Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): National Research, Development and Innovation Office Hungary Introduction Unexpected ischaemia-induced cardiac adverse events are major contributors to clinical trial discontinuation and drug attrition, and these effects remain undetected by routine preclinical cardiotoxicity assessment. On the other hand, the proarrhythmic effect of drug candidates is studied in healthy cells, tissues, or exclusively in healthy experimental animals. However, a number of experimental and clinical reports demonstrate that the proarrhythmic effects of most drugs are manifested only in pathological conditions. Thus, the aim is to develop a sensitized animal model that can reliably screen for the arrhythmogenic effects of a compound. Here we show that the selective COX- 2 inhibitor rofecoxib, which has been withdrawn over cardiovascular safety concerns, possesses cardiac electrophysiological adverse effects only revealed during ischaemia/reperfusion, a phenomenon we termed "hidden cardiotoxicity". Purpose Our group has previously reported the hidden cardiotoxic effect of rofecoxib on rat ventricular preparations. Given the significant differences in cardiac electrophysiological properties between rats and humans, the human extrapolation of arrhythmological results obtained from rats is limited, so our aim was to investigate the proarrhythmic effect of rofecoxib in a sensitized rabbit model. Methods Action potentials were registered from rabbit right ventricular papillary muscles using the conventional microelectrode technique. The following parameters were measured under normoxic and ischaemic conditions in the presence or absence of 10 μM rofecoxib: conduction time, action potential amplitude, action potential duration at 25%, 50%, 75% and 90% repolarization (APD25-90). The effects of rofecoxib upon test ischaemia and reperfusion were investigated. Results Rofecoxib (10 μM) did not alter electrophysiological parameters in normoxic conditions. However, following 30 minute ischaemia the APD90 was significantly decreased during reperfusion compared to APD90 in the vehicle-treated group: 161.96 ± 19.55 ms vs. 180.04 ± 23.21 ms (p<0.05, n=5), respectively. Following sI/R, a decrease in Vmax and AMPL parameters indicating impulse conduction velocity was also measured in the 10 μM rofecoxib- treated group, but the differences were not statistically significant. Conclusions Under pathological conditions, rofecoxib may increase the incidence of reperfusion arrhythmias due to the mechanism of delayed afterdepolarization (DAD) and reentrant arrhythmias. Consequently, the sensitized rabbit model detailed above may be suitable for investigating the "hidden cardiotoxic" effect of a drug candidate compound. Significant differences were observed in the effect of rofecoxib on repolarization between the rat and rabbit models. However, due to the well- known electrophysiological differences between the two species, the human relevance of the results obtained in rabbits is more reliable.

中文翻译：

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罗非昔布对兔心室制剂的潜在心脏毒性作用

资金致谢 资金来源类型：公共拨款——仅限国家预算。主要资金来源： 匈牙利国家研究、发展和创新办公室 引言 意外缺血引起的心脏不良事件是导致临床试验中止和药物损耗的主要因素，而常规临床前心脏毒性评估仍未发现这些影响。另一方面，在健康细胞、组织或仅在健康实验动物中研究候选药物的促心律失常作用。然而，许多实验和临床报告表明，大多数药物的促心律失常作用仅在病理条件下表现出来。因此，目的是开发一种致敏动物模型，可以可靠地筛选化合物的致心律失常作用。在这里，我们展示了选择性 COX-2 抑制剂罗非昔布，它已因心血管安全问题而被撤回，具有仅在缺血/再灌注期间显示的心脏电生理不良反应，我们称之为“隐藏的心脏毒性”现象。目的 我们小组之前曾报道过罗非昔布对大鼠心室制剂的潜在心脏毒性作用。鉴于大鼠和人类心脏电生理特性的显着差异，从大鼠获得的心律失常结果的人类外推是有限的，因此我们的目的是研究罗非昔布在致敏兔模型中的促心律失常作用。方法采用常规微电极技术对兔右心室乳头肌动作电位进行记录。在存在或不存在 10 μM 罗非昔布的情况下，在常氧和缺血条件下测量以下参数：传导时间、动作电位幅度、25%、50%、75% 和 90% 复极时的动作电位持续时间 (APD25-90)。研究了罗非昔布对受试缺血和再灌注的影响。结果 Rofecoxib (10 μM) 在常氧条件下没有改变电生理参数。然而，在缺血 30 分钟后，再灌注期间 APD90 与载体治疗组中的 APD90 相比显着降低：分别为 161.96 ± 19.55 ms 对 180.04 ± 23.21 ms (p＜0.05，n=5)。在 sI/R 之后，在 10 μM 罗非昔布治疗组中也测量了指示脉冲传导速度的 Vmax 和 AMPL 参数的降低，但差异无统计学意义。结论 在病理条件下，罗非昔布可能因延迟后除极（DAD）和折返性心律失常的机制增加再灌注心律失常的发生率。因此，上面详述的致敏兔模型可能适用于研究候选药物化合物的“隐藏的心脏毒性”效应。在大鼠和兔模型之间观察到罗非昔布对复极化的影响存在显着差异。然而，由于两个物种之间众所周知的电生理差异，在兔子中获得的结果与人类相关性更可靠。上面详述的致敏兔模型可能适用于研究候选药物化合物的“隐藏的心脏毒性”效应。在大鼠和兔模型之间观察到罗非昔布对复极化的影响存在显着差异。然而，由于两个物种之间众所周知的电生理差异，在兔子中获得的结果与人类相关性更可靠。上面详述的致敏兔模型可能适用于研究候选药物化合物的“隐藏的心脏毒性”效应。在大鼠和兔模型之间观察到罗非昔布对复极化的影响存在显着差异。然而，由于两个物种之间众所周知的电生理差异，在兔子中获得的结果与人类相关性更可靠。