Cardiovascular & Metabolism (Z.H.D., F.D., Q.L., M.B., M.C.) and Janssen Biotechnology (E.R.L.), Janssen Research & Development LLC, Spring House, Pennsylvania
Cardiovascular & Metabolism (Z.H.D., F.D., Q.L., M.B., M.C.) and Janssen Biotechnology (E.R.L.), Janssen Research & Development LLC, Spring House, Pennsylvania.
J Pharmacol Exp Ther. 2019 Nov;371(2):375-384. doi: 10.1124/jpet.119.261032. Epub 2019 Aug 26.
JNJ-64179375 (JNJ-9375) is a recombinant human IgG4 monoclonal antibody engineered to mimic an IgA antibody that was identified in a patient who exhibited markedly prolonged clotting times but without spontaneous bleeding episodes over several years of follow-up. The crystal structure of the JNJ-9375 antigen-binding fragment/thrombin complex showed an almost identical binding mode to that of the patient IgA. In the current study, we characterized the in vitro and in vivo properties of JNJ-9375. Surface plasmon resonance studies demonstrated that JNJ-9375 binds to -thrombin with high affinity and specificity ( : 0.8 nM for human thrombin). JNJ-9375 produced concentration-dependent prolongation of in vitro clotting assays in human plasma, including thrombin time (TT), ecarin clotting time, prothrombin time, and activated partial thromboplastin time, with EC values of 4.4, 12.4, 172.6, and 202.7 g/ml, respectively. JNJ-9375 inhibited thrombin-induced platelet aggregation in human plasma with an IC value of 52.6 nM (7.8 g/ml) and produced concentration-dependent prolongation of reaction time tested by thromboelastography. JNJ-9375 pretreatment resulted in dose-dependent reduction in thrombus formation in the rat arteriovenous (AV) shunt model of thrombosis. Robust efficacy was observed at 0.3 mg/kg accompanied by 1.5× of TT. Bleeding was increased at 3 mg/kg in a rat tail transection bleeding model demonstrating a therapeutic index of 10× compared with 1× for apixaban in the same models. Our data suggest that thrombin exosite I inhibition is efficacious against thrombosis in a pretreatment prevention animal model. SIGNIFICANCE STATEMENT: JNJ-9375 is a novel, fully human monoclonal antibody that binds to the exosite I region of thrombin with high affinity and specificity. JNJ-9375 concentration dependently prolonged clotting times and inhibited thrombin-induced platelet aggregation in in vitro assays based on its mechanism of action. In an in vivo rat AV shunt model, JNJ-9375 prevented thrombus formation in a dose-dependent fashion while demonstrating reduced bleeding risk. The present study demonstrated the antithrombotic effects of inhibiting the exosite I region of thrombin when given in a prevention mode in preclinical animal models.
JNJ-64179375（JNJ-9375）是一种重组人 IgG4 单克隆抗体，经过工程改造后可模拟在一名患者体内发现的 IgA 抗体，该患者的凝血时间明显延长，但在几年的随访中并未出现自发性出血事件。JNJ-9375 抗原结合片段/凝血酶复合物的晶体结构显示出与患者 IgA 几乎相同的结合模式。在本研究中，我们对 JNJ-9375 的体外和体内特性进行了表征。表面等离子体共振研究表明，JNJ-9375 与人凝血酶具有高亲和力和特异性的结合（对人凝血酶的 Kd 值为 0.8 nM）。JNJ-9375 在人血浆中可产生浓度依赖性的体外凝血试验延长，包括凝血酶时间（TT）、蝰蛇凝血酶时间、凝血酶原时间和活化部分凝血活酶时间，EC 值分别为 4.4、12.4、172.6 和 202.7 g/ml。JNJ-9375 以 52.6 nM（7.8 g/ml）的 IC 值抑制凝血酶诱导的人血浆血小板聚集，并通过血栓弹性描记术测试产生浓度依赖性的反应时间延长。JNJ-9375 预处理可导致大鼠动静脉（AV）分流模型中血栓形成的剂量依赖性减少。在 0.3 mg/kg 时观察到强大的疗效，同时 TT 延长 1.5 倍。在大鼠尾切断出血模型中，3 mg/kg 时出血增加，与同一模型中的阿哌沙班相比，治疗指数为 10×，而不是 1×。我们的数据表明，在预处理预防动物模型中，凝血酶外切酶 I 抑制对血栓形成有效。
JNJ-9375 是一种新型的、完全人源的单克隆抗体，它与人凝血酶的外切酶 I 区域具有高亲和力和特异性结合。根据其作用机制，JNJ-9375 在体外测定中浓度依赖性地延长凝血时间并抑制凝血酶诱导的血小板聚集。在大鼠 AV 分流模型中，JNJ-9375 以剂量依赖性方式预防血栓形成，同时降低出血风险。本研究在临床前动物模型中以预防模式给药时，证明了抑制凝血酶外切酶 I 区域的抗血栓形成作用。
