Donor-specific B cells in Transplantation

摘要 肾移植激发了强大的B细胞反应，产生移植特异性抗体 供体B细胞对同种异体HLA（群体反应性抗体，PRA）和对移植供体（供体-受体）的应答。 特异性抗体（DSA）与排斥反应和移植物丢失的风险相关，因此可作为排斥反应的诊断指标。 同种免疫产生DSA的B细胞反应引发最令人烦恼的排斥类型（抗体- 介导的排斥反应，ABMR）和慢性ABMR，因此对治疗剂的开发存在很大兴趣 专门针对B细胞。然而，一些B细胞反应可能与移植物相关，甚至促进移植物的生长。 生存（免疫调节、调节、操作耐受）。虽然有广泛的 对血液DSA的特异性和功能的调查几乎不知道关于 供者特异性B细胞。我们提出的研究将首次确定 肾移植受者的供者特异性B细胞反应，以确定与哪些特性相关 与排斥反应相反，可能有助于稳定移植物功能。作为第一个目标， 将通过测定供体特异性B细胞的重链和轻链IG序列来研究供体特异性B细胞应答。 通过单细胞测序鉴定特异性B细胞，通过RNA-seq鉴定其表型特征。我们将确定关键 IG的特性，如克隆多样性、体细胞超突变的程度或IG独立特性，如 在功能稳定或排斥的受者中产生细胞因子。作为第二个目标，供体特异性IG NGS获得的序列将在>一年（最多两年）的时间段内进行前瞻性研究。我们将 确定克隆型是否在预期的排斥反应中扩增和多样化，如T细胞的预期， 依赖性反应;以及供体特异性序列在排斥治疗后如何变化。我们还将 确定在移植物稳定的受体中，供体特异性B细胞是否编码具有不同特性的Ig。作为第三 目标，我们将进行关键属性（例如最大结合，亲合力，直接 供体细胞的活化和抗原特异性，在可能的情况下，使用移植物组织确认） 单克隆DSA对供体细胞，以确定这些属性是否可以解释差异， 致病性这项研究可能会揭示评估风险的新方法和生物学的新目标。 干预这项研究也可能启发合理的方法来修改移植中的B细胞反应 目的是促进保护性反应和拮抗 致病的

Abstract Kidney transplantation elicits powerful B cell responses that generate antibodies specific for the transplant donor. B cell responses to allogeneic HLA (panel reactive antibodies, PRA) and to the transplant donor (donor- specific antibodies, DSA) correlate with risk of rejection and graft loss and thus serve as diagnostic indices of allo-immunity. B cell responses that generate DSA initiate the most vexing types of rejection (antibody- mediated rejection, ABMR) and chronic ABMR and hence much interest exists in development of therapeutics that specifically target B cells. Some B cell responses however may correlate with or even promote graft survival (immune regulation, accommodation, operational tolerance). Although there has been extensive investigation of the specificity and function of the blood DSA hardly anything is known about the properties of donor-specific B cells. The research we propose will identify for the first time key characteristics of donor-specific B cell responses in kidney transplant recipients to determine what properties associate with and potentially contribute to stable graft function, as opposed to rejection. As a first objective, donor-specific B cell responses will be studied by determining Heavy and Light chain Ig sequences of donor- specific B cells by single cell sequencing and their phenotypic signature by RNA-seq . We will determine key properties of Ig, such as clonal diversity, extent of somatic hypermutation or Ig independent properties such as production of cytokines in recipients with stable function or rejection. As a second objective, donor-specific Ig sequences obtained by NGS will be studied prospectively over periods > a year (up to two years). We will determine whether clonotypes expand and diversify in anticipation of rejection as would be expected of T cell dependent responses; and how donor-specific sequences change following rejection treatment. We will also determine if in recipients with stable grafts, donor-specific B cells encode Igs with distinct properties. As a third objective, we will conduct the first investigation of key properties (e.g. maximum binding, avidity, direct activation of donor cells and antigen specificity, where possible, confirmed using graft tissues) of recombinant monoclonal DSA on donor cells to determine whether these properties could explain differences in pathogenicity. The research might reveal novel approaches to assessing risk and new targets for biological intervention. The research might also inspire rational approaches to modifying B cell responses in transplant recipients in a personalized manner with the goal of promoting protective responses and antagonizing pathogenic ones.