This study was designed as a randomized, controlled, single-blinded exploratory study comparing groups administered different doses of sugammadex, and was conducted in a single tertiary hospital located in Seoul, Republic of Korea. The study adhered to the tenets of the Declaration of Helsinki (revised in 2013), and the protocol was approved by the Institutional Review Board of Seoul National University Hospital (no.: 2002-148-1105, approval date: 03/25/2020) and the Ministry of Food and Drug Safety of the Republic of Korea (no.: 32,828, approval date: 04/13/2020). The study was registered at http://clinicaltrials.gov (NCT04347486, principal investigator: Hee-Soo Kim, published date: 04/14/2020) and was conducted in accordance with the Good Clinical Practice guidelines of the International Council for Harmonization and the Declaration of Helsinki. Participants were recruited between April 2020 and December 2021.

Children aged between 2 and 17 years who were scheduled to undergo elective brain or spinal surgery under general anesthesia were enrolled. Written informed consent was obtained from the parent(s) of each participant, as well as from the participants older than 6 years of age. The exclusion criteria were a history of hypersensitivity to any anesthetic agent, presence of cardiovascular or urological disease, presence of renal or hepatic impairment, use of a neuromuscular blocking agent or drugs that can affect the action of neuromuscular blocking agents prior to anesthetic induction, a history of malignant hyperthermia, anticipation of massive hemorrhage during surgery, and the refusal of one or both parents or legal guardians to permit enrollment.

When the participants arrived at the operating room, non-invasive blood pressure, oxygen levels (determined using pulse oximetry), and heart activity (determined using electrocardiography) were monitored. Anesthesia was induced by intravenous (IV) injection of sodium thiopental or propofol. After loss of consciousness, anesthesia was maintained with the continuous IV infusion of an opioid and propofol. To monitor neuromuscular blockade, a ToFscan^® device (IDMED, Marseille, France) was attached to each participant’s unilateral ulnar nerve. Train-of-four stimulation, which applies four twitch stimulations with an intensity of 50 mA and frequency of 2 Hz, was commenced and repeated at 15-s intervals until the end of surgery. The responses were measured via acceleromyography and automatically recorded on a computer using a vital recorder (Lee and Jung, 2018). Thereafter, 1 mg/kg of rocuronium was administered IV, followed by arterial catheterization at an extremity for continuous monitoring of invasive blood pressure. On the reappearance of the second twitch (T₂) in response to the train-of-four stimulation, the study drug was administered IV.

Allocation of patients to treatment groups was determined based on a randomization table obtained from https://sealedenvelope.com/. Having enrolled participants, they were allocated to one of the following groups: a control group (0.03 mg/kg neostigmine) or groups treated with 2, 4, or 8 mg/kg sugammadex. The study groups were assembled by a single anesthesiologist. The participants and their parents were blinded to group allocation.

Nine arterial blood samples were collected for pharmacokinetic measurements: 2 min after rocuronium injection, and immediately prior to and at 2, 5, 15, 60, 120, 240, and 480 min post-administration of the study drug. For each sampling time point, the plasma concentrations of rocuronium and sugammadex were measured. In all groups, the concentration of sugammadex was not measured at the first time point and at all points in the control group. When sampling was not performed at the exact scheduled time, the actual sampling time was recorded (Choi et al., 2013).

At each pharmacokinetic sampling point, 1 mL of arterial blood was withdrawn and immediately stored in a sodium heparin tube (BD Vacutainer^® sodium heparin [N] 75 USP Units; Becton Dickinson Korea, Seoul, Korea). After centrifuging at 3,000 rpm (1,167 × g) for 10 min, the supernatant was stored in a sterile internal cryogenic vial (Cryotain™; SCILAB Korea, Seoul, Korea). The collected specimens were stored in a freezer below −70°C until analysis. The plasma concentrations of sugammadex and rocuronium were determined using liquid chromatography–tandem mass spectrometry. The assays were conducted in compliance with Good Laboratory Practice regulations. As this assay cannot be used to discriminate the sugammadex–rocuronium complex from the free forms of these drugs, all plasma concentrations were considered total plasma concentrations. The internal standards used for sugammadex and rocuronium were phenformin-d₅ and rivastigmine-d₄ (Toronto Research Chemicals, North York, Canada), respectively.

The assay methods for rocuronium and sugammadex were validated previously and partial validation was performed based on a risk-based approach (Farenc et al., 2001; de Zwart et al., 2011; Briggs et al., 2014; de Moraes et al., 2014). Calibration curves for rocuronium were established in the ranges of 100–10000 ng/mL (r = 0.9965) and 2–200 ng/mL (r = 0.9980). Accuracy was between −3.0% and −1.0%, −8.8 and −7.5 in the range of 100–10000 and 2–200 ng/mL, respectively. The corresponding precisions were 2.0%–10.0% and 2.8%–5.0%, respectively. Accuracy and precision of the diluted samples were −1.0% and 7.0%. Carryover effect was not detected.

The assay method for sugammadex was also validated. Calibration curve for sugammadex was established in the range of 0.1–100 μg/mL (r = 0.9981). Accuracy was between −2.7% and −1.0% and precision was between 1.7% and 6.2%. Accuracy and precision of the diluted samples were 6.0% and 4.7%, respectively. Carryover effect was not detected.

The pharmacokinetic parameters of sugammadex and rocuronium were calculated based on non-compartmental analysis via Phoenix WinNonlin (Version 8.1; Certara United States, Princeton, NJ, United States). The maximum plasma concentration (C_max) and the time to reach C_max (T_max) were obtained directly based on the observed values. The area under the plasma concentration-time curve (AUC) to the last measurable concentration (AUC_last) was calculated using the linear up-log down trapezoidal method. AUC extrapolated to infinity (AUC_inf) was calculated as the sum of AUC_last and the last measurable concentration divided by the elimination phase constant (λ_z), which was regressed on the log-transformed concentration. The terminal phase half-life (t_1/2) was calculated as the natural logarithm of 2 divided by λ_z. The volume of distribution and clearance were calculated as dose/(λ_z × AUC_inf) and dose/AUC_inf, respectively. Dose-normalized C_max, AUC_last, and AUC_inf values were calculated as each parameter divided by the administered dose. We compared the obtained pharmacokinetic parameters with those of our previous study for reversal of intense neuromuscular blockade (Ji et al., 2023).

Responses to train-of-four stimulation were continuously monitored at 15-s intervals. When the number of twitches was fewer than four, only the number of twitches was recorded. After the appearance of the fourth twitch, the ratio of the fourth twitch to the first twitch (T₄/T₁) was recorded, as this is considered to represent receptor occupancy by rocuronium. As a T₄/T₁ ratio ≥90% is considered sufficient for extubation, the time from administration of the study drug to recovery of a T₄/T₁ ratio ≥90% was taken to be the primary pharmacodynamic outcome.

Electrocardiogram, mean blood pressure, pulse oximetry, and body temperature were monitored until 24 h after the end of surgery. We monitored the occurrence of any changes >30% of the baseline in heart rate or mean blood pressure, pulse oximetry of <92%, changes in body temperature to above 38.3°C or below 35.5°C, and other complications, including nausea, vomiting, urticaria, and any anaphylactic reactions.

Given the exploratory nature of the study, sample size was calculated empirically. We allotted eight children to each group on the basis of previously conducted pediatric pharmacokinetic studies for sugammadex that had assessed group sizes of 4–10 children (Plaud et al., 2009; Voss et al., 2022). Normality was assessed using the Shapiro–Wilk test. Baseline characteristics and recovery times of patients in the different study groups were compared using analysis of variance for normally distributed data, and the Kruskal–Wallis test with post hoc analysis using the Mann–Whitney U test with Bonferroni correction for non-normally distributed data. Analyses were performed using R ver. 4.0.3 (The R Foundation, http://r-project.org), with the acceptable alpha error rate set at 0.05.

For non-compartmental analysis, subgroup analysis was done for C_max, AUC_last, and AUC_inf.: parameters of rocuronium were compared between each dose group and parameters of sugammadex were compared between age group classified according to a recent pharmacodynamics study (Voss et al., 2022) using the Kruskal–Wallis test. Non-parametric post hoc evaluation was performed with the Dwass, Steel, and Critchlow–Fligner procedures. Parameters from non-compartmental analyses of this study and our previous study (Ji et al., 2023) were compared for both drugs using Student’s t-test or Wilcoxon rank sum test. These analyses were done via SAS software (version 9.3; SAS Institute, Inc., Cary, NC, United States).

Average (±Standard deviation) time interval between rocuronium administration and sugammadex administration was 48.7 (±11.4) minutes. We found that the time [median (interquartile range)] from administration of sugammadex to achieving a T₄/T₁ ratio of 0.9 in response to train-of-four stimulation differed significantly among the groups (p < 0.001). For the control group, we recorded a recovery time of 7.7 (5.3–21.0) min, whereas that for participants in the 2, 4, and 8 mg/kg sugammadex groups was 1.3 (1.0–1.9), 0.9 (0.8–0.9), and 0.6 (0.4–1.0) min, respectively. In the post hoc analysis, we detected a significant difference between the control group and each of the groups treated with sugammadex (p = 0.002, 0.001, and 0.001 for the groups treated with 2, 4, and 8 mg/kg sugammadex, respectively). However, there were no significant differences among the groups treated with sugammadex. Figure 3 shows box plots of the recovery time for each group.

Plasma sugammadex concentrations were found to be characterized by a biexponential decline, with a terminal half-life of 1.7–1.8 h. The systemic exposure to sugammadex was established to be proportional to the administered dose. Clearance (0.07 L/min) was consistent across dose levels and the volume of distribution ranged between 8.8 and 11.9 L (Figure 4; Table 2).

Age group was found to be significantly associated with pharmacokinetics parameters, with reductions in dose-normalized C_max, AUC_last, and AUC_inf being observed with increasing age. In contrast, both the clearance and volume of distribution were observed to be higher in older children, whereas the elimination half-life was consistent among all age groups. Subgroup analysis of data from our previous study (Ji et al., 2023) based on age group showed similar results (Table 3).

Plasma rocuronium concentration was also characterized by a biexponential decline, with a half-life of 1.6–1.7 h. Systemic exposure to rocuronium was comparable between the control and sugammadex groups, with clearance and volume values ranging between 0.11 and 0.16 L/min and 15.9 and 21.1 L, respectively (Figure 5; Table 2). Figure 3 shows the molar concentrations of sugammadex and rocuronium, along with the T₄/T₁ ratio of the individual participants.

There were no statistically significant difference between this study and our previous study in any of the parameters for sugammadex. For rocuronium, C_max and AUCs were similar, while t _1/2 and V_z tended to show larger values in this study (Table 2).

During the period between administration of the study drug and 24 h after the end of surgery, 20 of the 29 participants experienced a range of adverse side effects, including fever, hypothermia, hypotension, hypertension, tachycardia, bradycardia, headache, nausea, and vomiting. Among these, two hypotensive events were considered to be associated with the use of sugammadex, although these were self-limiting and no subsequent intervention was necessary. Other adverse events were considered to be surgery-related rather than attributable to the use of a neuromuscular reversal agent. Detailed profiles of the adverse events are presented in Table 4.