背景:大多数抗程序性细胞死亡1(PD-1)单克隆抗体(mAb)使用S228P突变IgG4作为结构基础,以避免免疫细胞或补体的激活。然而,IgG4和其他IgGFc片段之间的Fc-Fc相互作用可能导致不良反应的研究很少.Fc-无效IgG1框架是避免用IgG4观察到的不期望的Fc-Fc相互作用和Fc受体结合衍生效应的潜在更安全的替代方案。本研究提供了对这两种框架的抗PD-1mAb的综合评价。
方法:曲妥珠单抗和利妥昔单抗(均为IgG1),野生型IgG1和IgG4固定在硝酸纤维素膜上,涂在微孔板和生物传感器芯片上,并结合肿瘤细胞作为Fc-Fc相互作用的靶标。野生型IgG1和IgG4,抗PD-1单克隆抗体nivolumab(IgG4S228P),penpullimab(Fc-nullIgG1),评估和tislelizumab(Fc-nullIgG4S228P-R409K)与固定的IgG蛋白的结合反应,并获得定量动力学数据。评估两种抗PD-1单克隆抗体对联合治疗中曲妥珠单抗和利妥昔单抗介导的免疫反应的影响,我们采用经典的抗体依赖性细胞毒性免疫模型,抗体依赖性细胞吞噬作用,和补体依赖性细胞毒性。荷瘤小鼠模型,野生型和人源化,用于体内研究。此外,我们还检查了IgG1和IgG4对不同免疫细胞群体的影响。结果:实验证明野生型IgG4和nivolumab通过Fc-Fc相互作用与固定化IgG结合,减少抗体依赖性细胞介导的细胞毒性和吞噬反应。动力学参数的定量分析表明,纳武单抗和野生型IgG4在非变性和变性状态下都表现出与固定化IgG1相当的结合亲和力。IgG4对各种免疫细胞类型发挥抑制作用。野生型IgG4和nivolumab均促进野生型小鼠模型中的肿瘤生长。相反,野生型IgG1,penpulimab,和tislelizumab未显示类似的不良反应.
结论:通过避免IgG4的不利的Fc-Fc相互作用和Fc相关的免疫抑制作用,Fc-nullIgG1代表抗PD-1免疫疗法的更安全的选择。Fc-无效IgG4S228P-R409K和Fc-无效IgG1显示相似的结构特性和益处。这项研究有助于了解免疫疗法的耐药性和更安全的癌症免疫疗法的发展。
BACKGROUND: The majority of anti-programmed cell-death 1 (PD-1) monoclonal antibodies (mAbs) use S228P mutation IgG4 as the structural basis to avoid the activation of immune cells or complement. However, little attention has been paid to the Fc-Fc interactions between IgG4 and other IgG Fc fragments that could result in adverse effects. Fc-null IgG1 framework is a potential safer alternative to avoid the undesirable Fc-Fc interactions and Fc receptor binding derived effects observed with IgG4. This study provides a comprehensive evaluation of anti-PD-1 mAbs of these two frameworks.
METHODS: Trastuzumab and rituximab (both IgG1), wildtype IgG1 and IgG4 were immobilized on nitrocellulose membranes, coated to microplates and biosensor chips, and bound to tumor cells as targets for Fc-Fc interactions. Wildtype IgG1 and IgG4, anti-PD-1 mAb nivolumab (IgG4 S228P), penpulimab (Fc-null IgG1), and tislelizumab (Fc-null IgG4 S228P-R409K) were assessed for their binding reactions to the immobilized IgG proteins and quantitative kinetic data were obtained. To evaluate the effects of the two anti-PD-1 mAbs on immune responses mediated by trastuzumab and rituximab in the context of combination therapy, we employed classic immune models for
antibody-dependent cellular cytotoxicity,
antibody-dependent cellular phagocytosis, and complement dependent cytotoxicity. Tumor-bearing mouse models, both wildtype and humanized, were used for in vivo investigation. Furthermore, we also examined the effects of IgG1 and IgG4 on diverse immune cell populations RESULTS: Experiments demonstrated that wildtype IgG4 and nivolumab bound to immobilized IgG through Fc-Fc interactions, diminishing
antibody-dependent cell-mediated cytotoxicity and phagocytosis reactions. Quantitative analysis of kinetic parameters suggests that nivolumab and wildtype IgG4 exhibit comparable binding affinities to immobilized IgG1 in both non-denatured and denatured states. IgG4 exerted inhibitory effects on various immune cell types. Wildtype IgG4 and nivolumab both promoted tumor growth in wildtype mouse models. Conversely, wildtype IgG1, penpulimab, and tislelizumab did not show similar adverse effects.
CONCLUSIONS: Fc-null IgG1 represents a safer choice for anti-PD-1 immunotherapies by avoiding both the adverse Fc-Fc interactions and Fc-related immune inhibitory effects of IgG4. Fc-null IgG4 S228P-R409K and Fc-null IgG1 displayed similar structural properties and benefits. This study contributes to the understanding of immunotherapy resistance and the advancement of safer immune therapies for cancer.