Pai Rama, Ma Ning, Connor Anu V, Danilenko Dimitry M, Tarrant Jacqueline M, Salvail Dany, Wong Lisa, Hartley Dylan P, Misner Dinah, Stefanich Eric, Wu Yan, Chen Yongmei, Wang Hong, Dambach Donna M
*Safety Assessment;
IPS Therapeutique, Sherbrooke, Quebec, Canada.
Toxicol Sci. 2016 Jun;151(2):245-60. doi: 10.1093/toxsci/kfw037. Epub 2016 Feb 25.
PRO304186, a humanized monoclonal antibody targeting soluble interleukin-17 A and F, was developed for autoimmune and inflammatory disease indications. When administered to cynomolgus monkeys PRO304186 induced unexpected adverse effects characterized by clinical signs of hematemesis, hematochezia, and moribundity. Pathology findings included hemorrhage throughout the gastrointestinal tract without any evidence of vascular wall damage or inflammatory cellular infiltration. Mechanistic investigation of these effects revealed mild elevations of serum MCP-1 and IL-12/23 but without a classical proinflammatory profile in PRO304186-treated animals. In vitro studies demonstrated off-target effects on vascular endothelial cells including activation of nitric oxide synthase leading to production of nitric oxide (NO) accompanied by increased mitochondrial membrane depolarization, glutathione depletion, and increased paracellular permeability. Additionally, endothelial cell-PRO304186-conditioned medium reduced myosin light chain phosphorylation in vascular smooth muscle cells. Furthermore, an ex vivo study utilizing segments from cynomolgus aorta and femoral artery confirmed PRO304186-induced endothelium-dependent smooth muscle relaxation and vasodilation mediated via NO. Finally, a single dose of PRO304186 in cynomolgus monkeys induced a rapid and pronounced increase in NO in the portal circulation that preceded a milder elevation of NO in the systemic circulation and corresponded temporally with systemic hypotension; findings consistent with NO-mediated vasodilation leading to hypotension. These changes were associated with non-inflammatory, localized hemorrhage in the gastrointestinal tract consistent with hemodynamic vascular injury associated with intense local vasodilation. Together, these data demonstrate that PRO304186-associated toxicity in monkeys was due to an off-target effect on endothelium that involved regional NO release resulting in severe systemic vasodilation, hypotension, and hemorrhage.
PRO304186是一种靶向可溶性白细胞介素-17A和F的人源化单克隆抗体，用于自身免疫性和炎性疾病适应症。给食蟹猴给药后，PRO304186引发了意外的不良反应，其特征为呕血、便血和濒死的临床症状。病理检查结果包括全胃肠道出血，未发现血管壁损伤或炎性细胞浸润的证据。对这些效应的机制研究显示，在接受PRO304186治疗的动物中，血清MCP-1和IL-12/23轻度升高，但没有典型的促炎特征。体外研究表明，PRO304186对血管内皮细胞有脱靶效应，包括激活一氧化氮合酶导致一氧化氮（NO）生成，同时伴有线粒体膜去极化增加、谷胱甘肽耗竭和细胞旁通透性增加。此外，内皮细胞-PRO304186条件培养基可降低血管平滑肌细胞中的肌球蛋白轻链磷酸化。此外，一项利用食蟹猴主动脉和股动脉节段进行的体外研究证实，PRO304186可诱导内皮依赖性平滑肌舒张和通过NO介导的血管舒张。最后，给食蟹猴单次注射PRO304186可导致门静脉循环中NO迅速显著增加，随后全身循环中NO轻度升高，并在时间上与全身低血压相对应；这些发现与NO介导的血管舒张导致低血压一致。这些变化与胃肠道非炎性局部出血有关，这与强烈局部血管舒张相关的血流动力学血管损伤一致。总之，这些数据表明，PRO304186在猴子身上的毒性是由于对内皮的脱靶效应，该效应涉及局部NO释放，导致严重的全身血管舒张、低血压和出血。
