Fielden Mark R, Hassani Mary, Uppal Hirdesh, Day-Lollini Patricia, Button Donald, Martin Renee S, Garrido Rosario, Liu Xingrong, Kolaja Kyle L
Department of Nonclinical Safety, Roche Palo Alto LLC, Palo Alto, CA 94051, USA.
Exp Toxicol Pathol. 2010 Nov;62(6):607-13. doi: 10.1016/j.etp.2009.08.009. Epub 2009 Sep 24.
A number of drugs and drug candidates, including fenfluramine and ergot derivatives, are associated with valvulopathy in humans; however, these responses are poorly predicted from animal studies. In vitro and in vivo evidence suggests that these compounds exert their pathological effect through activation of serotonin 2B receptor (5HT2BR) signaling. However, the variable effect of fenfluramine and other 5HT2BR agonists in rodents has cast doubt on the relevance of animal findings to predicting human risk. Herein, a candidate compound, RO3013, induced subendocardial cell proliferation in the mitral and tricuspid valves in rats after only 3 days of daily dosing. Additionally, there was a treatment-related increase in immunostaining of the proliferation marker Ki67, and phosphorylated Smad3 in the heart indicative of TGFβ signaling co-localized with 5HT2BR expression. To substantiate the hypothesis that RO3013-induced valvular proliferation is secondary to 5HT2BR activation, the compound was evaluated in vitro and found to bind to the human 5HT2BR with a K(i) of 3.8μM; however, it was virtually devoid of agonist activity in a functional assay in human cells. By contrast, RO3013 bound to the rat 5HT2BR with a K(i) of 1.2μM and activated the receptor with an EC50 of 0.5μM. This agonist potency estimate is in good agreement with the free plasma concentrations of RO3013 at which valvular proliferation was observed. These results suggest that the rat may be susceptible to 5HT2BR-mediated valvular proliferation similar to humans; yet, the significant differences between binding and functional activities can be a possible explanation for the observed species-selective receptor responses.
包括芬氟拉明和麦角衍生物在内的多种药物及候选药物与人类瓣膜病有关；然而，动物研究对这些反应的预测效果不佳。体外和体内证据表明，这些化合物通过激活血清素2B受体（5HT2BR）信号传导发挥其病理作用。然而，芬氟拉明和其他5HT2BR激动剂在啮齿动物中的作用存在差异，这使人对动物研究结果与预测人类风险的相关性产生怀疑。在此，一种候选化合物RO3013，在每日给药仅3天后就诱导大鼠二尖瓣和三尖瓣的心内膜下细胞增殖。此外，增殖标志物Ki67的免疫染色出现了与治疗相关的增加，并且心脏中指示TGFβ信号传导的磷酸化Smad3与5HT2BR表达共定位。为了证实RO3013诱导的瓣膜增殖继发于5HT2BR激活这一假设，对该化合物进行了体外评估，发现它与人5HT2BR结合，解离常数（K(i)）为3.8μM；然而，在人类细胞的功能试验中它几乎没有激动剂活性。相比之下，RO3013与大鼠5HT2BR结合，K(i)为1.2μM，并以0.5μM的半数有效浓度（EC50）激活该受体。这种激动剂效力估计与观察到瓣膜增殖时RO3013的游离血浆浓度高度一致。这些结果表明，大鼠可能与人类一样易受5HT2BR介导的瓣膜增殖影响；然而，结合活性和功能活性之间的显著差异可能是观察到的物种选择性受体反应的一个可能解释。
