An antibody mimetic known as Fab–PEG–Fab (FpF) is a stable bivalent molecule that may have some potential therapeutic advantages over IgG antibodies due to differences in their binding kinetics as determined by surface plasmon resonance. Here we describe the thermodynamic binding properties to vascular endothelial growth factor (VEGF) of the FpF antibody mimetics derived from bevacizumab and ranibizumab. Bevacizumab is an IgG antibody and ranibizumab is an antibody fragment (Fab). Both are used clinically to target VEGF to inhibit angiogenesis. FpF_beva displayed comparable binding affinity (KD) and binding thermodynamics (ΔH = −25.7 kcal mole⁻¹ and ΔS = 14 kcal mole⁻¹) to bevacizumab (ΔH = −25 kcal mole⁻¹, ΔS = 13.3 kcal mole⁻¹). FpF_rani interactions with VEGF were characterised by large favourable enthalpy (ΔH = −42 kcal mole⁻¹) and unfavourable entropy (ΔS = 31 kcal mole⁻¹) changes compared to ranibizumab (ΔH = −18.5 kcal mole⁻¹ and ΔS = 6.7 kcal mole⁻¹), which being a Fab, is mono-valent. A large negative entropy change resulting in binding of bivalent FpF to homodimer VEGF might be due to the conformational change of the flexible regions of the FpF upon ligand binding. Mono-valent Fab (i.e. ranibizumab or the Fab derived from bevacizumab) displayed a larger degree of freedom (smaller unfavourable entropy) upon binding to homodimer VEGF. Our report describes the first comprehensive enthalpy and entropy compensation analysis for FpF antibody mimetics. While the FpFs displayed similar thermodynamics and binding affinity to the full IgG (i.e. bevacizumab), their enhanced protein stability, slower dissociation rate and lack of Fc effector functions could make FpF a potential next-generation therapy for local tissue-targeted indications.

中文翻译：

---

下一代 VEGF 抗体模拟物溶液中的比较热力学分析

称为 Fab-PEG-Fab (FpF) 的抗体模拟物是一种稳定的二价分子，由于表面等离子共振确定的结合动力学差异，可能比 IgG 抗体具有一些潜在的治疗优势。在这里，我们描述了源自贝伐单抗和雷珠单抗的 FpF 抗体模拟物与血管内皮生长因子 (VEGF) 的热力学结合特性。贝伐单抗是一种 IgG 抗体，雷珠单抗是一种抗体片段 (Fab)。两者都在临床上用于靶向 VEGF 以抑制血管生成。FpF _beva与贝^伐单抗( Δ H _ ^_ _= -25 kcal 摩尔^-1，Δ S = 13.3 kcal 摩尔^-1 )。与^雷珠单抗（ΔH = -18.5 _kcal摩尔^-1和_ ^_ _ Δ S = 6.7 kcal 摩尔^-1 )，它是一种Fab，是一价的。导致二价 FpF 与同源二聚体 VEGF 结合的大的负熵变化可能是由于配体结合后 FpF 的柔性区域的构象变化。单价Fab（即雷珠单抗或源自贝伐单抗的 Fab）在与同型二聚体 VEGF 结合时表现出更大的自由度（更小的不利熵）。我们的报告描述了 FpF 抗体模拟物的第一个综合焓和熵补偿分析。虽然 FpF 显示出与完整 IgG（即贝伐单抗）相似的热力学和结合亲和力，但它们增强的蛋白质稳定性、较慢的解离速率和缺乏 Fc 效应功能可能使 FpF 成为局部组织靶向适应症的潜在下一代疗法。