Mass Balance and Metabolism of ¹⁴C‐Atogepant in Healthy Male Participants: Findings of a Phase 1 Clinical Trial

ABSTRACT

Atogepant, a calcitonin gene‐related peptide (CGRP) receptor antagonist, is approved for the preventive treatment of migraine in adults. This open‐label, phase 1 study examined radiolabeled atogepant (¹⁴C‐atogepant) metabolism, elimination, and mass balance after a single dose (50 mg, ~200 μCi) in healthy adult males. Blood, urine, and feces were collected ≤ 28 days for atogepant, metabolite, and radioactivity level measurement. Six participants were enrolled (mean age: 33.3 years; body mass index: 28.8 kg/m²); five completed the study (one withdrew on Day 7 for family emergency). For atogepant, median time to maximum concentration (T _max) was 1 h and mean terminal elimination half‐life (T _1/2) was 18.5 h; for total radioactivity, T _max was 1.5 h and mean T _1/2 was 11.6 h. Unchanged atogepant was the major circulating species in plasma (mean atogepant/total radioactivity area under the curve ratio: ~0.75). In the 14 days after dosing, ~81% and ~8% of the radioactive dose were recovered in feces and urine, respectively. In feces, parent drug (unabsorbed drug, biliary excretion, and/or intestinal secretion) accounted for 42% of administered radioactivity. At least 11 metabolites were identified in feces, and each represented < 10% of administered radioactivity. In plasma, atogepant and metabolite M23 were the only radiometric peaks detected. Metabolite M23, tentatively characterized as dioxygenated methylated glucuronide of atogepant, represented ~15% of plasma radioactivity and was short‐lived. There were no treatment‐emergent adverse events or clinically meaningful changes in laboratory values, vital signs, or the electrocardiogram.

Study Highlights.

• What is the current knowledge on the topic?

  • ○
    Atogepant pharmacokinetics have been well characterized, but full metabolic characterization and mass balance have not yet been reported.
  • ○
    An improved understanding of atogepant's metabolic/elimination pathways and metabolites is of particular importance, as first generation gepants had liver toxicity concerns likely related to hepatotoxic metabolites.
  • ○
    Second‐generation gepants, including atogepant, lack hepatotoxic chemical functionalities, but the importance of metabolite evaluation remains.
• What question did this study address?

  • ○
    Here, we report the full metabolic profile, elimination, and mass balance of radiolabeled atogepant (¹⁴C‐atogepant) based on a single 50 mg oral dose (~200 μCi) in healthy male participants.
• What does this study add to our knowledge?

  • ○
    Circulating radioactivity consisted mainly of atogepant (parent drug, ~75%) and metabolite M23 (dioxygenated methylated glucuronide of atogepant, ~15%), which was short‐lived and likely pharmacologically inactive.
  • ○
    No unique metabolites were observed in humans, and based on in vivo rat and monkey metabolism studies, there are no disproportionate human metabolites.
  • ○
    Feces was the predominant route of excretion (~81%), but some urinary excretion did occur (~8%).
  • ○
    There were no treatment‐emergent adverse events and no clinically meaningful changes in laboratory values, vital signs, or the electrocardiogram.
• How might this change clinical pharmacology or translational science?

  • ○
    Study findings support the lack of atogepant hepatotoxicity and justify the decision not to conduct a dedicated renal impairment pharmacokinetic study.

1. Introduction

Atogepant (QULIPTA/AQUIPTA), an oral calcitonin gene‐related peptide (CGRP) receptor antagonist indicated for migraine prophylaxis in adults [1], can safely and effectively reduce the number of migraine days in patients with episodic [2, 3] and chronic (≥ 15 headache days/month) [4] migraine. Preclinical studies demonstrated that atogepant has a high affinity, potency, and selectivity for the human CGRP receptor [5]. Atogepant is a substrate of organic anion transporting polypeptide (OATP), P‐glycoprotein (P‐gp), and breast cancer resistance protein (BCRP) transporters and is mainly eliminated by the liver via cytochrome P450 (CYP) 3A4 metabolism [1, 6]. Significant drug–drug interactions (DDIs) were observed in healthy phase 1 study participants. Steady–state itraconazole (strong CYP3A4 inhibitor) and single‐dose rifampin (OATP inhibitor) co‐administration significantly increased atogepant exposure [7]. Conversely, multiple‐dose rifampin (strong CYP3A4 and P‐gp inducer) co‐administration significantly decreased atogepant exposure [1, 7]. However, no clinically meaningful DDIs were observed between atogepant (single 60 mg dose) and other commonly used migraine treatments including acetaminophen (1000 mg) [8], naproxen (500 mg) [8], sumatriptan (100 mg) [9], and ubrogepant (100 mg) [10]. This was also true for atogepant co‐administration with topiramate (100 mg twice daily), a mild CYP3A4 inducer [11], and quinidine, a P‐gp inhibitor [12].

Atogepant pharmacokinetics (PK) have been well characterized, but full metabolic characterization and mass balance have not yet been reported. Here, we report the full metabolic profile, elimination, and mass balance of radiolabeled atogepant (¹⁴C‐atogepant) based on a single 50 mg oral dose (~200 μCi) in healthy male participants.

2. Methods

This phase 1, single‐center, open‐label, single‐dose study examined the mass balance, plasma concentration‐time profile, metabolism, and safety of a single dose of ¹⁴C‐atogepant. The study protocol, informed consent forms, and recruitment materials were reviewed and approved by the Chesapeake Institutional Review Board (Columbia, MD) prior to participant enrollment. All participants provided written informed consent to participate, and all study conduct was in accordance with the International Conference for Harmonization guidelines, applicable regulations, and the Declaration of Helsinki of 1975 (as revised in 1983). The current mass balance study was conducted in accordance with US regulatory guidelines [13].

2.1. Participants

This study enrolled 6 healthy male participants aged 19–55 years. All participants were non‐smokers, had a body mass index (BMI) of 18–32 kg/m², and had a sitting pulse rate of 45–100 bpm at Screening. Participants who were not sterilized for ≥ 1 year agreed to use effective contraception and not to impregnate their partner during the study. Key exclusion criteria included clinically significant abnormal blood pressure, abnormal electrocardiogram (ECG), pre‐dose HIV (type 1 or 2) or hepatitis (B or C) positivity, history of alcohol/substance abuse ≤ 5 years, concomitant medication use ≤ 14 days of Screening (including over‐the‐counter), clinically significant disease, or known hypersensitivity to atogepant or another CGRP receptor antagonist.

2.2. Study Design

2.2.1. Study Objectives and Outcome Measures

Study primary objectives were to investigate routes of elimination and mass balance of ¹⁴C‐atogeptant, assess atogepant and radioactivity concentration‐time profiles, and identify major metabolites in biological specimens after oral administration of a single 50 mg (~200 μCi) dose of ¹⁴C‐atogepant. Secondary objective was to assess safety/tolerability. Primary outcome measures included plasma PK parameters, urine PK parameters, mass balance, and metabolite profiling in plasma, urine, and feces. Atogepant safety/tolerability was assessed via adverse events (AEs), clinical labs, vital signs, ECG, and physical examination.

2.2.2. Study Procedures and Examinations

All clinical assessments and sample collections were performed at Celerion (Lincoln, NE). Study duration was ≤ 31 days, with Day 1 defined as the day of atogepant dosing (Figure 1). Briefly, Screening occurred ≤ 21 days before dosing and included medical/surgical history (including medication use), physical exam, vital signs assessment, 12‐lead ECG, blood and urine collection (for hematology, coagulation, chemistry, urinalysis, serology, drug abuse screen), and AE assessment. Pre‐dose (Day −1) examination included AE assessment, concomitant medication assessment, safety laboratory testing (hematology, coagulation, and chemistry), urinalysis, drug abuse screen, and fecal sample. A single 50 mg ¹⁴C‐atogepant dose (~200 μCi) was orally administered on Day 1 after a ≥ 10‐h fast, with the last potential PK sampling planned on Day 29 and at Follow‐up (Day 30). ¹⁴C‐atogepant was dosed in 2.5 mL of solution (20 mg/mL in PEG400 containing Ora‐Sweet) and followed by a 10 mL placebo wash. Further dosing solution detail is provided in the Supplement.

FIGURE 1.

Study design. A single dose of 50 mg ¹⁴C‐atogepant was orally administered on Day 1 after ≥ 10 h fast. Last PK sampling was done on Day 8 or until ≥ 1 discharge (d/c) criterion was met (> 90% administered total radioactivity excreted or radioactivity in urine and feces combined < 1% of total administered dose on two consecutive days).

Study assessments occurred on Days 1–8 and included AE assessment and PK blood, urine, and fecal sample collection. Day 1 examinations also included ECG and vital sign assessment; Day 2 examinations also included safety laboratory testing and vital sign assessment. Participants were housed at the study center between Day −1 and Day 8 and were discharged on or after Day 8 if ≥ 1 of the following were met: > 90% of the administered radioactive dose had been excreted or radioactivity was < 1% (urine and feces combined) on 2 consecutive days when both urine and fecal samples were collected. In participants who did not meet a discharge criterion, blood sampling continued at 3‐day intervals and urine and fecal sampling continued at 24‐h intervals, respectively, until ≥ 1 criterion was met or Day 29 was reached. Beginning 14 days prior to Day 1 through End‐of‐Study, participants avoided grapefruit‐containing products and vegetables in the mustard green family. Participants abstained from caffeine and alcohol‐containing products for 48‐ and 72‐h prior to Day 1, respectively, through End‐of‐Study. During study housing, participants were provided daily meals and snacks at the same times.

Combustion and liquid scintillation counting (LSC) was performed at the Celerion ADME Laboratory on one aliquot of each sample for plasma, urine, and fecal total radioactivity measurement. Another aliquot was used to quantify plasma and urine atogepant concentrations using validated liquid chromatography‐mass spectrometry/mass spectrometry (LC–MS/MS) assays (Keystone Bioanalytical Inc., North Wales, PA) [11]. A third aliquot of each sample was used for plasma, urine, and fecal metabolite profiling via high‐resolution mass spectrometry and radiometric detection at the study sponsor's drug metabolism and pharmacokinetics laboratory (AbbVie DMPK Laboratory, Irvine, CA). Further detail is provided below.

2.3. Pharmacokinetic Assessments

Atogepant concentration measurements were conducted by Keystone Bioanalytical Inc. All samples were frozen prior to shipping (−20°C or less), received frozen and in good condition, and stored at −70°C until analysis.

2.3.1. Blood and Plasma

Blood samples for drug concentration measurement were collected on Day 1 at 0 h (pre‐dose), 20 and 40 min post‐dose, and 1, 1.5, 2, 3, 4, 6, 12, 24, 36, 48, 72, 96, 120, 144, and 168 h post‐dose. If ≥ 1 discharge criterion was not met at 168 h (Day 8), PK blood samples were obtained in 72‐h intervals until ≥ 1 criterion was met or until Day 29 (End‐of‐Study, 28 days post‐dose). Within 30 min of collection, samples were centrifuged at ≥ 2500 g for 10 min at ~4°C. Plasma was then harvested and transferred to 3 prechilled, labeled polypropylene tubes and stored upright at −70°C or colder. A validated LC–MS/MS assay was used for atogepant quantification in K₂EDTA human plasma (range of quantification: 1–1000 ng/mL). Methodological detail and measurement acceptance criteria are provided in Table S1.

2.3.2. Urine

Urine was collected for PK and metabolite measurements pre‐dose and at the following post‐dose intervals: 0–4 h, 4–8 h, 8–12 h, 12–24 h, 24–48 h, 48–72 h, 72–96 h, 96–120 h, 120–144 h, 144–168 h, and every 24 h thereafter until ≥ 1 discharge criterion was met or End‐of‐Study. Urine samples from each participant were combined at the end of each collection interval, thoroughly mixed, and weighed. All samples were kept refrigerated during collection intervals, and collected urine was treated with Tween20 (surfactant) solution for a final concentration of 0.25% Tween20 in urine. Three urine aliquots were taken from each collection interval, placed in labeled polypropylene storage tubes, and stored upright at −20°C or colder. Total radioactivity in urine samples was determined within 24 h of sample collection until discharge criteria were met. A validated LC–MS/MS assay was used for atogepant quantification in urine (range of quantification: 1–1000 ng/mL). Methodological detail and measurement acceptance criteria are provided in Table S1.

2.3.3. Feces

Pre‐dose fecal samples were obtained ≤ 48 h before ¹⁴C‐atogepant dosing and were used to determine background radioactivity. After dosing, total fecal radioactivity through 120 h was measured and then every 24 h thereafter until ≥ 1 discharge criterion was met or the End‐of‐Study. All fecal samples were collected during confinement. At the end of each collection interval, samples were weighed and homogenized. The homogenate was once again weighed, and a single aliquot (~25 mL) was taken for radioactive measurements. Two additional aliquots were collected; one frozen (−20°C or colder) and sent for metabolite profiling, the other stored (−20°C or colder) at the study site as back‐up.

2.4. Atogepant Metabolite Profiling/Characterization

2.4.1. Plasma

Samples with sufficient radioactivity were analyzed for metabolite profiling/characterization. All chemicals used in the analysis were reagent grade or better; all solvents HPLC grade or better. Diclofenac acyl‐glucuronide, estriol 3‐glucuronide, and axitinib N‐glucuronide were purchased from Cayman Chemical (Ann Arbor, MI); hydroxylamine solution (50% solution in water) from Sigma‐Aldrich (Saint Louis, MO). Limits of quantification (LOQs) for metabolites in profiling radiochromatograms were determined for each sample. LOQ was defined as 5‐fold higher than background radiochromatogram counts. Further detail on the metabolite profiling/characterization is provided in Table S2.

Plasma samples of the 0.33–36 h timepoints from each participant were pooled using the Hamilton (area under the curve [AUC]) pooling scheme [14], except for one whose samples were pooled from 0.33–24 h (36‐h sample omitted due to low signal intensity). Pooled plasma samples (4 mL) were extracted twice with 12 mL of acetonitrile, and extracts were evaporated to dryness and reconstituted in 50% acetonitrile in water prior to analysis. Individual plasma samples of all 6 participants that were collected through 312 h post‐dose were also analyzed using an LC–MS/MS method for metabolite M23 (Table S3). Briefly, 0.1 mL plasma aliquots were extracted with 0.3 mL acetonitrile. Samples were vortexed and centrifuged, 0.15 mL aliquots of supernatant were transferred to vials in 96‐well format, and 0.15 mL of water was added to each well prior to LC–MS/MS analysis. Based on Hamilton (AUC) pooling scheme, the percent of radioactive exposure (AUC_0–t) for each metabolite in plasma was calculated (Equation 1):

$$\%\text{Metabolite}{\mathrm{AUC}}_{0-\mathrm{t}}=\frac{{\left(\text{Metabolite peak area}\right)}_{0-\mathrm{t}}}{{\left(\text{Total peak area}\right)}_{0-\mathrm{t}}}\times 100$$ (1)

where 0–t indicates pooled sample from time zero to time of last analyzed sample; peak area obtained from HPLC‐RAD radiochromatograms. Extraction efficiency was > 93%, as determined using thrice acetonitrile extraction on representative samples from a single participant.

2.4.2. Metabolite M23 Characterization

Further characterization of the only major metabolite identified (M23) in plasma was conducted. Full methodological details are provided in Table S3. Briefly, plasma samples at the 4‐h post‐dose timepoint were obtained following a single 300 mg dose of atogepant administered as part of a separate study [15]. High performance liquid chromatography coupled with high‐resolution mass spectrometry (HPLC‐HRMS) was used to detect hydroxamic acid derivatives of M23 (Table S4). Samples were analyzed using extracted ion scan (XIC) in positive electrospray mode at specific m/z ranges based on the glucuronide conjugate and the molecular weight of the expected hydroxamic acid derivatives. Aliquots (0.1 mL) of 4‐h post‐dose plasma samples were treated with 0.3 mL acetonitrile. After being vortexed and centrifuged, 0.2 mL aliquots of supernatants were treated with equal volumes of 50% aqueous hydroxylamine solution. Additional 0.1 mL aliquots of supernatants were fortified with an equal volume of water and served as stability control samples. After incubating at room temperature for 0, 47, 94, and 1541 min, aliquots were analyzed by HPLC coupled with HPLC‐HRMS. Positive and negative controls were analyzed for further confidence in experimental results. Diclofenac acyl‐glucuronide was used as a positive control, while estriol 3‐glucuronide (O‐glucuronide) and axitinib N‐glucuronide were used as negative controls. An untreated blank human plasma aliquot was also included to verify the absence of background peaks. Control compounds were added to blank human plasma at 0.1 mM, and reactions were conducted as described above for M23 plasma sample analysis with the exception of incubation times. Incubations for controls were done at room temperature but were conducted for up to 60 min (diclofenac acyl‐glucuronide), 120 min (estriol 3‐glucuronide and axitinib N‐glucuronide), and overnight (blank human plasma). After incubation, aliquots were analyzed by HPLC‐HRMS.

2.4.3. Urine

Urine samples from each participant of 0–4, 4–8, 8–12, and 12–24 h were pooled in proportion to their sample weight and analyzed for metabolite profiling/characterization. Each pooled urine sample (one per participant) was centrifuged, and supernatants were transferred to vials. The percentage of radioactive dose for each metabolite was calculated (Equation 2):

$$\%\text{Metabolite radioactive dose}=\frac{\%\text{Radioactive dose}\times \%\text{Peak area}}{100}$$ (2)

where %radioactive dose was total sample radioactivity divided by total‐dose radioactivity; %peak area was the metabolite: total peak area (as determined on HPLC‐radiographic detector [RAD] radiochromatograms). Column recovery was examined using the 0–4 h pooled urine sample from one participant and was determined to be 93.1% for the participant and 98.4% for the ¹⁴C‐atogepant standard (acceptable range: 80%–120%).

2.4.4. Feces

Fecal samples were homogenized with water prior to the shipping of aliquots. Fecal samples with ≥ 2% of the radioactive dose through 312 h timepoints were selected for each participant. Aliquots (~5 g) of fecal homogenate were weighted and extracted two times with 15 mL of acetonitrile, then again with 5 mL of acetonitrile. Resulting supernatants were combined, evaporated to dryness, and reconstituted in 50% acetonitrile in water prior to analysis. The % radioactive dose for each metabolite was calculated as described above for urine (Equation 2). Extraction efficiency after thrice acetonitrile extraction was ~72%.

2.5. Atogepant Safety

The safety population included all participants who received study drug. Safety was assessed via AE reporting, physical exam, ECG, vital sign, and laboratory parameter monitoring. AEs were coded using MedDRA terminology (version 20.1 or newer). Treatment‐emergent AEs (TEAEs) were defined as any new or worsening AE that occurred ≤ 30 days following study drug dosing.

2.6. Data Analysis

Atogepant and radioactivity plasma concentration measurements were used for PK parameter evaluation including atogepant and radioactivity exposures over all time (AUC_0–∞), AUC_0–t, C _max, T _max, terminal elimination rate constant (Kel), terminal elimination half‐life (T _1/2), apparent total clearance from plasma (CL/F), apparent volume of distribution during the terminal phase (V_Z/F), and atogepant:total plasma radioactivity ratio. Principal plasma PK parameters were calculated using noncompartmental analysis (Phoenix WinNonlin [version 8.0]; Certara, Radnor, PA). Actual sampling times were used in PK calculations. Atogepant plasma concentrations below the LOQ (BLQ; < 1 ng/mL) were treated as zero for PK analyses. Sample radioactivity that was BLQ (plasma: < 73 dpm/mL, urine: < 89 dpm/mL, feces: < 81 dpm/g) was assigned a zero value for analyses. All post‐dose timepoints for which no sample was collected were treated as missing and no value was imputed.

Urine atogepant and radioactivity concentrations were used to calculate total urine atogepant excretion (urine Ae_0–t), renal clearance (CL_R), and proportion of dosed radioactivity excreted in urine. Urine atogepant concentrations measured as BLQ (< 1 ng/mL) were treated as zero for PK analyses. Fecal atogepant and radioactivity concentrations were used to calculate total fecal atogepant excretion (fecal Ae_0–t) and proportion of dosed radioactivity excreted in feces. The mass balance for atogepant was performed by the Celerion ADME laboratory using total radioactivity recovered in urine and feces.

2.7. Statistical Analysis

Sample size calculations were not performed for this Phase 1 study. However, it is common practice to include six participants in human mass balance studies [16] and the study population size was suitable to achieve study objectives. The safety population included all participants who received atogepant; the PK population included all who received atogepant and had evaluable PK data.

Descriptive statistics were used to describe demographic, vital sign, clinical laboratory, PK, and mass balance parameters. Data are presented as mean ± standard deviation (SD) or median (range), as appropriate. However, if extrapolated AUC was > 20%, AUC_0–∞, CL/F, and Vz/F were listed by participant and excluded from descriptive statistics.

3. Results

3.1. Study Population

Six healthy male participants (mean age: 33.3 ± 7.9 years; race: 66.7% white, 33.3% Black/African‐American; ethnicity: 50.0% Hispanic/Latino; weight: 87.9 ± 10.9 kg; BMI: 28.8 ± 2.21 kg/m²) were enrolled in the study and received a single 50 mg dose (~200 μCi) of oral ¹⁴C‐atogepant. Five participants completed the study. The remaining participant withdrew from the study on Day 7 (family emergency), completed Early Termination assessments, and agreed to return for the Follow‐up visit. All participants had evaluable PK data and were included in both PK and safety populations. All participants had plasma, urine, and fecal samples collected beyond 168 h post‐dose, except for the one who withdrew early, but PK samples were collected through 144 h post‐dose.

3.2. Safety

No TEAEs were reported and no meaningful changes in clinical laboratory values, vital signs, or ECG traces were noted. No serious TEAEs, AE‐related study discontinuations, or deaths occurred. No cases of Hy's Law occurred.

3.3. Pharmacokinetics

Atogepant plasma concentrations were measurable up to 120 h after dosing; radioactivity was quantifiable up to 48 h after dosing. Plasma concentration and measured radioactivity over time after a single 50 mg (~200 μCi) ¹⁴C‐atogepant dose are shown in Figure 2. Key PK parameters are summarized in Table 1. Briefly, mean peak atogepant plasma concentration (C _max) was 367 ± 189 ng/mL, occurring at a median (range) of 1 h (0.98–2.98) after dose administration. In close agreement, mean peak radioactivity was 349 ± 82.6 ng‐Eq/mL, occurring at a median of 1.5 h (1–2.98) after dose administration. Mean atogepant AUC_0–t and AUC_0–∞ were 2180 ± 960 and 2220 ± 966 ng·h/mL, respectively; radioactivity AUC_0–t and AUC_0–∞ were 2710 ± 759 and 2970 ± 779 ng‐Eq·h/mL, respectively. Circulating atogepant accounted for ~75% of measured plasma radioactivity (mean atogepant: total radioactivity ratio: AUC_0–t: 0.776 ± 0.167, AUC_0–∞: 0.728 ± 0.172). Mean T _1/2 was longer for atogepant vs. total radioactivity (18.5 ± 11.9 vs. 11.6 ± 6.84 h), likely because atogepant was measurable for a longer period of time following dosing (120 vs. 48 h).

FIGURE 2.

Mean measured atogepant concentration and radioactivity in plasma following a single 50 mg dose of oral ¹⁴C‐atogepant. Both linear and log‐scale concentration profiles are shown. Atogepant and radioactivity were detectable up to 120 and 48 h, respectively. Error bars represent standard deviation.

TABLE 1.

Key atogepant and total radioactivity pharmacokinetic parameters in healthy male participants receiving a single 50 mg dose of ¹⁴C‐atogepant.

	Atogepant (N = 6)	Total radioactivity (N = 6)	Atogepant/Total radioactivity ratio (N = 6)
C max, ng/mL or ng‐Eq/mL, mean ± SD	367 ± 189	349 ± 82.6	0.997 ± 0.286
T max, hours, median (range)	1 (0.98–2.98)	1.5 (1–2.98)	—
AUC0–t, ng·h/mL or ng‐Eq/mL, mean ± SD	2180 ± 960	2710 ± 759	0.776 ± 0.167
AUC0–∞, ng·h/mL or ng‐Eq·h/mL, mean ± SD	2220 ± 966	2970 ± 779	0.728 ± 0.172
T 1/2, hours, mean ± SD	18.5 ± 11.9	11.6 ± 6.84	—
Total proportion excreted, mean ± SD	Not measured	88.5% ± 2.40%	Not measured
Urine	4.84% ± 1.63%	7.86% ± 1.83%	0.605 ± 0.109
Feces	Not measured	80.6% ± 3.35%	Not measured
CLR, L/h, mean ± SD	1.18 ± 0.33	Not calculated	—

Abbreviations: AUC_0–∞, drug exposure from dosing through all time; AUC_0–t, drug exposure from dosing through End‐of‐Study (Day 29); CL_R, renal clearance; C _max, maximum concentration; SD, standard deviation; T _1/2, terminal elimination half‐life; T _max, time to reach maximum concentration after dosing.

Approximately 5% of the administered dose was recovered as parent drug (atogepant) in urine up to 360 h post‐dose (mean ± SD Ae_0–t: 2.42 ± 0.82 mg; Table 1). Mean (±SD) atogepant CL_R was 1.18 ± 0.33 L/h.

3.4. Atogepant Mass Balance and Metabolic Profiling

Total radioactivity recovery through the last specimen collection was high (mean: feces: 80.6% ± 3.35%, urine: 7.86% ± 1.83%; Table 1, Figure 3). Therefore, ~89% of radioactivity was recovered, with excretion primarily via the feces.

FIGURE 3.

Cumulative recovered radioactivity in the urine, feces, and overall. All participants were healthy males (N = 6) and were administered a single 50 mg dose of ¹⁴C‐atogepant (~200 μCi). Time zero measurements (pre‐dose) were assumed to have zero concentration and were added for illustrative purposes.

The metabolites of ¹⁴C‐atogepant (all primary from the parent except M23, which is a secondary glucuronide of the primary oxidized metabolite) were successfully profiled and characterized (Table 2, Figure 4). In plasma, two radiometric peaks were identified; one due to parent drug (atogepant), one from a metabolite tentatively characterized as dioxygenated methylated glucuronide of atogepant (Metabolite M23; Figure 5). M23 accounted for ~15% of plasma radioactivity exposure (AUC_0–t: 15.3% ± 9.2%; Table 2) and was not long‐lasting (not detected in plasma after 96 h, similar parallel plasma concentration‐time profile as atogepant in the elimination phase [Figure S1]). Urine was a minor elimination pathway (8% of total radioactivity recovery), as detailed above. The only radiometric peak of interest in urine (≥ 1% of urine radioactive recovery) was that of atogepant, which accounted for ~66% of urine radioactive recovery (5% of total radioactivity dose; Table 2). Feces was the major elimination pathway (81% of administered radioactivity), and the parent drug accounted for ~52% of fecal radioactivity recovery (42% of total radioactivity dose) due to unabsorbed drug, biliary excretion, and/or intestinal secretion. Nevertheless, at least 11 metabolites were detected in feces (≥ 1% of fecal radioactivity), but each accounted for < 10% of the total radioactivity dose (Table 2).

TABLE 2.

Atogepant and identified metabolites in plasma, urine, and feces following a single oral administration of ¹⁴C‐atogepant to healthy male participants (N = 6).

	RT min	MW g/mol	Proposed structure	Plasma %AUC0–t	Urine %DE	Feces %DE
				Mean ± SD	Mean ± SD	Mean ± SD
Parent	33.6	603.5	Atogepant (P: C29H23F6N5O3)	84.7 ± 9.2	5.21 ± 1.69	42.2 ± 4.2
M3/M4 a	22.2	651	Trioxygenated atogepant (P + 3O)	ND	ND	5.32 ± 1.04
M5	23.2	651	Trioxygenated atogepant (P + 3O)	ND	ND	1.29 ± 0.81
M6/M7 a	25.2	635	Dioxygenated atogepant (P + 2O)	ND	ND	1.94 ± 0.59
M20	26.4	621	Dioxygenated demethylated atogepant (P + 2O‐CH2)	ND	ND	1.39 ± 0.51
M21	27.4	621	Dioxygenated demethylated atogepant (P + 2O‐CH2)	ND	ND	3.19 ± 1.04
M22	28.0	607	Dioxygenated de‐ethylated atogepant (P + 2O‐C2H4)	ND	ND	2.19 ± 0.48
M11	31.3	607	Monooxygenated atogepant (P + O)	ND	ND	1.66 ± 0.57
M18	31.9	619	Monooxygenated atogepant (P + O)	ND	ND	1.38 ± 0.34
M23	32.2	619	Dioxygenated methylated glucuronide of atogepant b	15.3 ± 9.2	ND (±~0.3)	ND
M2	32.8	825	Monooxygenated atogepant (P + O)	ND	ND	8.99 ± 1.84
R1	34.0–35.5	NA	Unknown	ND	ND	1.23 ± 0.35

Note: Each metabolite (M2–M23) represented a single detected metabolite based on mass spectrometry data. N = 6 for plasma, urine, and feces analyses.

Abbreviations: AUC_0–t, area under the radioactivity‐time curve from time zero to last measurable concentration (0–36 h except 0–24 h for one participant with low 36‐h intensity); DE, dose excreted (urine: 0–24 h [n = 6], feces: 0–312 h [n = 1], 0–120 h [n = 3], 0–96 h [n = 2]); MW, molecular weight; NA, not applicable; ND, not detected or < 1% of dose or AUC_0–t; R1, region with multiple coeluting metabolites; RT, retention time; SD, standard deviation.

^a Metabolite M3/M4 and M6/M7 coeluted in radiometric chromatograms.

^b (P + 2O + CH₂ + Gluc).

FIGURE 4.

Proposed metabolic scheme of atogepant following a single oral administration of ¹⁴C‐atogepant (50 mg) in healthy male participants. ‘*’ denotes ¹⁴C label position. In vitro reaction phenotyping experiments indicated that CYP3A4, and to a lesser extent CYP2D6, were involved in forming metabolite M2 in liver microsomes [17]. The other minor metabolites were not detected in vitro in liver microsome incubations. Therefore, the specific enzymes involved in their formation have not been determined but likely include CYP3A4 based on the effects of CYP3A4 inhibition on atogepant clinical pharmacokinetics. CYP, cytochrome P450; F, feces; Gluc, glucuronic acid (C₆H₈O₆); M, metabolite; MW, molecular weight; P, plasma; U, urine; UGT, uridine 5′‐diphospho‐glucuronosyltransferase.

FIGURE 5.

Representative radiochromatogram from pooled plasma (0–36 h, A), urine (0–24 h, B), and feces (72–96 h, C) specimens following administration of a single oral dose of 50 mg ¹⁴Catogepant. M, metabolite; R1, region with multiple coeluting metabolites.

Investigations using human plasma samples from a prior study following administration of a single 300 mg oral atogepant dose [15] showed that the reaction of M23 (in representative plasma samples) with hydroxylamine did not result in the formation of a hydroxamic acid derivative chromatographic peak at m/z 665. Therefore, M23 is not an acyl‐glucuronide. Positive and negative control results were as expected.

4. Discussion

Human mass balance and metabolite characterization studies are essential to the drug development process. Improved understanding of an investigational drug's metabolic and elimination pathways and identified metabolites also informs on potential renal and/or hepatic toxicities, as well as DDIs [16]. Mass balance studies with metabolite characterization are of particular importance for gepants, as liver toxicity concerns arose during clinical trials of first‐generation molecules (telcagepant [18], MK‐3207 [19]). Though not fully understood, metabolic oxidation of telcagepant and MK‐3207 is thought to produce potentially hepatotoxic reactive intermediates [20]. This discovery guided the development of second‐generation gepants, including atogepant, that lack hepatotoxic chemical functionalities [20]. In a Phase 3 atogepant clinical trial, the proportion of patients who received atogepant (10, 30, or 60 mg/day) vs. placebo had a lower rate of alanine transaminase (ALT) and aspartate transaminase (AST) elevations (≥ 3× upper limit of normal; 0.9%, 0.9%, 0.4% vs. 1.8%) [3]. A single 50 mg dose of atogepant was well‐tolerated by healthy participants in the current study (N = 6), with no ALT or AST elevations observed. Similar findings were observed at much higher atogepant doses (single 300 mg dose [15], 170 mg/day for 28 days [21]) in healthy participants.

Atogepant is predominantly metabolized via CYP3A4 with minor involvement of CYP2D6, as revealed by an in vitro study using human liver microsomes [17]. An in vitro study identified metabolites M1 (N‐oxide on the azaoxindole moiety) and M2 (5‐hydroxylation on the azaoxindole moiety) following incubation of rat, monkey, and human liver microsomes [17]. Of note, the observed in vitro metabolites were similar between species, and no unique metabolites were observed in humans. Similarly, a comparison of this study to in vivo rat and monkey studies confirmed no disproportionate human metabolites. Therefore, further study to investigate human metabolite safety was not needed per FDA guidance [22].

The current atogepant PK/mass balance study provides important additional details on atogepant disposition. Following a single oral dose of 50 mg ¹⁴C‐atogepant (~200 μCi) in healthy male participants, median T _max for atogepant and total radioactivity was 1 h and 1.5 h after dosing, respectively. Further, the obtained mean T_½ was 18.5 h and 11.6 h for atogepant and total radioactivity, respectively, suggesting that there were no long‐lasting circulating metabolites. The longer T _1/2 of atogepant vs. total radioactivity was likely because atogepant was measurable for a longer period of time following dosing (120 vs. 48 h). Parent atogepant and M23 were the only two radiometric peaks detected in plasma.

Metabolite M23 is proposed to be a glucuronide conjugate (dioxygenated methylated glucuronide of atogepant) based on mass spectral data (Figure 5) and, therefore, is unlikely to be pharmacologically active [23]. Of note, M23 is not an acyl‐glucuronide [17], which carries particular toxicological concern [24]. Because of its probable inactivity [23] and low circulating levels in plasma, the likelihood of M23‐related toxicities and/or DDIs is low [23]. Further, relative exposures for M23 in human plasma versus those in preclinical simian toxicology studies indicate that simian exposures were more than sufficient to assess safety. Simian exposure was 3‐fold higher following a 10 mg/kg dose [17], and an estimated 64‐fold higher at the no‐observed‐adverse‐effect level (300 mg/kg) in chronic toxicology studies (internal data). Mean atogepant/total radioactivity ratio for systemic exposure (AUC_0–∞) was ~0.75, indicating that the parent drug was the main circulating species. In the current study, M23 represented ~15% of radioactivity exposure in plasma and was not long‐lasting.

Fecal excretion was the main route of atogepant elimination. Approximately 89% of total radioactivity was recovered, with ~81% recovered in the feces over 336 h (14 days) and ~8% recovered in the urine. Though total recovery was just below the 90% threshold recommended by US regulatory guidance [13], mean radioactivity recovery was 89% in 171 industry‐conducted mass balance studies [25]. Proposed reasons for low recovery included prolonged fecal recovery [25], which could have contributed here. In feces, 52% of recovered radioactivity (42% of total dose) was as the parent drug due to unabsorbed drug, biliary excretion, and/or intestinal secretion. At least 11 metabolites of atogepant were detected in feces, with each representing < 10% of the total radioactive dose. In urine, the only peak that accounted for > 1% of total radioactive dose was that of atogepant (5% of administered dose). Urine was a minor route of elimination with a CL_R of ~1.2 L/h, accounting for only ~5% of radioactive recovery. This was consistent with a prior study showing a CL_R of ~0.9–1.5 L/h following administration of a single 10–60 mg dose of atogepant in healthy Japanese and White adults [26].

This study had limitations. First, the single 50 mg atogepant dose, although within the clinically relevant dose range for migraine prevention (10–60 mg/day) [3], does not reflect the intended once daily use. However, atogepant does not accumulate with daily dosing [21, 27], and this study followed standard mass balance practices of testing single doses in humans [13, 16]. Second, though atogepant is commercially available as a tablet, ¹⁴C‐atogepant was administered as a liquid solution (in PEG400) to accommodate radioactive compound dosing. PEG is a P‐gp efflux inhibitor [28, 29], which could increase atogepant exposure. However, atogepant bioavailability is higher with tablet vs. PEG solution dosing [17] and a clinical DDI study with quinidine, a strong P‐gp inhibitor, showed no clinically relevant increase in atogepant exposure [12]. Therefore, any potential marginal increase in atogepant exposure due to P‐gp inhibition by PEG400 is unlikely of significance. In support, PK parameters obtained here (liquid formulation) and in late phase clinical trials (tablet formulation) [1] were similar, indicating similar atogepant absorption/disposition. Third, M23 could not be fully characterized so was not synthesized for direct determination of half‐life, bioactivity, and DDI potential. However, based on mass spectrometry data, M23 was proposed to be a glucuronide conjugate, known to be rapidly eliminated and have generally very low pharmacological activity [23].

In conclusion, a single 50 mg dose of ¹⁴C‐atogepant was well tolerated in this small study of healthy males (no TEAEs). ¹⁴C‐atogepant radioactivity recovery was high, with total recovery at ~89% over 14 days. Median T _max was 1 and 1.5 h post‐dose, and mean T _1/2 was 18.5 and 11.6 h for atogepant and total radioactivity, respectively. Atogepant was mainly excreted via the feces (81% total radioactive dose) as unchanged parent drug, with some urinary excretion (8% total radioactive dose). Based on plasma measurements, the parent compound accounted for ~75% of total radioactive exposure, indicating that atogepant was the main circulating species. The only atogepant metabolite detected in plasma was M23 (dioxygenated methylated glucuronide of atogepant), which was short‐lived. Being a glucuronide conjugate, M23 is unlikely to be pharmacologically active. Several other metabolites were identified in feces, but each accounted for < 10% of the total radioactive dose administered. Importantly, these results guided the conduct of an atogepant hepatic impairment study and led to the conclusion that a dedicated renal impairment PK study was not required.

Author Contributions

R.R.B., P.C., and J.M.R. wrote the manuscript; R.R.B. and J.M.R. designed the research; P.C. and J.M.R. performed the research; R.R.B., P.C., and J.M.R. analyzed the data.

Conflicts of Interest

All authors are employees of AbbVie Inc. and may hold stock or options in the company.

Supporting information

Appendix S1: cts70382‐sup‐0001‐AppendixS1.docx.

Boinpally R. R., Chandrasekar P., and Rowe J. M., “Mass Balance and Metabolism of ¹⁴C‐Atogepant in Healthy Male Participants: Findings of a Phase 1 Clinical Trial,” Clinical and Translational Science 18, no. 11 (2025): e70382, 10.1111/cts.70382.

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