Single Cycle Structure-Based Humanization of an Anti-Nerve Growth Factor Therapeutic Antibody

Most forms of chronic pain are inadequately treated by present therapeutic options. Compelling evidence has accumulated, demonstrating that Nerve Growth Factor (NGF) is a key modulator of inflammatory and nociceptive responses, and is a promising target for the treatment of human pathologies linked to chronic and inflammatory pain. There is therefore a growing interest in the development of therapeutic molecules antagonising the NGF pathway and its nociceptor sensitization actions, among which function-blocking anti-NGF antibodies are particularly relevant candidates. In this respect, the rat anti-NGF αD11 monoclonal antibody (mAb) is a potent antagonist, able to effectively antagonize rodent and human NGF in a variety of in vitro and in vivo systems. Here we show that mAb αD11 displays a significant analgesic effect in two different models of persistent pain in mice, with a remarkable long-lasting activity. In order to advance αD11 mAb towards its clinical application in man, anti-NGF αD11 mAb was humanized by applying a novel single cycle strategy based on the a priori experimental determination of the crystal and molecular structure of the parental Fragment antigen-binding (Fab). The humanized antibody (hum-αD11) was tested in vitro and in vivo, showing that the binding mode and the NGF neutralizing biological activities of the parental antibody are fully preserved, with even a significant affinity improvement. The results firmly establish hum-αD11 as a lead candidate for clinical applications in a therapeutic area with a severe unmet medical need. More generally, the single-cycle structure-based humanization method represents a considerable improvement over the standard humanization methods, which are intrinsically empirical and require several refinement cycles.当前治疗手段难以充分应对绝大多数慢性疼痛病症。已有大量确凿证据表明，神经生长因子（Nerve Growth Factor，NGF）是炎症与伤害感受反应的关键调节因子，也是治疗慢性疼痛及炎症相关性疼痛相关人类疾病的极具潜力的靶点。因此，学界对开发能够拮抗NGF通路及其伤害感受器敏化作用的治疗性分子的兴趣与日俱增，其中功能阻断型抗NGF抗体便是极具相关性的候选药物之一。就此而言，大鼠抗NGF αD11单克隆抗体（monoclonal antibody，mAb）是一种强效拮抗剂，可在多种体外（in vitro）及体内（in vivo）实验体系中有效拮抗啮齿类与人类的NGF。本研究证实，αD11单克隆抗体在两种不同的小鼠持续性疼痛模型中展现出显著的镇痛效果，且镇痛活性持久不衰。为推动αD11单克隆抗体应用于人类临床治疗，研究团队基于亲本抗原结合片段（Fragment antigen-binding，Fab）的晶体结构与分子结构的先验实验测定结果，采用一种全新的单循环策略对该抗NGF αD11单克隆抗体进行了人源化改造。研究人员对该人源化抗体（hum-αD11）开展了体外（in vitro）与体内（in vivo）实验，结果显示亲本抗体的结合模式与NGF中和生物学活性均得到完整保留，甚至还实现了亲和力的显著提升。本研究结果明确证实，hum-αD11可作为一款先导候选药物，用于存在严重未满足医疗需求的治疗领域的临床应用。更广泛而言，这种基于结构的单循环人源化方法相较于传统的经验性、需多轮优化的标准人源化方法，实现了显著改进。