Center of Excellence for Companion Animal Cancer, Department of Veterinary Pathology, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, Thailand.
Center of Excellence in Systems Biology, Faculty of Medicine, Chulalongkorn University, Bangkok, Thailand.
Vet Q. 2023 Dec;43(1):1-9. doi: 10.1080/01652176.2023.2240380.
Immune escape is the hallmark of carcinogenesis. This widely known mechanism is the overexpression of immune checkpoint ligands, such as programmed cell death protein 1 and programmed death-ligand 1 (PD-1/PD-L1), leading to T cell anergy. Therefore, cancer immunotherapy with specific binding to these receptors has been developed to treat human cancers. Due to the lack of cross-reactivity of these antibodies in dogs, a specific canine PD-1/PD-L1 antibody is required. The aim of this study is to develop mouse anti-canine PD-L1 (cPD-L1) monoclonal antibodies and characterize their properties. Six mice were immunized with recombinant cPD-L1 with a fusion of human Fc tag. The hybridoma clones that successfully generated anti-cPD-L1 antibodies and had neutralizing activity were selected for monoclonal antibody production. Antibody properties were tested by immunosorbent assay, surface plasmon resonance, and immunohistochemistry. Four hybridomas were effectively bound and blocked to recombinant cPD-L1 and cPD-1-His-protein, respectively. Candidate mouse monoclonal antibodies worked efficiently on formalin-fixed paraffin-embedded tissues of canine cancers, including cutaneous T-cell lymphomas, mammary carcinomas, soft tissue sarcomas, squamous cell carcinomas, and malignant melanomas. However, functional assays of these anti-cPD-L1 antibodies need further investigation to prove their abilities as therapeutic drugs in dogs as well as their applications as prognostic markers.
免疫逃逸是癌症发生的标志。这种广泛知晓的机制是免疫检查点配体的过度表达，如程序性细胞死亡蛋白 1 和程序性死亡配体 1（PD-1/PD-L1），导致 T 细胞失能。因此，开发了针对这些受体的特异性结合的癌症免疫疗法来治疗人类癌症。由于这些抗体在狗中缺乏交叉反应性，因此需要一种特异性的犬 PD-1/PD-L1 抗体。本研究旨在开发抗犬 PD-L1（cPD-L1）的单克隆抗体并对其特性进行鉴定。用融合了人 Fc 标签的重组 cPD-L1 免疫了 6 只小鼠。选择具有中和活性并成功产生抗 cPD-L1 抗体的杂交瘤克隆进行单克隆抗体生产。通过免疫吸附测定、表面等离子体共振和免疫组织化学来检测抗体特性。有 4 个杂交瘤能有效结合并阻断重组 cPD-L1 和 cPD-1-His 蛋白。候选的鼠单克隆抗体能有效作用于犬癌症的福尔马林固定石蜡包埋组织，包括皮肤 T 细胞淋巴瘤、乳腺肿瘤、软组织肉瘤、鳞状细胞癌和恶性黑色素瘤。然而，这些抗 cPD-L1 抗体的功能检测还需要进一步研究，以证明它们在犬中作为治疗药物的能力以及作为预后标志物的应用。
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