DISCUSSION
Herein, we describe a single-centre, prospective, double-blind, randomised, placebo-controlled, two-treatment, two-sequence, two-period crossover study with 400 mg zamicastat/placebo, in healthy male subjects. The cold stimulus caused a marked sympathetic response evidenced by a significant increase in BP and HR during CPT, at baseline (day -1) and after a 10-day treatment with zamicastat or placebo.
CPT was selected to explore the effect of oral zamicastat, at steady-state conditions, on the overdrive of the sympathetic system and consequent effects on the cardiovascular system. This test has been used both clinically and experimentally to evaluate non-baroreflex-mediated sympathetic neural control in humans, cardiovascular reactivity to stress in normotensive and hypertensive subjects, and the efficacy of lifestyle and pharmacological interventions on BP and vascular reactivity.
Even though our results followed the tendencies presented in other reports, showing increased BP and HR, the increment in HR was lower than in other studies, in which the increase could be higher than 20 bpm, after a 90-second immersion in cold water. However, in other studies CPT showed to have lower or no effect on HR. On the other hand, the change in BP was higher than in previous studies, reaching about 30 mm Hg. These variations may be related to differences in the baseline BP values of the participants as well as to other demographic and behavioural patterns such as age, gender, weight, and exercise habits, which have been shown to influence reactivity to cold-induced stress. Despite the differences, our results on day 10 illustrate the absence of habituation or learning from exposure to the cold stimulus. In fact, both BP and HR kept the same increasing trend with similar magnitude, after 10 days. This observation combined with the referred influence of demographic and behavioural patterns on the reactivity to CPT can guide the definition of future selection criteria that can guarantee the inclusion of subjects with more robust baseline conditions to promote adaptation and learning during cold exposure. This would enrich the cohort by reducing selection bias. However, this endeavour requires a more detailed analysis of the correlation of the baseline characteristics of our participants with habituation to the cold stressor.
Women were not eligible for this study to avoid the potential influence of fluctuation of sex hormones throughout the menstrual cycle with neurohumoral variations with impact on the cardiovascular system.
Compared to placebo, zamicastat significantly decreased SBP and MAP response to cold stimulus during CPT, following 10 days of treatment, evidencing its effect on the overdrive sympathetic response to cold stimulus. Zamicastat reduced SBP (-4.62 mmHg) and DBP (-1.86 mmHg) when compared with placebo. The HR slightly increased (0.81 bpm) which may be related to compensatory hemodynamic effects. The one-sided 95% confidence interval (-7.77 mmHg) was below the established clinical margin for SBP (-5.5 mmHg) supported by a statistically significant difference (p=0.020) between zamicastat and placebo.
The measurement of plasma and urine catecholamines proved to be a valid tool to evaluate the impact of zamicastat on the SNS. These data showed that zamicastat 400 mg, given once daily, inhibited DβH plasmatic activity, increased the 24-h urinary excretion of HVA, and significantly decreased the 24-h urinary excretion of EPI, NE, and VMA, when compared to placebo. The decrease of NE in urine 24h explains the reduction of its metabolite VMA, which reflects a decrease in the total production of catecholamines caused by zamicastat treatment. On the other hand, the levels of DA in urine 24h were similar with zamicastat and placebo, but its metabolite HVA increased, which may imply that a higher amount of DA was metabolized. Our results confirm that the 24h urinary catecholamines is a collector of plasmatic levels of catecholamines and, as such, is more assertive as a potential biomarker of SNS modulation than the sporadic assessment of catecholamines in plasma, which is more affected by fluctuations and interferences than an integrated measurement over 24 h.
Overall, the profile of AEs showed that placebo and zamicastat treatments were similarly well tolerated.
Our results suggest that mechanism of action mediated by zamicastat can be a promising therapeutic option for morbidities characterized by excessive activation of the SNS, such as PAH, ischemic disease, among others.
In conclusion, our study confirmed the effect of zamicastat on the SNS through the decrease of SBP and MAP response to cold stimulus during CPT and inhibition of DβH plasmatic activity. In addition, the effect of zamicastat in 24 h urine catecholamines and their metabolites proved to be clinically relevant, thereby confirming its potential as SNS modulator.