Characterization of the Canine MHC Class I DLA-88*50101 Peptide Binding Motif as a Prerequisite for Canine T Cell Immunotherapy

There are limitations in pre-clinical settings using mice as a basis for clinical development in humans. In cancer, similarities exist between humans and dogs; thus, the dog patient can be a link in the transition from laboratory research on mouse models to clinical trials in humans. Knowledge of the peptides presented on MHC molecules is fundamental for the development of highly specific T cell-based immunotherapies. This information is available for human MHC molecules but is absent for the canine MHC. In the present study, we characterized the binding motif of dog leukocyte antigen (DLA) class I allele DLA-88*50101, using human C1R and K562 transfected cells expressing the DLA-88*50101 heavy chain. MHC class I immunoaffinity-purification revealed 3720 DLA-88*50101 derived peptides, which enabled the determination of major anchor positions. The characterized binding motif of DLA-88*50101 was similar to HLA-A*02:01. Peptide binding analyses on HLA-A*02:01 and DLA-88*50101 via flow cytometry showed weak binding of DLA-88*50101 derived peptides to HLA-A*02:01, and vice versa. Our results present for the first time a detailed peptide binding motif of the canine MHC class I allelic product DLA-88*50101. These data support the goal of establishing dogs as a suitable animal model for the evaluation and development of T cell-based cancer immunotherapies, benefiting both dog and human patients.

以小鼠作为人类临床开发的临床前研究模型存在局限性。在癌症领域，人类与犬类存在诸多相似性，因此患犬可作为衔接小鼠模型实验室研究与人类临床试验的桥梁。主要组织相容性复合体（Major Histocompatibility Complex, MHC）分子呈递的肽段相关知识，是开发高特异性基于T细胞的癌症免疫疗法的核心基础。此类信息在人类MHC分子中已有完善收录，但犬类MHC却尚无相关数据。本研究利用表达犬白细胞抗原（dog leukocyte antigen, DLA）I类等位基因DLA-88*50101重链的转染人C1R与K562细胞，对该等位基因的结合基序进行了表征。通过MHC I类免疫亲和纯化技术，我们共获取了3720个源自DLA-88*50101的肽段，借此确定了其主要锚定位点。所表征的DLA-88*50101结合基序与人类白细胞抗原（Human Leukocyte Antigen, HLA）-A*02:01高度相似。通过流式细胞术对HLA-A*02:01与DLA-88*50101开展肽段结合分析，结果显示源自DLA-88*50101的肽段与HLA-A*02:01仅存在弱结合，反之亦然。本研究首次报道了犬MHC I类等位基因产物DLA-88*50101的详细肽段结合基序。本研究数据支持将犬作为评估与开发基于T细胞的癌症免疫疗法的合适动物模型，此举将同时惠及患犬与人类患者。