pH-switchable shark-derived single domain antibodies for biomedical applications

几十年来，抗体一直是包括诊断和治疗领域在内的各种应用中不可替代的分子。同源抗原的高特异性与fc介导的免疫效应功能相结合，使抗体成为医疗应用的吉祥工具。除了具有经典的重链和轻链组成的抗体外，鲨鱼的适应性免疫库还包括仅具有重链的IgNAR同型，其中抗原结合完全由称为vNAR的小而高度稳定的结构域介导。近年来，vNAR片段已发展成为一种有前途的靶向结合支架，可用于医学和生物技术。这些结构域包括高亲和力和特异性、小尺寸、高物理化学稳定性和低生产成本等有益特性。在拟议的项目中，我们着手生成和利用ph响应的vNAR域用于生物治疗应用。这种ph可切换的vNAR结构域可能对酸性肿瘤微环境中抗原的特异性靶向具有吸引力。因此，在酸性pH下对肿瘤相关抗原具有较高亲和力，但在生理条件下具有明显较低亲和力的变体可以减少常规抗体治疗中经常观察到的脱靶效应。此外，将开发ph依赖性和抗独特型vNAR变体，旨在提高现有治疗性抗体的靶向疗效。这些掩蔽vnas在生理pH值下对抗体具有高亲和力，但在微酸性条件下亲和力较低。这种作用会在健康组织中阻断治疗性抗体，而在到达酸性肿瘤微环境后被揭开。

For decades, antibodies have been irreplaceable molecules for various applications encompassing the field of diagnostics as well as therapy. The high specificity for a cognate antigen combined with Fc-mediated immune effector functions renders antibodies auspicious tools for medical applications. In addition to antibodies with the classical composition of heavy and light chains, the adaptive immune repertoire of sharks also includes a heavy-chain only isotype named IgNAR, where antigen binding is mediated exclusively by a small and highly stable domain referred to as vNAR. In recent years, vNAR fragments have evolved as promising, target-binding scaffolds that can be tailored for applications in medicine and biotechnology. These domains comprise beneficial properties such as high affinity and specificity, small size, high physicochemical stability and a low cost of production. In the proposed project we set out to generate and utilize pH-responsive vNAR domains for biotherapeutic applications. Such pH-switchable vNAR domains might be attractive for the specific targeting of antigens in the acidic tumor microenvironment. As such, variants comprising higher affinity for a tumor-associated antigen at acidic pH but significantly lower affinity under physiological conditions could reduce off-target effects which are often observed for conventional antibody therapeutics. In addition, pH-dependent and anti-idiotypic vNAR variants will be developed aimed at increasing the on-target efficacy of already available therapeutic antibodies. These masking vNARs comprise high affinity for the antibody at physiological pH, but low affinity under slightly acidic conditions. This effect would block the therapeutic antibody in healthy tissue, while being unmasked upon reaching the acidic tumor microenvironment.