Identification of Canine Minor Histocompatibility Antigens

犬次要组织相容性抗原的鉴定

• 批准号: 8240004
• 负责人: Jay Ashok Shendure
• 金额: $ 31.28万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-08 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8240004
• 关键词: Acute leukemia; Address; Age; Alleles; Allogenic; Allografting; Animals; Antigens; Biological Assay; Breeding; Canis familiaris; Cell Transplantation; 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; high risk; high throughput screening; improved; in vivo; leukemia; mycophenolate mofetil; next generation; novel; novel strategies; peripheral blood; pre-clinical; prevent; pup; response; tumor

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已经开发了一种DLA相同的犬造血细胞移植（HCT）方法， 导致稳定的混合供体-宿主嵌合体。持续的宿主造血可以作为实验性的 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。 混合嵌合体向全供者嵌合体的转化可以通过注射具有以下特征的供者淋巴细胞来实现： 对外周血单核细胞上表达的宿主次要组织相容性抗原敏感 （PBMC），然而，代价往往是致命的移植物对宿主病（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细胞应答 针对普遍存在的和造血特异性次要抗原的组合，有助于消除 残留的宿主造血细胞以类似于移植物对在人体中观察到的白血病效应 患者因此，鉴定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结果。这一知识将有助于提高认识， 靶向宿主造血细胞同时保留GVHD靶组织的策略。然而，当狗 同种异体HCT模型是新型HCT疗法临床前开发的最佳选择，没有犬次要， 迄今为止，已经描述了组织相容性抗原， 识别是低效。为了解决这个问题，我们提出了一种新的方法， 狗的发现这种方法利用下一代测序技术来定义蛋白质编码 受者特有的并在PBMC中表达的变异，其将用于敏化HCT供体。 然后将致敏的供体T细胞注射到相应的受体中，目的是将混合的T细胞转化为受体。 完全供者嵌合体导致移植物抗宿主病转换完成后，T细胞将被 从接受者中收获，并使用一种新的、高- 通量T细胞测定。阳性反应将确定真正的次要组织相容性抗原。使用 qRT-PCR，然后我们将识别那些在造血细胞中高度表达但在造血细胞中不表达的次要抗原 在GVHD靶组织中。接下来，将使用相关的次要抗原肽来使供体T细胞与 目的是将混合嵌合体转化为完全供体嵌合体，而不发生GVHD（项目1）。最终，这一概念将成为 在项目1的急性白血病犬模型中进行了测试。对公共卫生的好处：综合起来， 本项目中提出的研究和项目1中提出的体内研究有可能 开发新的有效方法，使接受治疗的持续/复发性恶性肿瘤患者受益 在项目3和项目4下通过同种异体HCT。