-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8459332
关键词：
Acute leukemia Address Age Alleles Allogenic Allografting Animals Antigens Biological Assay Breeding Canis familiaris Cells Chimerism Clinic Code Collaborations Comorbidity Custom Cyclosporine Data Development Disease Donor Lymphocyte Infusion Experimental Models Functional RNA Genes Genetic Genetic Polymorphism Genetic Variation Genomics Genotype Goals Harvest Hematologic Neoplasms Hematopoiesis Hematopoietic Histocompatibility Antigens Human Immunization Immunosuppression Immunosuppressive Agents Injection of therapeutic agent Knowledge Lymphocyte Malignant Neoplasms Methodology Minor Minor Histocompatibility Antigens Minority Modeling Molecular Outcome Patients Peptides Peripheral Blood Mononuclear Cell Pharmaceutical Preparations Price Procedures Proteins Public Health Reaction Regimen Relapse Residual state Reverse Transcriptase Polymerase Chain Reaction Risk Role Shotgun Sequencing Single Nucleotide Polymorphism Stem cells T cell response T-Lymphocyte Technology Testing Tissues Translating Transplant Recipients Transplantation Variant base conditioning cost design experience graft vs host disease hematopoietic cell transplantation high risk high throughput screening improved in vivo leukemia mycophenolate mofetil next generation sequencing novel novel strategies peripheral blood pre-clinical prevent pup response tumor

项目摘要

PROJECT 2: IDENTIFICATION OFCANINE MINOR HISTOCOMPATIBILITY ANTIGENS Project 1 has developed an approach at DLA-identical canine hematopoietic cell transplantation (HCT) that results in stable mixed donor-host chimerism. Persistent host hematopoiesis can serve as an experimental model of persistent hematologic malignancy seen in some patients transplanted under Projects 3 and 4. Conversion of mixed to all-donor chimerism can be achieved with injection of donor lymphocytes that have been sensitized to host minor histocompatibility antigens expressed on peripheral blood mononuclear cells (PBMC), however at the price of often fatal graft-vs.-host disease (GVHD). T-cell responses directed against ubiquitously expressed minor antigens are thought to be responsible for GVHD, while T-cell responses against a combination of ubiquitous and hematopoietic-specific minor antigens contribute to elimination of residual host hematopoietic cells in a manner analogous to the graft-vs.-leukemia effect observed in human patients. The identification of minor antigens restricted to hematopoietic cells therefore holds great promise for improving allogeneic HCT outcomes. That knowledge would facilitate the development of sensitization strategies that target host hematopoietic cells while sparing GVHD target tissues. However, while the dog model of allogeneic HCT is optimal for preclinical development of novel HCT therapies, no canine minor, histocompatibility antigens have been described to date, and existing methodologies for minor antigen identification are inefficient. To address this problem, we propose a novel approach to minor antigen discovery in the dog. This approach utilizes next generation sequencing technology to define protein coding variations unique to the recipient and expressed in PBMC which will be used for sensitizing the HCT donor. Sensitized donor T-cells will then be injected into the respective recipients with the aim of converting mixed to full-donor chimerism and causing GVHD. After conversion has been accomplished, T-cells will be harvested from recipients and tested for responses against candidate minor antigens using a novel, high- throughput T-cell assay. Positive responses would define genuine minor histocompatibility antigens. Using qRT-PCR, we will then identify those minor antigens that are highly expressed in hematopoietic cells but not in GVHD target tissues. Next, relevant minor antigen peptides will be used to sensitize donor T-cells with the aim of converting mixed to full-donor chimerism without GVHD (Project 1). Eventually, this concept will be tested in a canine model of acute leukemia in Project 1. Benefits to Public Health: Taken together, the studies proposed in this Project and the in vivo studies proposed in Project 1 have the potential of developing new and effective approaches benefiting patients with persisting/relapsing malignancies treated by allogeneic HCT under Projects 3 and 4.

项目2：犬次要组织相容性抗原的鉴定项目 1 开发了一种与 DLA 相同的犬造血细胞移植 (HCT) 方法导致稳定的混合供体-宿主嵌合。持续宿主造血可以作为实验在项目 3 和 4 下移植的一些患者中观察到的持续性血液恶性肿瘤模型。混合嵌合体向全供体嵌合体的转化可以通过注射具有对外周血单核细胞表达的宿主次要组织相容性抗原敏感（PBMC），然而，其代价往往是致命的移植物抗宿主病（GVHD）。 T细胞反应针对普遍表达的次要抗原被认为是 GVHD 的原因，而 T 细胞反应对抗普遍存在的造血特异性次要抗原的组合有助于消除以类似于在人类中观察到的移植物抗白血病效应的方式残留宿主造血细胞患者。因此，鉴定仅限于造血细胞的次要抗原具有很大的前景用于改善同种异体 HCT 的结果。这些知识将有助于提高认识靶向宿主造血细胞同时不伤害 GVHD 靶组织的策略。然而，虽然狗同种异体 HCT 模型是新型 HCT 疗法临床前开发的最佳选择，无犬类未成年人、迄今为止，已经描述了组织相容性抗原，并且现有的次要抗原方法学识别效率低下。为了解决这个问题，我们提出了一种针对次要抗原的新方法在狗身上的发现。该方法利用下一代测序技术来定义蛋白质编码受者特有的变异并在 PBMC 中表达，这些变异将用于使 HCT 捐赠者敏感。然后，致敏的供体 T 细胞将被注射到各自的受体中，目的是转化混合的 T 细胞。完全供体嵌合并引起 GVHD。转化完成后，T细胞将被从接受者身上收获并使用一种新型的高抗原测试针对候选次要抗原的反应吞吐量T细胞测定。阳性反应将定义真正的次要组织相容性抗原。使用 qRT-PCR，然后我们将鉴定那些在造血细胞中高表达但不表达的次要抗原 GVHD 靶组织中。接下来，相关的次要抗原肽将用于使供体 T 细胞敏感目标是在没有 GVHD 的情况下将混合嵌合转化为完全供体嵌合（项目 1）。最终，这个概念将被在项目 1 中的急性白血病犬模型中进行了测试。对公共健康的益处：总而言之，本项目中提出的研究和项目 1 中提出的体内研究具有以下潜力：开发新的有效方法，使接受治疗的持续性/复发性恶性肿瘤患者受益通过项目 3 和 4 下的同种异体 HCT。