A Randomized Trial on the Combined Effect of Ponesimod and Propranolol on Heart Rate, Cardiac Safety, and Pharmacokinetics in Healthy Adults

ABSTRACT

The objective of this phase 1 study was to evaluate the pharmacokinetics (PK), pharmacodynamics, and cardiac effect following administration of ponesimod (a selective sphingosine‐1‐phosphate receptor modulator) and propranolol in healthy adults. In treatment period (TP) 1, participants received ponesimod (2 mg). In TP2, if resting heart rate (HR) was ≥ 55 bpm, the ponesimod up‐titration regimen was initiated. Participants were randomized to TP2A (placebo plus ponesimod up‐titration) or TP2B (80 mg propranolol plus ponesimod up‐titration). Concomitant administration resulted in an increased bradyarrhythmic effect on HR versus ponesimod alone. The mean maximum difference in mean hourly HR from time‐matched baseline for TP2B compared with TP2A during the first 12 h post‐dose was −12.4 bpm. This was observed after the first dose of ponesimod, persisted for the first 4 doses, then decreased to −7.4 bpm post‐up‐titration. The lowest mean of the HR_nadir in TP2B was 48.9 bpm (95% CI: 46.4–51.3). There was no significant difference in the occurrence of 1st degree AV block between groups and no occurrences of 2nd or higher degree AV block. No clinically relevant changes were observed in the PK of ponesimod or propranolol. Overall, 88.5% of participants experienced ≥ 1 AE during the study. In TP2, the most reported TEAEs (≥ 5%) considered related to ponesimod were fatigue (12 [25.5%]) and dizziness (10 [21.3%]). No deaths were reported. Co‐administration of ponesimod with propranolol resulted in a greater HR‐lowering effect compared to ponesimod alone, without significant changes in PK parameters or serious cardiac adverse events in healthy adults.

Study Highlights.

• What is the current knowledge on the topic?

  • ○
    Ponesimod is approved for the treatment of relapsing forms of multiple sclerosis with active disease defined by clinical or imaging features.
• What question did this study address?

  • ○
    This randomized, double‐blind, parallel‐group, 2‐period, placebo‐controlled, phase 1 study in healthy individuals was designed to identify any drug–drug interactions with the combined use of ponesimod and propranolol.
• What does this study add to our knowledge?

  • ○
    Results from the study show there was no clinically relevant PK drug–drug‐interaction between ponesimod and propranolol. The combined treatment of propranolol and ponesimod resulted in an additive effect on HR, and this HR effect was reduced with subsequent doses. ECG abnormalities were observed with combined treatment but were not considered clinically relevant.
• How might this change clinical pharmacology or translational science?

  • ○
    Although the HR effect observed with the coadministration of ponesimod uptitration regimen with propranolol did not lead to related serious AE's or clinically significant ECG abnormalities, results from this study suggest that resting HR should be considered when ponesimod is initiated in patients receiving treatment with a beta‐blocker because of the additive effects on lowering heart rate. Temporary interruption of the beta‐blocker treatment may be needed in some patients prior to initiation of ponesimod.

Abbreviations

∆HR_max

maximum decrease from time‐matched baseline in mean hourly heart rate on each day

AE

adverse event

AESI

adverse event of special interest

AUC_24h

area under the plasma analyte concentration‐time curve (AUC) from time 0 to 24 h postdose, calculated by linear‐linear trapezoidal summation

AUC_τ

AUC during a dosing interval (τ) at steady state, calculated by linear‐linear trapezoidal summation

AV

atrioventricular

BP

blood pressure

bpm

beats per minute

CL_SS/F

total apparent oral clearance at steady state, calculated as dose/AUC_τ

C_max

maximum observed plasma analyte concentration

C_trough

observed analyte concentration just prior to the beginning of a dosing interval

HR

heart rate

HR_nadir

minimum of the mean hourly heart rate for each day

mHR

mean hourly heart rate

PD

pharmacodynamic

PK

pharmacokinetic

QTcF

QT interval corrected for heart rate according to Fridericia's formula

S1PR

sphingosine‐1‐phosphate receptor

t_max

actual sampling time to reach the maximum observed plasma analyte concentration

1. Introduction

Ponesimod is a selective sphingosine‐1‐phosphate receptor (S1PR) modulator that blocks the egress of lymphocytes from lymphoid tissue, thereby reducing the number of circulating effector T cells that can invade target organs [1, 2, 3, 4]. Ponesimod is approved for the treatment of adults with active relapsing multiple sclerosis (MS) in the United States and European Union [5, 6, 7, 8].

Beta‐blockers are widely used for the treatment of hypertension and are known to decrease heart rate (HR) in some people. Several studies suggest that the co‐administration of S1PR modulators with beta‐blockers should be considered carefully [9]. Specifically, a study evaluating the combined effect of the beta blocker, atenolol, with the S1PR modulator, fingolimod, showed that heart rate nadir (HR_nadir) was decreased when the two drugs were combined compared with fingolimod alone [10]. In a separate study, concomitant administration of atenolol (50 mg once daily for 5 days) with a single dose of 10 mg ponesimod (without gradual up‐titration) had an additive pharmacodynamic (PD) effect on HR [11]. Bradycardia‐related serious adverse events (SAEs) led to termination of the study for safety reasons.

Studies show that the initiation of S1PR modulators, such as ponesimod, can cause a transient, dose‐dependent decrease in HR [12, 13]. To mitigate the first‐dose bradyarrhythmic effect, a gradual up‐titration regimen of ponesimod was evaluated in separate studies [8, 14, 15, 16, 17]. Compared to a starting dose of 10 mg for 1 week followed by a single‐step up‐titration to 20 mg, a gradual up‐titration of ponesimod was found to elicit fewer and less pronounced cardiodynamic effects [15, 16]. Studies confirm that an up‐titration regimen has been found to considerably attenuate the bradycardic effects during ponesimod initiation [17, 18]. Model‐based simulations indicate that the incidence of bradycardia at the initiation of ponesimod treatment is expected to substantially decrease in participants with a baseline HR > 55 beats per minute (bpm) with the gradual up‐titration regimen [16, 17].

Given that beta blockers and S1P1 receptor (S1P1R) modulators can affect HR, the goal of this study was to evaluate the pharmacokinetic (PK), PD, and cardiac effects following administration of ponesimod gradual up‐titration in healthy adult participants receiving propranolol at steady state.

2. Methods

2.1. Study Design

This was a randomized, double‐blind, parallel‐group, 2‐period, placebo‐controlled study (Figure 1). A computer‐generated randomization schedule was prepared prior to study start. Randomization was balanced by using randomly permuted blocks and was stratified by sex. The sample size was determined based on the estimators of the various endpoints of interest, with a focus on the primary endpoint: the change from baseline in mean hourly heart rate (ΔHR). Participants received treatments under fed conditions with a washout period of at least 7 days and a maximum of 14 days between the 2 treatment periods. Treatment period (TP) 1 was an open‐label treatment period (Days −1 to 2), and all participants received a single oral dose of 2 mg ponesimod. TP2 was a double‐blind treatment period (Days −2 to 26), and participants were randomized to 1 of 2 treatments (TP2A or TP2B). In TP2A, participants received placebo propranolol once daily from Day 1 to 19 plus an up‐titration regimen of ponesimod (2, 2, 3, 3, 4, 4, 5, 6, 7, 8, 9, 10, 10, 10, and 20 mg/day) from Day 5 to 19. In TP2B, participants received 80 mg propranolol once daily from Day 1 to 19 plus the same up‐titration regimen of ponesimod as in TP2A. The study took place in Belgium from March to August of 2019.

FIGURE 1.

Study design. This study consisted of 3 phases: a screening phase, a 2‐period treatment phase, and an end‐of‐study phase. Randomization was stratified by sex. Treatment period (TP) 1 was an open‐label treatment. In the morning of Day 1, participants received a single dose of 2 mg ponesimod, and HR and ECG parameters were monitored. In TP2 (double‐blind treatment period), participants were randomized to 1 of 2 treatments (TP2A or TP2B). In TP2A, participants received an up‐titration regimen of ponesimod from Day 5 to 19 plus placebo propranolol from Day 1 to 19. In TP2B, participants received a similar regimen of ponesimod (as for TP2A) plus 80 mg propranolol from Day 1 to 19. BMI, body mass index; bpm, beats per minute; ECG, electrocardiogram; HR, heart rate; PD, pharmacodynamics; PK, pharmacokinetics; TP, treatment phase.

In both TP2A and TP2B, the up‐titration regimen was initiated if the pre‐dose 12‐lead electrocardiogram (ECG) criteria on Day 5 were met (HR ≥ 55 bpm, HR > 70% of baseline, and no clinically relevant ECG abnormalities ongoing).

The study protocol and amendments were reviewed by an independent ethics committee (Commissie voor Medische Ethiek—ZNA/OCMW Antwerpen, Belgium), and all applicable regulatory requirements were followed (Protocol ID: AC‐058‐117; approval date: 13 March 2019). This study was conducted following the ethical principles outlined in the Declaration of Helsinki and is consistent with the International Conference on Harmonization and Good Clinical Practice guidelines [19, 20]. Participants provided their written informed consent to participate in the study (Clinicaltrials.gov, NCT03882255).

2.2. Inclusion and Exclusion Criteria

Eligible participants were healthy, between 18 and 55 years of age, and had a blood pressure (BP) of 90–140 mmHg systolic and 50–90 mmHg diastolic with normal cardiac conduction and function as determined by 12‐lead safety ECG. Other required criteria were a HR of 55–100 bpm, QT interval corrected for HR using Fridericia's formula (QTcF) of ≤ 450 ms for males and ≤ 470 ms for females, a QRS interval of < 120 ms, a PR interval ≤ 200 ms, and ECG morphology consistent with healthy cardiac conduction and function.

Participants with any cardiac condition or illness (including ECG abnormalities) were excluded. Participants were discontinued if at any time during the study, the participants' HR was < 40 bpm as measured by any 12‐lead safety ECG, HR was < 45 bpm for more than 1 h on cardiac telemetry, in cases of symptomatic bradycardia (e.g., presyncope/syncope associated with bradycardia, confusion, chest pain), or if participants developed a 2^nd or 3^rd degree AV block (12‐lead safety ECG).

2.3. Pharmacokinetic Evaluations

Blood samples for PK assessments were taken over a 24‐h PK sampling period on Day 4 (last day of propranolol single agent treatment), Day 5 (at initiation of ponesimod treatment at the dose of 1 mg), and Day 19 (last day of ponesimod treatment, at the dose of 20 mg) (Figure 1). Plasma was analyzed using a validated, specific, and sensitive achiral liquid chromatography coupled to tandem mass spectrometry method using a lower limit of quantification of 1 ng/mL [4, 14]. PK analyses were performed using the validated computer program Phoenix WinNonlin (version 6.2.1). Non‐compartmental analysis was applied for the PK analysis.

PK parameters were derived for ponesimod on Day 5 (maximum observed plasma analyte concentration [C_max], actual sampling time to reach the maximum observed plasma analyte concentration [t_max], and area under the plasma analyte concentration‐time curve from time 0–24 h postdose [AUC_24h]) and on Day 19 (observed analyte concentration just prior to the beginning of a dosing interval [C_trough], C_max, t_max, and AUC during a dosing interval (τ) at steady state [AUC_τ]). Propranolol and 4‐hydroxypropranolol (TP2B only) were determined on Day 4 and Day 5 (C_trough, C_max, t_max, and AUC_τ, and total apparent oral clearance at steady state, calculated as dose/AUC_τ [CL_ss/F]) and on Day 19 (C_trough, C_max, t_max, and AUC_τ, and CL_ss/F).

2.4. Heart Rate Evaluations

PD parameters were investigated on Day 1 of TP1 and from Day 1 to 19 of TP2 (Figure 1). PD endpoints included HR and bradyarrhythmia and were evaluated using Holter ECG recordings. The primary endpoint was the change from baseline in mean hourly HR (ΔHR). Resting HR was also measured using 10‐s 12‐lead ECGs and 10‐ to 20‐s vital signs measurements. For both methods, participants remained in a supine position for at least 5 min before measurement to ensure HR stability.

2.5. Lymphocyte Evaluations

The effect of ponesimod on lymphocyte count was evaluated throughout the study, including absolute lymphocyte count and the percent change from baseline to absolute lymphocyte count.

2.6. Safety Assessments

Safety and tolerability were evaluated throughout the study and included all participants who received at least 1 dose of ponesimod. Single 12‐lead safety ECGs were recorded and assessed immediately. Telemetry was performed continuously starting ~1 h prior to study drug intake on Day 1 of TP1 until 12 h after study drug intake and continuously starting ~1 h prior to study drug intake on Day 1 of TP2 until 24 h after the last study drug intake on Day 19.

2.7. Statistical Analysis

Statistical analysis was performed for (log‐transformed) ponesimod C_max and AUC_24h (Day 5), and C_max and AUC_τ (Day 19) of TP2, and propranolol and 4‐hydroxypropranolol C_max and AUC_τ in TP2. The least square means (LS means) of the log‐transformed primary parameters of ponesimod were estimated with a linear mixed effects model, with treatment (TP2A and TP2B) and Day (5 and 19) as fixed effects and subject as a random effect. For each PK parameter, the data were fitted in one statistical model from which the required pairwise comparisons were obtained (Day 5 of TP2B versus Day 5 of TP2A and Day 19 of TP2B versus Day 19 of TP2A). For the propranolol analyses, the LS means of the log‐transformed primary parameters was estimated with a linear mixed effects model, with treatment day as a fixed effect and subject as a random effect. For each PK parameter, the data were fitted in one statistical model from which the required pairwise comparisons were obtained (TP2B: Day 5 vs. Day 4 and Day 19 vs. Day 4). A 90% confidence interval (CI) was constructed around the difference between the LS means on the treatments/days being compared. Both the difference between the LS means and the 90% CIs were retransformed to the original scale. All observations for test and reference were included in the statistical analysis.

The ΔHR_max (maximum decrease from time‐matched baseline in mean hourly HR on each day), HR_nadir (minimum of the mean hourly HR for each day), mHR (mean hourly heart rate), and ΔHR (change in heart rate) were compared between TP2A and TP2B for assessment days using 2‐sample t‐tests or paired t‐tests. The mean and the mean difference between treatment groups were reported along with the 95% CI and 90% CI, respectively.

3. Results

3.1. Participant Demographics and Study Completion

A total of 52 participants were enrolled in TP1 (Figure S1). The median age was 36.5 years (range: 20–54), the median body mass index (BMI) was 25.1 (range: 18.9–29.7), and 27 participants (51.9%) were male (Table S1). During TP1, a total of 5 participants (9.6%) terminated the study prematurely; 3 participants (5.8%) were inadvertently terminated from the study as the pre‐dose exclusion criterion of HR < 55 bpm was incorrectly applied as a post‐dose criterion, 1 participant (1.9%) was discontinued due to an AE of supraventricular arrhythmia (atrial escape rhythm); and 1 participant (1.9%) withdrew consent.

Forty‐seven participants (90.3%) completed TP1 and were enrolled in TP2, and 38 of the TP2 participants (80.9%) completed the study (21 participants [87.5%] in TP2A and 17 participants [73.9%] in TP2B).

During TP2A, 3 participants (12.5%) terminated the study prematurely: 1 (4.2%) withdrew consent; 1 (4.2%) had an AE of ventricular tachycardia (non‐sustained ventricular tachycardia) that was considered not related to study treatment; and 1 (4.2%) met the discontinuation criterion of HR < 45 bpm for more than 1 h.

During TP2B, a total of 6 participants (26.1%) terminated the study prematurely: 1 (4.3%) had an AE of redness due to ECG patches, and 5 (21.7%) had HR < 45 bpm for more than 1 h. In addition, 4 participants who entered the up‐titration regimen on Day 5 had a pre‐dose HR < 55 bpm and therefore did not meet the criteria for up‐titration but were inadvertently dosed on Day 5 and subsequent days. Three of these participants discontinued the study on Days 7, 8, and 12 as they met the bradycardia discontinuation criterion (HR < 45 bpm for a period of more than 1 h), and 1 participant completed the study.

3.2. Pharmacokinetics

The PK of ponesimod and propranolol are summarized in Table 1. Briefly, median ponesimod t_max was 4–5 h post‐dose (range of 2–8 h), both in the absence and presence of propranolol. Based on the geometric mean ratios (GMRs), on Day 5, ponesimod C_max and AUC_0–24h were similar in TP2B compared to TP2A. On Day 19, the GMRs of TP2B versus TP2A for ponesimod C_max and AUC_24h were 111% (90% CI: 102.98–120.48) and 110% (90% CI: 101.75–119.32), respectively.

TABLE 1.

Pharmacokinetics of ponesimod and propranolol on Days 4, 5 and 19 for TP2A (placebo plus ponesimod) and TP2B (propranolol plus ponesimod).

Parameter	Day 4	Day 5	Day 19
Pharmacokinetics of ponesimod in TP2A
N	NA	22	21
Ponesimod (mg)	0	2	20
Ctrough (ng/mL)	NA	NA	71.4 (13.9)
Cmax (ng/mL)	NA	12.3 (1.65)	188 (24.2)
tmax (h)	NA	5.00 (2.00–8.00)	4.00 (3.00–5.00)
AUC0–24h (ng.h/mL)	NA	173 (16.4)	3126 (450)
Pharmacokinetics of ponesimod in TP2B
N	NA	22 a	17
Ponesimod (mg)	0	2	20
Ctrough (ng/mL)	NA	NA	78.7 (20.6)
Cmax (ng/mL)	NA	12.9 (2.31)	211 (35.1)
tmax (h)	NA	5.00 (2.00–5.07)	4.00 (3.00–5.00)
AUC0–24h (ng.h/mL)	NA	185 (29.6)	3448 (534)
Pharmacokinetics of propranolol in TP2B
N	23	22	17
Propranolol (mg)	80	80	80
Ctrough (ng/mL)	12.6 (8.60)	10.7 (7.52)	11.5 (5.00)
Cmax (ng/mL)	44.3 (29.5)	36.6 (18.8)	41.6 (16.9)
tmax (h)	8.00 (5.00–12.00)	6.18 (4.98–12.00)	8.00 (4.98–10.00)
AUC0–24h (ng.h/mL)	583 (370)	534 (290)	566 (211)

Note: Data for C_trough, C_max, and AUC_0–24 is presented as mean (SD); data for t_max, is presented as median (range). TP2A, participants received placebo propranolol once daily from Day 1 to 19 plus an up‐titration regimen of ponesimod (2, 2, 3, 3, 4, 4, 5, 6, 7, 8, 9, 10, 10, 10 and 20 mg) from Day 5 to 19; TP2B, participants received 80 mg propranolol once daily from Day 1 to 19 plus the same up‐titration regimen of ponesimod as in TP2A.

Abbreviations: AUC_0–24h, area under the plasma analyte concentration‐time curve (AUC) from time 0 to 24 h post dose, calculated by linear‐linear trapezoidal summation; C_max, maximum observed plasma analyte concentration; C_trough, observed analyte concentration just prior to the beginning of a dosing interval; N, number of participants with data; NA, not applicable; t_max, actual sampling time to reach the maximum observed plasma analyte concentration.

^a n = 21 for AUC_0–24h.

Median propranolol t_max was achieved 8 h post‐dose on Day 4 (propranolol alone), Day 5 (2 mg ponesimod plus propranolol), Day 6, and Day 19 (20 mg ponesimod plus propranolol), with similar ranges on all treatment days. Mean propranolol C_trough levels were comparable on Day 4, 5, and 19. Propranolol GMR of Day 5 versus Day 4 for C_max and AUC_24h were 89% (90% CI: 77–101) and 93% (90% CI: 85–102), respectively, and for Day 19 versus Day 4 for C_max and AUC_24h 108% (90% CI 93–125) and 111% (90% CI 100–123).

3.3. Effect on Heart Rate

The mHR on Holter for all participants remained above 40 bpm at all time points during both treatments. A statistical comparison of ΔHR_max and HR_nadir for TP2B versus TP2A using the time‐matched baseline is shown in Table 2.

TABLE 2.

Comparison of ∆HR_max using time‐matched baseline and HR_nadir for TP2B (propranolol plus ponesimod) versus TP2A (placebo plus ponesimod).

	Day	Ponesimod (mg)	TP2B (propranolol + ponesimod)		TP2A (placebo + ponesimod)		Comparison for TP2B vs. TP2A
			N	Mean (95% CI)	N	Mean (95% CI)	Mean (90% CI)
∆HRmax over 12 h (bpm)	5	2	21	−23.5 (−26.39; −20.66)	20	−11.2 (−13.96; −8.34)	−12.4 (−15.61; −9.14)
	6	2	20	−24.4 (−27.30; −21.40)	19	−12.1 (−14.86; −9.35)	−12.2 (−15.50; −8.99)
	7	3	20	−25.9 (−29.02; −22.78)	20	−15.6 (−18.04; −13.16)	−10.3 (−13.49; −7.11)
	8	3	18	−25.7 (−29.40; −22.04)	20	−16.1 (−18.39; −13.81)	−9.6 (−13.02; −6.22)
	16	10	16	−22.9 (−26.34; −19.41)	20	−14.0 (−16.92; −10.98)	−8.9 (−12.57; −5.28)
	19	20	16	−21.9 (−25.57; −18.30)	20	−14.6 (−17.18; −11.92)	−7.4 (−10.89; −3.88)
HRnadir over 12 h (bpm)	5	2	21	52.2 (49.65; 54.73)	20	62.7 (59.30; 66.10)	−10.5 (−13.90; −7.12)
	6	2	20	50.9 (48.67; 53.13)	19	62.3 (58.67; 65.85)	−11.4 (−14.72; −8.01)
	7	3	20	48.9 (46.43; 51.27)	20	58.5 (55.60; 61.40)	−9.7 (−12.69; −6.61)
	8	3	18	50.4 (48.17; 52.72)	20	60.3 (57.48; 63.02)	−9.8 (−12.73; −6.89)
	16	10	16	53.6 (51.68; 55.57)	20	60.2 (57.38; 63.02)	−6.6 (−9.48; −3.67)
	19	20	16	54.1 (51.72; 56.53)	20	60.3 (57.38; 63.12)	−6.1 (−9.23; −3.02)

Note: TP2A, participants received placebo propranolol once daily from Day 1 to 19 plus an up‐titration regimen of ponesimod (2, 2, 3, 3, 4, 4, 5, 6, 7, 8, 9, 10, 10, 10 and 20 mg) from Day 5 to 19; TP2B, participants received 80 mg propranolol once daily from Day 1 to 19 plus the same up‐titration regimen of ponesimod as in TP2A.

Abbreviations: ∆HR_max, maximum decrease from baseline in mean hourly heart rate from each day; bpm, beats per minute; HR_nadir, minimum absolsute value of the observed mean hourly heart rate on each day; N, number of participants with data; TP2A, treatment period 2A; TP2B, treatment period 2B.

3.4. Individual Effect of Ponesimod

The first dose of ponesimod on Day 1 in TP1 resulted in a transient decrease in the mHR versus the time‐matched baseline of 8.4 bpm at 3 h post‐dose. The mean nadir of the mHR was 61.1 bpm observed at 4 h post‐dose. The minimum value for mHR at 4 h post‐dose was 52 bpm.

The first dose of ponesimod on Day 5 of TP2A produced an initial maximum decrease in mean of the mHR of 6.1 bpm at 3 h post‐dose, followed by another decrease of 6.6 bpm at 10 h.

The first dose of ponesimod 2 mg alone also resulted in the lowest mHR mean of 65.5 bpm, at 4 h post‐dose (Figure 2A). The average minimum of the individual mHR (HR_nadir) on day 5 in TP2A was 62.7 bpm (Table 2).

FIGURE 2.

Mean of the mHR over time for TP2B (propranolol plus ponesimod). (A) Mean of the mHR over time for TP2B at baseline (Day −1) following repeated doses of ponesimod in combination with placebo (TrtA) or propranolol (Trt B). (B) Mean of the mHR over time for TP2B at baseline (Day −1) following a single dose of propranolol (Day 1) and at steady state (Day 4). Bpm, beats per minute; mHR, mean heart rate; TP, treatment phase; Trt, treatmen.

With increasing ponesimod doses, the lowest average value of the mHR was 64.5, 60.0, 63.2, 63.4, and 62.0 bpm at 4 h post‐dose on Day 6, 7, 8, 16, and 19, respectively (Figure 2A) and the corresponding values of the HR_nadir were 62.3, 58.5, 60.3, 60.2, and 60.3 bpm, respectively (Table 2).

3.5. Individual Effect of Propranolol

On Day 1 of TP2B, a single dose of propranolol produced an initial maximum decrease versus time‐matched baseline in the mHR of 8.0 bpm at 6 h post‐dose. On Day 4, after repeated doses of propranolol, a greater decrease of 16.5 bpm was observed at 10 h post‐dose. The lowest value of the mHR was 62.2 bpm observed at 4 h post‐dose after a single dose of propranolol on Day 1 and 59.6 bpm at 10 h after repeated doses on Day 4 (Figure 2B).

3.6. Combined Effect of Propranolol and Ponesimod

Comparisons were also made within TP2B evaluating the effect of the combination ponesimod and propranolol, propranolol alone (Day 4, Period 2), or ponesimod alone (Day 1, period 1). The comparisons for ∆HR_max and HR_nadir over 12 h are summarized in Table 3.

TABLE 3.

Comparison of ∆HR_max and HR_nadir within TP2B using time‐matched baseline.

	Days	Dose a	TP2B (propranolol + ponesimod)		TP2A (placebo + ponesimod)		Comparison Within TP2B
			N	Mean (95% CI)	N	Mean (95% CI)	Mean (90% CI)
∆HRmax over 12 h (bpm)	5 vs. 1	PON(2 mg) + PRO vs. PON(2 mg)	21	−23.5 (−26.39; −20.66)	22	−13.4 (−15.81; −11.01)	−10.6 (−11.88; −9.26)
	5 vs. 4	PON(2 mg) + PRO vs. PRO	21	−23.5 (−26.39; −20.66)	22	−18.9 (−21.67; −16.06)	−4.6 (−5.84; −3.40)
	6 vs. 4	PON(2 mg) + PRO vs. PRO	20	−24.4 (−27.30; −21.40)	22	−18.9 (−21.67; −16.06)	−5.9 (−6.70; −5.00)
	7 vs. 4	PON(3 mg) + PRO vs. PRO	20	−25.9 (−29.02; −22.78)	22	−18.9 (−21.67; −16.06)	−7.4 (−8.52; −6.28)
	8 vs. 4	PON(3 mg) + PRO vs. PRO	18	−25.7 (−29.40; −22.04)	22	−18.9 (−21.67; −16.06)	−7.3 (−8.67; −5.89)
	16 vs. 4	PON(10 mg) + PRO vs. PRO	16	−22.9 (−26.34; −19.41)	22	−18.9 (−21.67; −16.06)	−5.0 (−6.72; −3.28)
	19 vs. 4	PON(20 mg) + PRO vs. PRO	16	−21.9 (−25.57; −18.30)	22	−18.9 (−21.67; −16.06)	−4.1 (−6.03; −2.09)
HRnadir over 12 h (bpm)	5 vs. 1	PON(2 mg) + PRO vs. PON(2 mg)	21	52.2 (49.65; 54.73)	22	59.6 (57.27; 62.00)	−7.8 (−9.56; −5.97)
	5 vs. 4	PON(2 mg) + PRO vs. PRO	21	52.2 (49.65; 54.73)	22	57.5 (55.39; 59.61)	−5.6 (−6.73; −4.41)
	6 vs. 4	PON(2 mg) + PRO vs. PRO	20	50.9 (48.67; 53.13)	22	57.5 (55.39; 59.61)	−6.7 (−7.86; −5.44)
	7 vs. 4	PON(3 mg) + PRO vs. PRO	20	48.9 (46.43; 51.27)	22	57.5 (55.39; 59.61)	−8.7 (−10.02; −7.38)
	8 vs. 4	PON(3 mg) + PRO vs. PRO	18	50.4 (48.17; 52.72)	22	57.5 (55.39; 59.61)	−7.6 (−9.10; −6.12)
	16 vs. 4	PON(10 mg) + PRO vs. PRO	16	53.6 (51.68; 55.57)	22	57.5 (55.39; 59.61)	−5.0 (−6.84; −3.16)
	19 vs. 4	PON(20 mg) + PRO vs. PRO	16	54.1 (51.72; 56.53)	22	57.5 (55.39; 59.61)	−4.5 (−6.72; −2.28)

Note: TP2A, participants received placebo propranolol once daily from Day 1 to 19 plus an up‐titration regimen of ponesimod (2, 2, 3, 3, 4, 4, 5, 6, 7, 8, 9, 10, 10, 10 and 20 mg) from Day 5 to 19; TP2B, participants received 80 mg propranolol once daily from Day 1 to 19 plus the same up‐titration regimen of ponesimod as in TP2A.

Abbreviations: ∆HR_max, maximum decrease from baseline in mean hourly heart rate from each day; bpm, beats per minute; HR_nadir, minimum absolute value of the observed mean hourly heart rate on each day; N, number of participants with data; PON, ponesimod; PRO, propranolol; TP2A, treatment period 2A; TP2B, treatment period 2B.

^a The propranolol dose used was 80 mg.

For ΔHR_max over 12 h, a comparison of TP2B (propranolol plus ponesimod) versus TP2A (placebo plus ponesimod) showed that the combination exhibited a 12.4 bpm (90% CI: 9.14–15.61) decrease in mean HR on Day 5 (after the first dose of ponesimod 2 mg) and a 7.4 bpm (90% CI: 3.88–10.89) decrease in mean HR on Day 19 (the day of ponesimod 20 mg dose) compared to ponesimod alone (Table 2, Figure 3).

FIGURE 3.

Change from time‐matched baseline in mean hourly HR after the first dose of ponesimod (Day 5) or after the up‐titration regimen (Day 19) in TP2A (placebo plus ponesimod) and TP2B (propranolol plus ponesimod). bpm, beats per minute; HR, heart rate; h, hour; mHR, mean heart rate; TP, treatment phas.

The lowest mean of the HR_nadir in TP2B was 48.9 bpm (95% CI: 46.43–51.27) observed on Day 7 (third day of the ponesimod up‐titration regimen) and increased to 54.1 bpm (95% CI: 51.72–56.53) on Day 19 (Table 2). For HR_nadir over 12 h, a comparison of TP2B (combination of ponesimod + propranolol) versus TP2A (ponesimod plus placebo) had a 10.5 bpm (90% CI: 7.12–13.90) decrease in the mean HR_nadir on Day 5 (after the first dose of ponesimod 2 mg); the largest decrease in the mean HR_nadir was observed on Day 6 (after the second dose of ponesimod 2 mg) an 11.4 bpm (90% CI: 8.01–14.72) and a 6.1 bpm (90% CI: 3.02–9.23) decrease in the mean HR_nadir was observed on Day 19 (the day of ponesimod 20 mg dose).

In TP2B, compared with propranolol (80 mg) alone (Day 4), when the ponesimod 2 mg dose was added to propranolol (Day 5), there was a −4.6 bpm (90% CI: −5.8 to −3.4) difference in mean ∆HR_max. The largest difference of −7.4 bpm (90% CI: −8.5 to −6.3) was observed on Day 7 after the third dose of ponesimod (3 mg), and on Day 19 at the first dose of ponesimod 20 mg after up‐titration, a −4.1 bpm (90% CI: −6.0 to −2.12) difference in mean ∆HR_max was observed (Table 3).

3.7. Effect on Lymphocyte Counts

Ponesimod treatment resulted in a decrease in circulating lymphocytes. This decrease was similar in TP2A and TP2B. A maximum decrease of approximately 50% from baseline in total lymphocyte count was observed in both treatments following administration of 10 mg ponesimod (on Day 16 for both TP2A and TP2B) (Figures S2 and S3). Lymphocyte count started to increase on Day 21, within 2 days of the last dose administered on Day 19. The lymphocyte count returned to near baseline values by Day 26 and was back to above normal values in all participants.

3.8. Overall Safety Analysis and Study Discontinuation

In total, 46 of 52 participants (88.5%) experienced at least one AE during the study. The most common TEAEs (≥ 5% of participants) are summarized in Table S2, and all observed AEs are listed in Table S3. Briefly, the most commonly reported AEs were fatigue (22 [42.3%]) and headache (20 [38.5]). Ten participants (19.2%) had AEs that were not resolved at the end of the study. None were considered serious or clinically significant. All persistent AEs, except for fatigue observed in 1 participant, were skin irritation or itchiness around the ECG patches. One participant (4.2%) experienced a SAE of breast neoplasm (benign fibroma on pathology) during the study. The event was assessed to be of moderate severity and was considered not related to the study drug by the investigator.

Overall, 3 (5.8%) participants were discontinued from the study due to AEs (Table 4). In TP1, 1 participant experienced an AE of supraventricular arrhythmia (atrial escape rhythm) of mild intensity ~3 h after the first dose of ponesimod. This event was considered related to the study drug by the investigator. In TP2A, 1 participant experienced an AE of non‐sustained ventricular tachycardia of mild intensity. The event was not considered to be related to the study drug. In TP2B, 1 participant experienced an AE of redness due to ECG patches of mild intensity. There were no deaths reported in this study.

TABLE 4.

Summary of HR, AV block and QTcF interval analysis (12‐lead ECG).

Criteria	TP1: 2 mg ponesimod	TP2A: placebo + ponesimod	TP2B: propranolol + ponesimod
N	52	24	23
Overall summary of AEs, N (%)
1 or more TEAE	22 (42.3)	21 (87.5)	21 (91.3)
AE leading to death	0 (0.0)	0 (0.0)	0 (0.0)
Persistent AEs	4 (7.7)	3 (12.5)	4 (17.4)
Serious AEs	0 (0.0)	1 (4.2)	0 (0.0)
AEs leading to discontinuation	1 (1.9)	1 (4.2)	1 (4.3)
AE of special interest	4 (7.7)	9 (37.5)	7 (30.4)
Abnormal HR (bpm), N (%)
Participants with at least 1 finding	31 (59.6)	22 (91.7)	23 (100.0)
HR < 45 bpm	0 (0.0)	2 (8.3)	13 (56.5)
HR > 100 bpm	0 (0.0)	1 (4.2)	0 (0.0)
Decrease from baseline > 5 bpm	12 (23.1)	8 (33.3)	4 (17.4)
Decrease from baseline > 10 bpm	17 (32.7)	11 (45.8)	11 (47.8)
Decrease from baseline > 20 bpm	2 (3.8)	3 (12.5)	4 (17.4)
Decrease from baseline > 30 bpm	0 (0.0)	0 (0.0)	4 (17.4)
Occurrences of AV‐block from safety 12‐lead ECG, N (%)
AV block, 1st degree a	1 (1.9)	6 (25.0)	7 (30.4)
QTcF interval aggregate, N (%)
> 450–480 msec, measured value	1 (1.9)	2 (8.3)	1 (4.3)
> 480–500 msec, measured value	0 (0.0)	0 (0.0)	0 (0.0)
> 500 msec, measured value	0 (0.0)	0 (0.0)	1 (4.3)
Increase from baseline, > 30–60 msec	2 (3.8)	4 (16.7)	8 (34.8)
Increase from baseline, > 60 msec	1 (1.9)	0 (0.0)	1 (4.3)
Total	4 (7.7)	5 (20.8)	9 (39.1)

Note: TP1, all participants received a single dose of 2 mg ponesimod. TP2A, participants received placebo propranolol once daily from Day 1 to 19 plus an up‐titration regimen of ponesimod (2, 2, 3, 3, 4, 4, 5, 6, 7, 8, 9, 10, 10, 10 and 20 mg) from Day 5 to 19; TP2B, participants received 80 mg propranolol once daily from Day 1 to 19 plus the same up‐titration regimen of ponesimod as in TP2A. If the outcome of an adverse event is not resolved or not recovered, then the adverse event is persistent.

Abbreviations: AE, adverse event; AV, atrioventricular; bpm, beats per minute; HR, heart rate; QTcF, QT interval corrected for HR according to Fridericia's formula; TEAE, treatment‐emergent adverse event.

^a There were no reports of 2^nd or 3^rd degree AV block.

3.9. Cardiac‐Related Safety Analysis

A summary of abnormal HR values over time for the 12‐lead ECGs is shown in Table 4. There were no occurrences of HR < 45 bpm in TP1 (2 mg ponesimod), 2 (8.3%) in TP2A (placebo propranolol plus ponesimod) and 13 (56.5%) in TP2B (propranolol plus ponesimod). Bradycardia was observed between 2 and 4 h post‐dose in most participants; in 3 participants, this was at 8 h, and in 1 individual, this was at 10 h post‐dose. In TP2B, 3 participants had a HR < 40 bpm (38 bpm on Day 8, 39 bpm on Day 7, and 37 bpm on Day 7). Of these, 2 participants did not meet the pre‐dose HR criteria. All 3 participants discontinued the study as they met the discontinuation criterion of HR < 40 bpm on a single timepoint during treatment with ponesimod and propranolol in steady state. Two participants (both randomized to TP2B) exhibited bradycardia (HR < 45 bpm) during unscheduled assessments only and have not been accounted for in the 15 cases. On the recorded 12‐lead ECGs, 4 participants (7.7%) in the TP2B group experienced a maximum decrease in HR from pre‐dose baseline values of > 30 bpm, and 7 participants (13.5%) experienced a maximum decrease in HR from pre‐dose baseline values of > 20 bpm.

While no AEs of symptomatic bradycardia were reported, 7 participants reported symptoms possibly indicative of bradycardia during the up‐titration phase of ponesimod (including fatigue, somnolence, headache, dizziness, palpitations). Six were assigned to TP2B and 1 to TP2A.

3.10. Treatment‐Emergent ECG Abnormalities

A review of the summary statistics revealed that 28 participants (53.8%) in TP1, 20 (83.3%) in TP2A, and 22 (95.7%) in TP2B had at least one treatment‐emergent ECG abnormality. The total number of treatment‐emergent 12‐lead ECG abnormalities was higher in TP2B (360) compared to TP2A (162). These ECG abnormalities in TP2 were not clinically relevant. Seventy‐five treatment‐emergent 12‐lead ECG abnormalities were observed during TP1.

There is no significant difference in the occurrence of 1^st degree AV block in TP2A (25%) versus TP2B (30.4%) (Table 4). There were no reports of 2nd or 3rd degree AV block. During the study, 1 participant (1.9%) had a measured aggregate QTcF interval of > 500 msec.

4. Discussion

Although the gradual up‐titration regimen has been found to considerably attenuate the bradycardic effects during ponesimod initiation [17, 18], studies show that the initiation of S1PR modulators can cause a transient, dose‐dependent decrease in HR [12, 13]. Since beta blockers can also affect HR, it is suggested that the combined use of S1P1 modulators with beta blockers be carefully considered. Propranolol was selected for this study due to its established negative chronotropic effects [21], frequent clinical use for tremors in the MS population [22], and favorable safety profile in healthy adults [23], unlike atenolol, which has been associated with serious bradycardic events in a prior study with ponesimod without the gradual up‐titration regimen [11]. This drug–drug interaction study was designed to evaluate the PK, PD, and safety of the concomitant administration of ponesimod with propranolol in healthy adults.

PK results show that there was no clinically relevant drug–drug interaction between ponesimod and propranolol. The median ponesimod t_max was achieved 4–55 h post‐dose in the absence or presence of propranolol. Similarly, based on geometric mean ratios, on Day 5 ponesimod C_max and AUC_24h were similar when ponesimod was administered both in the absence or presence of propranolol. Mean propranolol C_trough levels were comparable on Days 4, 5, and 19, indicating that steady state was achieved before Day 5. These PK findings are not only relevant to propranolol but may be extrapolated to other beta‐blockers. This is because ponesimod is neither an inhibitor nor an inducer of metabolizing enzymes and does not affect renal drug transporters [8]. Consequently, a PK‐based drug–drug interaction is not expected with other beta‐blockers.

The minimum value of mHR from Holter in different ponesimod treatments (single dose ponesimod, up‐titration with propranolol placebo, and up‐titration with propranolol steady state) is seen at 4 h post‐dose, which is earlier than or equal to the t_max of ponesimod, which was 5 h on Day 5 and 4 h on Day 19 of the study.

Following the administration of ponesimod alone in Treatment Period 1, a maximum decrease versus time‐matched baseline in the mean of the mean hourly HR of 8.4 bpm was observed at 3 h post‐dose on Day 1. A similar decrease was observed following the first dose of ponesimod administered on Day 5 during TP2A.

Overall, concomitant administration of ponesimod with propranolol resulted in an additive effect on HR as measured by Holter. The largest difference in ΔHR_max between the combination of ponesimod with propranolol compared to ponesimod alone occurred on Day 5 (1st dose of 2 mg ponesimod) and was −12.4 bpm. The difference between treatments decreased with subsequent doses, reaching −7.4 bpm on Day 19 (the first 20 mg dose after up‐titration). This is in line with the known tolerance to the ponesimod HR effect on repeated dosing due to internalization of the S1P receptor. The lowest value of the HR_nadir in the combination arm was observed on Day 7 (3rd day of the ponesimod up‐titration regimen) and was 48.9 bpm and increased to 54.1 bpm on Day 19. The transient HR‐lowering effect of ponesimod is mechanistically linked to its initial agonistic effect at the S1P1R. Activation of this receptor in atrial myocytes upon initial ponesimod administration leads to the opening of G‐protein‐coupled inwardly rectifying potassium channels, causing membrane hyperpolarization and a temporary reduction in excitability, leading to bradycardia [24]. Upon repeated dosing, S1P1Rs undergo internalization and desensitization, thereby reducing this negative chronotropic response (functional antagonism) [6]. This PD profile supports the importance of up‐titration during ponesimod initiation to mitigate early HR effects, particularly in individuals with low resting HR or those receiving concomitant beta‐blockers [16, 17].

No AEs of symptomatic bradycardia were reported with the maximum HR reduction (Day 7). Safety 12‐lead ECGs and Holter did not show any 2^nd degree or higher AV block or clinically significant sinus pause (> 3 s). During the up‐titration regimen of ponesimod, and as expected, a maximum mean decrease of approximately 50% from baseline in total lymphocytes was observed. The lymphocyte counts returned to near baseline values by Day 26 after stopping ponesimod dosing on Day 19 and were back to above normal values in all participants. No significant difference was observed in lymphocyte reductions between ponesimod given alone or in combination with propranolol.

PK‐PD simulations suggest that co‐administering propranolol to ponesimod given at the clinical dose at steady state (instead of giving propranolol at the start of the ponesimod titration—as done in this study) would result in reduced negative chronotropic effects than those observed in this study [16]. This is because, with ponesimod at steady state, the tolerance to the initial negative chronotropic effect of ponesimod is fully developed. This has already been demonstrated by Biswal et al. from the interaction study between Siponimod and propranolol [25]. While this study evaluated propranolol, the additive HR‐lowering effect observed may be extrapolated to other beta‐blockers, as all agents in this class exert negative chronotropic effects [26]. However, the magnitude of the PD interaction could vary depending on the beta‐blocker's cardioselectivity, lipophilicity, intrinsic sympathomimetic activity, and dosing.

A limitation of this study is that it lacks a true placebo group and a ponesimod‐only group. As there was no placebo group throughout the study, comparisons across days within a treatment group are confounded by not having a baseline without drug on the corresponding days. For the primary objective of the study, which is the assessment of the effect of the addition of ponesimod in participants receiving propranolol at steady state (TP2B), time‐matched comparisons were made to participants receiving propranolol plus placebo (TP2A); therefore, the lack of placebo‐alone or ponesimod‐alone groups was mitigated. A further potential limitation of this study is that it was conducted in healthy adults rather than in patients with relapsing multiple sclerosis (RMS), the intended treatment population for ponesimod. However, it is important to note that the transient effect of ponesimod on HR in patients with MS might be less pronounced than that in healthy adults due to the disease's impact on the autonomic nervous system [16, 17]. Consequently, this study provided a controlled evaluation of PK, PD, and cardiac safety, with HR effects observed in a more sensitive population (healthy adults) than the RMS population. Therefore, the HR effects observed in this study are fully representative of those that might be achieved in patients with RMS.

To safe‐guard participants in the study, resting HR was required to be ≥ 55 bpm on steady‐state propranolol on Day 4 before introducing ponesimod up‐titration on Day 5. Therefore, the study was not designed to assess the effect of the combination in participants with resting HR < 55 bpm. However, 4 participants had a pre‐dose HR on Day 5 of < 55 bpm on propranolol alone but were erroneously dosed with the ponesimod up‐titration regimen. Two of these participants had a HR < 40 bpm with the combination and were later discontinued from the study. If resting HR is < 55 bpm, temporary interruption of the beta‐blocker treatment may be needed prior to the initiation of ponesimod.

Although combined administration of ponesimod with propranolol had an additive effect on HR, this effect did not lead to related serious AEs or clinically significant ECG abnormalities with the uptitration regimen. In addition, no clinically relevant effects on BP parameters and no AEs attributable to hypotension were observed with ponesimod alone or in combination with propranolol. Results from this study suggest that resting HR should be considered when ponesimod is initiated in patients receiving treatment with a beta‐blocker, and that temporary interruption of beta‐blocker treatment may be needed in patients with resting HR < 55 bpm, due to the additive effects on lowering HR.

Author Contributions

S.O.‐M., T.H., I.P., E.V., F.R., J.A., J.N., M.v.N.‐V., A.V., J.J.P.‐R., and T.S. wrote the manuscript. S.O.‐M., I.P., J.J.P.‐R., J.N., A.V., M.v.N.‐V., and T.S. designed the research. S.O.‐M., I.P., J.J.P.‐R., J.N., A.V., F.R., M.v.N.‐V., and T.S. performed the research. S.O.‐M., T.S., A.V., T.H., E.V., F.R., J.N., and J.A. analyzed the data. All authors read and approved the final manuscript.

Conflicts of Interest

Italo Poggesi was an employee of Janssen‐Cilag Italy, Part of Johnson & Johnson at the time the study and may hold stock or stock options in Johnson & Johnson. Dr. Poggesi is currently employed at GSK, Italy. Tessa Hosman was an employee of Janssen Biologics B.V, Leiden, the Netherlands, Part of Johnson & Johnson at the time the study and may hold stock or stock options in Johnson & Johnson. Dr. Hosman is currently employed at Wilhelmina Kinderziekenhuis, Utrecht, Netherlands. Tatiana Sidorenko was an employee of Actelion Pharmaceuticals Ltd., Part of Johnson & Johnson and may hold stock or stock options in the company. Dr. Sidorenko is currently an independent consultant. All other authors are employees or contractors of Johnson & Johnson and may hold stock or stock options in Johnson & Johnson.

Supporting information

Data S1. cts70341‐sup‐0001‐DataS1.docx

Ouwerkerk‐Mahadevan S., Hosman T., Poggesi I., et al., “A Randomized Trial on the Combined Effect of Ponesimod and Propranolol on Heart Rate, Cardiac Safety, and Pharmacokinetics in Healthy Adults,” Clinical and Translational Science 18, no. 9 (2025): e70341, 10.1111/cts.70341.