Development of tibulizumab, a tetravalent bispecific antibody targeting BAFF and IL-17A for the treatment of autoimmune disease
Development of tibulizumab, a tetravalent bispecific antibody targeting BAFF and IL-17A for the treatment of autoimmune disease
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DOI：
10.1080/19420862.2019.1624463
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发表时间：
2019-06-12
期刊：
影响因子：
5.3
通讯作者：
Allan, Barrett W.
中科院分区：
文献类型：
--
作者：
Benschop, Robert J.;Chow, Chi-Kin;Allan, Barrett W.
我们描述了一种针对人B细胞活化因子（BAFF）和白细胞介素17A （IL-17）的双特异性双拮抗剂抗体。来自ixekizumab （Taltz (R)）的抗il -17单链可变片段（scFv）通过富含甘氨酸的连接物与抗baff的tabalumab融合。IgG-scFv同时结合BAFF和IL-17，其化学计量与亲本单克隆抗体相同。对初始IgG-scFv的稳定性研究显示，在两种亲本抗体中均未观察到化学降解和聚集。差示扫描量热法显示，抗il -17 scFv具有较高的熔融温度（Tm）（73.1℃），但也具有浓度依赖性，初步可逆，蛋白质自结合。为了设计scFv的稳定性，采用了三种平行的方法：用稳定的、亲和中性的氨基酸取代不稳定的互补决定区（CDR）残基，平衡CDR电荷分布，引入H44-L100界面二硫键。二硫化物稳定的scFv的Tm基本未受干扰，但在高蛋白质浓度下仍保持单分散。荧光染料结合滴定表明，疏水残基的溶剂暴露减少，蛋白质水解敏感性降低，两者都表明构象稳定性增强。H44-L100 scFv （PDB id: 6NOU）和ixekizumab抗原结合片段（PDB id: 6NOV）晶体结构的叠加显示框架和CDR环的取向几乎相同。稳定的双特异性分子LY3090106 （tibulizumab）在细胞和体内小鼠模型中均能有效拮抗BAFF和IL-17。在食蟹猴中，它抑制B细胞的发育和存活，并在循环中保持功能完整，半衰期延长。总之，我们设计了一种有效的双特异性抗体，靶向参与人类自身免疫的两种关键细胞因子，适合临床发展。
We describe a bispecific dual-antagonist antibody against human B cell activating factor (BAFF) and interleukin 17A (IL-17). An anti-IL-17 single-chain variable fragment (scFv) derived from ixekizumab (Taltz (R)) was fused via a glycine-rich linker to anti-BAFF tabalumab. The IgG-scFv bound both BAFF and IL-17 simultaneously with identical stoichiometry as the parental mAbs. Stability studies of the initial IgG-scFv revealed chemical degradation and aggregation not observed in either parental antibody. The anti-IL-17 scFv showed a high melting temperature (Tm) by differential scanning calorimetry (73.1 degrees C), but also concentration-dependent, initially reversible, protein self-association. To engineer scFv stability, three parallel approaches were taken: labile complementary-determining region (CDR) residues were replaced by stable, affinity-neutral amino acids, CDR charge distribution was balanced, and a H44-L100 interface disulfide bond was introduced. The Tm of the disulfide-stabilized scFv was largely unperturbed, yet it remained monodispersed at high protein concentration. Fluorescent dye binding titrations indicated reduced solvent exposure of hydrophobic residues and decreased proteolytic susceptibility was observed, both indicative of enhanced conformational stability. Superimposition of the H44-L100 scFv (PDB id: 6NOU) and ixekizumab antigen-binding fragment (PDB id: 6NOV) crystal structures revealed nearly identical orientation of the frameworks and CDR loops. The stabilized bispecific molecule LY3090106 (tibulizumab) potently antagonized both BAFF and IL-17 in cell-based and in vivo mouse models. In cynomolgus monkey, it suppressed B cell development and survival and remained functionally intact in circulation, with a prolonged half-life. In summary, we engineered a potent bispecific antibody targeting two key cytokines involved in human autoimmunity amenable to clinical development.