The AAPS Journal (2023) 25:67
`https://doi.org/10.1208/s12248-023-00826-1
`
`RESEARCH ARTICLE
`
`Integration of Biorelevant Pediatric Dissolution Methodology
`into PBPK Modeling to Predict In Vivo Performance and Bioequivalence
`of Generic Drugs in Pediatric Populations: a Carbamazepine
`Case Study
`
`Gopal Pawar1 · Fang Wu2 · Liang Zhao2 · Lanyan Fang2 · Gilbert J. Burckart3 · Kairui Feng2 · Youssef M. Mousa2 ·
`Abdullah Al Shoyaib2 · Marie-Christine Jones1 · Hannah K. Batchelor4
`
`Received: 5 January 2023 / Accepted: 25 May 2023 / Published online: 29 June 2023
`© The Author(s) 2023
`
`Abstract
`This study investigated the impact of gastro-intestinal fluid volume and bile salt (BS) concentration on the dissolution of car-
`bamazepine (CBZ) immediate release (IR) 100 mg tablets and to integrate these in vitro biorelevant dissolution profiles into
`physiologically based pharmacokinetic modelling (PBPK) in pediatric and adult populations to determine the biopredictive
`dissolution profile. Dissolution profiles of CBZ IR tablets (100 mg) were generated in 50–900 mL biorelevant adult fasted
`state simulated gastric and intestinal fluid (Ad-FaSSGF and Ad-FaSSIF), also in three alternative compositions of biorelevant
`pediatric FaSSGF and FaSSIF medias at 200 mL. This study found that CBZ dissolution was poorly sensitive to changes in the
`composition of the biorelevant media, where dissimilar dissolution (F2 = 46.2) was only observed when the BS concentration
`was changed from 3000 to 89 μM (Ad-FaSSIF vs Ped-FaSSIF 50% 14 BS). PBPK modeling demonstrated the most predictive
`dissolution volume and media composition to forecast the PK was 500 mL of Ad-FaSSGF/Ad-FaSSIF media for adults and
`200 mL Ped-FaSSGF/FaSSIF media for pediatrics. A virtual bioequivalence simulation was conducted by using Ad-FaSSGF
`and/or Ad-FaSSIF 500 mL or Ped-FaSSGF and/or Ped-FaSSIF 200 mL dissolution data for CBZ 100 mg (reference and generic
`test) IR product. The CBZ PBPK models showed bioequivalence of the product. This study demonstrates that the integration
`of biorelevant dissolution data can predict the PK profile of a poorly soluble drug in both populations. Further work using
`more pediatric drug products is needed to verify biorelevant dissolution data to predict the in vivo performance in pediatrics.
`
`Keywords carbamazepine tablets · in vitro dissolution · PBPK · pediatric biorelevant dissolution media ·
`virtual bioequivalence
`
`Introduction
`
`Dissolution is commonly used as an in vitro testing
`method for orally administered drug products, such as
`tablets. Typically, biopredictive dissolution methods can
`
`use biorelevant media that is designed to simulate the
`composition of gastrointestinal (GI) fluids, such as fasted
`state simulated gastric fluid (FaSSGF) and fasted state
`simulated intestinal fluid (FaSSIF) which are both derived
`from the  characterization of healthy adults GI media
`
` * Gopal Pawar
`G.Pawar@bham.ac.uk
` * Fang Wu
`Fang.Wu@fda.hhs.gov
` * Hannah K. Batchelor
`Hannah.batchelor@strath.ac.uk
`
`1 School of Pharmacy, Institute of Clinical Sciences,
`University of Birmingham, Edgbaston, Birmingham B15 2TT, UK
`2 Division of Quantitative Methods and Modelling, Office
`of Research and Standard, Office of Generic Drug Products,
`
`Center for Drug Evaluation and Research, United States Food
`and Drug Administration, Silver Spring, Maryland 20993,
`USA
`3 Office of Clinical Pharmacology, Office of Translational
`Science, Center for Drug Evaluation and Research, United
`States Food and Drug Administration, Silver Spring,
`Maryland 20993, USA
`Strathclyde Institute of Pharmacy and Biomedical
`Sciences, University of Strathclyde, 161 Cathedral Street,
`Glasgow G4 0RE, UK
`
`4
`
`CELGENE 2048
`1 3
`Vol.:(0123456789)
`APOTEX v. CELGENE
`IPR2023-00512
`
`

`

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` 67 Page 2 of 18
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`The AAPS Journal (2023) 25:67
`
`composition (1, 2). However, the solubility of drug sub-
`stances (and hence dissolution profiles of oral drug products)
`in pediatric GI fluid may be different from those in adults due
`to inherent differences in GI fluid volume (3) and composi-
`tion, for example in terms of bile salt (BS) concentrations (4).
`Biorelevant pediatric Ped-FaSSGF media has been
`proposed previously, based on literature-reported BS
`concentration (5, 6), where values were lower in neonates
`(20  μM) and infants (60  μM) in comparison to adults
`(80 μM). Moreover, due to the lack of accurate data on
`pediatric BS concentrations, simulated Ped-FaSSIF media
`with BS concentrations corresponding to 50 and 150% (1.5
`and 4.5 mM) of adult content (3 mM) were proposed as
`an alternative media for exploratory studies (5). Recent
`studies that characterized pediatric gastric and intestinal
`fluids measured a lower BS concentration in pediatric
`fasted state intestinal fluids compared to adults (0.18 mM
`vs 3 mM) (4, 7–9) providing new data on which pediatric
`simulated media can be developed.
`The volume of dissolution media is a critical factor in
`designing the in vitro dissolution method for poorly solu-
`ble drug substances. Generally, the recommended dissolu-
`tion volume in USP I/II apparatus is 500 mL, which more
`closely resembles the GI volume in adults (10). The vol-
`ume of fluids in pediatric populations has been reported
`in two recent studies where the fasted gastric volume was
`reported to be a maximum of 8 mL in a population from 0
`to 16 years (n = 32) (3) although an alternative study sug-
`gested a median volume of 5.0 mL in infants and 26.6 mL in
`adolescents (11). The corresponding fasted state intestinal
`fluids were reported to reach a maximum of 51 mL in a
`population from 0 to 16 years (n = 32) (3) or have a median
`volume of 23.9 mL in infants and 62.9 mL in adolescents
`(11). When considering the dissolution volume to use, an
`important consideration is the fluid volume administered
`with a tablet which is typically 240 mL in adults whereas
`previous data on relative bioavailability studies suggest a
`volume of 120 mL in children (12, 13).
`Physiologically based pharmacokinetics (PBPK) mode-
`ling uses mathematical models and simulations to combine
`animal or human physiological data with drug character-
`istics to mechanistically describe the PK behaviors of a
`drug (14). PBPK has been particularly useful in pediatric
`drug development as it can factor ontogeny into relevant
`predictions (15–17), making quantitative predictions in
`pregnancy and in the fetus (18, 19), and in examining
`pediatric predictions in drug absorption in varying age
`groups (20). PBPK models can also incorporate in vitro
`dissolution data to predict the in vivo performance of oral
`formulations and to verify the clinical relevance of in vitro
`dissolution data (21–23). The identification of a biopredic-
`tive dissolution method for pediatric populations and sub-
`sequent integration of the generated dissolution data into
`
`1 3
`
`PBPK modeling would aid in de-risking pediatric clinical
`programs, specifically with reference to relative bioavail-
`ability studies or bioequivalence (BE) studies for generic
`drug products. Furthermore, virtual bioequivalence (VBE)
`trials using PBPK modeling could provide a powerful tool
`to predict and compare the in vivo performance of test
`drug products and reference listed drug (RLD) products
`by integrating the dissolution data generated by using
`the biorelevant medias and the inclusion of inter-subject
`variabilities.
`Carbamazepine (CBZ) is an anti-convulsant drug used
`in adults/pediatric populations to control seizures and as a
`mood-stabilizing drug in patients suffering from bipolar dis-
`order (trigeminal neuralgia), attention-deficit hyperactivity
`disorder (ADHD) and schizophrenia (24, 25). The solubil-
`ity of CBZ in water was measured to range from 0.14 to
`0.27 mg/L at 25°C (26) According to the Biopharmaceutics
`Classification System (BCS), CBZ is a BCS class II (poorly
`soluble; highly permeable) compound where dissolution is
`the rate limiting factor for its absorption (27). Due to its
`high pKa (11.8 or 14), no ionization of CBZ is expected
`within the physiological pH range (28). A previous study (5)
`showed that CBZ solubility in neonatal or infant FaSSGF,
`where the BS concentration was, respectively, 25% (20 μM)
`or 75% (60 μM) of adult values (80 μM), was significantly
`reduced at the lower BS concentrations. However, the impact
`on solubility of CBZ upon varying the BS concentration
`in FaSSIF media from 50 to 150% (1.5–4.5 mM) was not
`significant (29). The impact of gastric fluid volume (135 mL
`in neonates, 120 mL in infants) on the dissolution of the
`reference product, Tegretol© (carbamazepine) 200 mg tab-
`lets, was reported using the USP4 Flow-through-apparatus
`and media volumes selected to match those in neonates
`and infants. In all cases, dissolution was incomplete due to
`volume-limited dissolution (29). No data is available on dis-
`solution of CBZ pediatric suspensions which may be the
`more commonly used dosage form for such young patients.
`To better understand the impact of biorelevant dissolution
`conditions on the release of CBZ from a tablet formulation
`and to integrate these dissolution profiles into PBPK soft-
`ware to evaluate the impact on the predicted PK profile in
`pediatric and adult populations, this work seeks to explore
`the following objectives:
`
`• To compare in vitro dissolution profiles of CBZ IR tab-
`lets using industry recommended USP media as well as
`biorelevant simulated adult and pediatric medias.
`• To explore the impact of dissolution media volume on
`the dissolution profiles of CBZ tablet.
`• To compare the dissolution profiles of the RLD and
`generic CBZ 100 mg IR tablets using USP media, adult
`biorelevant, and our new proposed pediatric biorelevant
`media for PBPK-based virtual BE testing.
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`The AAPS Journal (2023) 25:67
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`Page 3 of 18 67
`
`• PBPK model development and validation for CBZ
`to identify the absorption factors that could result in
`bioinequivalence between pediatric and adult populations
`or between branded or generic CBZ product by integrat-
`ing the biorelevant dissolution data into a CBZ PBPK
`model.
`
`Materials and Methods
`
`Chemicals
`
`Bile Salts Cholic acid (CA); glycocholic acid (GC); glyc-
`ochenodeoxycholic acid (GCDC); glycodeoxycholic acid
`(GDC); glycoursodeoxycholic acid (GUDC); taurocholic
`acid (TC); taurochenodeoxycholic acid (TCDC); taurode-
`oxycholic acid (TDC); tauroursodeoxycholic acid (TUDC);
`taurolithocholic acid (TLC); deoxycholic acid (DC), litho-
`cholic acid (LC), and ursodeoxycholic acid (UDC) were
`purchased from either Sigma Aldrich (Gillingham, UK) or
`Acros Organics (Fisher Scientific, Loughborough, UK).
`
`CBZ powder, sodium-lauryl-sulfate (SLS), lecithin, pep-
`sin, acetonitrile HPLC grade, sodium hydroxide (NaOH),
`and hydrochloric acid (HCL) were purchased from Sigma
`Aldrich (Gillingham, UK). Biorelevant powder (Ad-
`FaSSGF/Ad-FaSSIF) was purchased from Biorelevant
`(https:// biore levant. com/; London, UK).
`Tegretol© 100 mg tablets (Novartis Pharmaceuticals UK
`Limited, Batch No-B00964; Exp-07/2023) were used as
`RLD and Carbamazepine Medreich 100 mg tablets (Medre-
`ich PLC, Feltham, UK) as a generic product. Tegretol©
`was purchased from Queen Elizabeth Hospital Pharmacy
`(Birmingham, UK) and generic product from New Castle
`Healthcare NHS pharmacy.
`
`Preparation of Dissolution Media
`
`USP Media SLS (1% w/v) was added to Milli-Q Type-1
`(18.2 MΩ cm–1) water and stirred at 40°C for 2 h. The pH
`was adjusted to enable comparison with biorelevant dissolu-
`tion media. The pH of the USP media was adjusted (but not
`buffered) to either 1.2 (to mimic gastric pH) by adding 0.1
`N HCl or to 6.5 (to mimic intestinal pH) by adding 0.1 M
`NaOH before use. All pH-adjusted USP dissolution media
`were prepared on the day of the experiment and used within
`4 h of preparation (30, 31).
`
`Adult Biorelevant Media Ad-FaSSGF.V2 (pH 1.2) and Ad-
`FaSSIF.V2 (pH 6.5) (9, 32) were prepared according to the
`protocol provided by the supplier, Biorelevant (London, UK).
`The pH of the media Ad-FaSSGF.V2 was adjusted (but not
`
`buffered) to either 1.2 by adding 0.1 N HCl or to 6.5 by add-
`ing 0.1 M NaOH before use. Media was used within 48 h of
`preparation. FaSSIF V2 media is the most widely used simu-
`lated intestinal fluid despite more recent versions and reviews
`of further characterization of adult intestinal fluids (8, 32, 33).
`
`In the literature, FaSSGF-V2 has a pH value of 1.6, but we
`modified it to pH 1.2 for this study as described above. The
`traditional medium to simulate gastric conditions in the
`fasted state was USP simulated gastric fluid (SGF) which has
`a pH of 1.2. Note that due to the pH independent solubility
`of CBZ, this slight difference in pH would not have a sig-
`nificant impact on this study (34). In addition to this, a range
`of ~ 1 to 3 has been reported for gastric pH in fasted healthy
`humans after they have ingested a glass of water (35).
`In this study, Ad-FaSSGF contained taurocholate to represent
`the bile salt concentration (0.08 mM), phospholipids (0.02 mM),
`sodium (34 mM), and chloride (59 mM). The Ad-FaSSIF media
`contained taurocholate (3 mM), phospholipids (0.75 mM),
`sodium (148 mM), chloride (106 mM), and phosphate (29 mM).
`
`Pediatric Biorelevant Media
`
`Ped-FaSSGF/FaSSIF (14BS) Media Ped-FaSSGF containing
`14 bile salts (14BS) and Ped-FaSSIF (14BS) were prepared
`based on the median concentrations (mM) (0.016 mM for
`Ped-FaSSGF and 0.178 mM for Ped-FaSSIF) of bile acids
`present in the gastric or intestinal fluid samples aspirated
`from children reported previously (4). All the ingredients
`listed in Supplementary Table S1 were added to Milli-Q
`Type-1 (18.2 MΩ cm–1) water and stirred for 2 h until com-
`pletely dissolved; the pH of each media was then adjusted to
`either 1.2 by adding 0.1 N HCL or to 6.5 by adding NaOH
`before use. Ped-FaSSGF and Ped-FaSSIF were stored at 8°C
`and used within 48 h of preparation.
`
`Ped-FaSSGF/FaSSIF (50% 14BS) Media This media was pre-
`pared to contain 50% of the concentration of each BS reported
`previously (4) by diluting the Ped-FaSSGF/FaSSIF media
`containing the 14 BS with Milli-Q Type-1 water. As a result
`of dilution, the concentration of other components such as
`sodium chloride, lecithin, and pepsin was also reduced to 50%
`of the originally reported concentration. Finally, the pH of the
`prepared 50% 14BS media was adjusted to either pH 1.2 or
`6.5 by adding 0.1 N HCL or NaOH, respectively. This 50%
`version was included to provide a “worst-case” scenario as
`there was large variability in the measured bile salt concentra-
`tions from the GI fluids of pediatric participants (4).
`
`Ped-FaSSGF/FaSSIF (Na TCA) Media Ad-FaSSGF and Ad-
`FaSSIF contained only a single bile salt, Na TCA (3 mM
`in Ad-FaSSIF; 0.08  mM in Ad-FaSSGF); therefore,
`
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` 67 Page 4 of 18
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`The AAPS Journal (2023) 25:67
`
`Ped-FaSSGF/FaSSIF media containing only Na TCA were
`also prepared. Na TCA concentrations mimicked the total
`bile acid concentration found in the 14 BS pediatric media.
`
`High Performance Liquid Chromatography (HPLC)
`Method Development
`
`HPLC methodology was based on previous methods (36, 37)
`where the accuracy and specificity were previously reported.
`In brief, HPLC analysis of CBZ was carried out using an
`Agilent (1260 infinity 2) HPLC equipped with binary pump,
`DAD detector UV at 285 nm, and C18 column (Ascentis:
`25 cm, 5 μm, 4.6 mm). The mobile phase was water and
`acetonitrile at a ratio of 6:4 (v/v) which ran in isocratic mode
`at a flow rate of 1.0 mL/min. The injection volume was 5
`μL, and the run time was 10 min (min), followed by 1-min
`post run to avoid any co-elution or carry-over of the analyte.
`
`Calibration Standards CBZ Calibration standards
`(0.002–1 mg/mL) were prepared in all the different adult/
`pediatric FaSSGF/FaSSIF media to exclude any interference
`or matrix effect (CBZ HPLC peak and calibration standards
`provided in Supplementary Figures S1A to S1K).
`
`In Vitro Dissolution Study
`
`Dissolution studies were conducted in accordance with USP
`dissolution (General chapter 711) guidelines (38) using USP
`apparatus 2 (Copley Scientific, Nottingham, UK). The tem-
`perature of dissolution medium in vessels was maintained
`at 37.0°C ± 0.5°C. Six replicates/units were used for each
`experiment. The size of the dissolution vessel was changed
`for smaller volumes (200 mL, 100 mL, and 50 mL) to match
`the hydrodynamics of the standard vessels. To scale down
`the volumes, low-volume conversion kits (Copley, UK)
`comprising vessels of either 100- or 200-mL capacity with
`appropriate mini-paddles, vessel cover, and centering ring
`assemblies were used. One mL of dissolution medium was
`withdrawn without replacement at each pre-specified time
`point (15, 30, 60, 90, 120, and 150 min) and filtered using
`Agilent’s 13-mm syringe filters (polypropylene 0.45 μm pore
`size) before analysis using the validated HPLC method. A
`summary of the dissolution conditions is presented in Sup-
`plementary Table S2. The dissolution studies were all con-
`ducted using a 100 mg CBZ tablet. This tablet strength is not
`available in the USA but is available in the UK (25). This
`strength was selected to use the lowest dose to be applicable
`to pediatric populations and also to minimize the solubil-
`ity effects when evaluating the dissolution conditions (30);
`the lower strength enabled the maximum discrimination
`between the dissolution conditions and also accounts for the
`high permeability of CBZ to best enable integration into the
`PBPK modeling software.
`
`1 3
`
`Dissolution Data Analysis
`
`Similarity between two dissolution profiles was determined
`by the difference factor (F1) and similarity factor (F2) (39)
`calculated as follows:
`∑t=n
`||Rt − Tt
`||
`∑ t = n
`
`F1 =
`
`t=1
`
`× 100
`
`Rt
`
`t = 1
`
`(1)
`
`F2 = 50 × log[
`
`√
`1 +
`
`100
`
`∑ t = n
`
`]
`
`t)2
`
`(R
`t − T
`t = 1
`where Rt = Dissolution (% label claim) for reference (R) for-
`mulation at time t
`
`(2)
`
`Tt
`n
`
` Dissolution (% label claim) for test (T) formulation at
`time t.
` number of time points
`
`Generally, three or more dissolution sampling timepoints
`with one measurement after 85% dissolution are required to
`calculate F1 and F2. The compared dissolution profiles are
`considered similar when F1 ≤ 15 and F2 ≥ than 50.
`
`CBZ PBPK Model
`
`A PBPK model for CBZ was developed using SimCyp®
`Simulator (Version 21, Release 1; Certara UK Limited, Shef-
`field, UK) that incorporated drug-dependent and system-
`related input parameters obtained from SimCyp’s internal
`compound library (Supplementary Table S4). The in vitro
`dissolution datasets generated in the “In Vitro Dissolution
`Study” section were incorporated into SimCyp® Simula-
`tor Advanced Dissolution, Absorption and Metabolism
`(ADAM) model; full details of the dissolution data that was
`integrated are provided in Supplementary Table S3.
`The dissolution profiles using various BS concentrations
`and dissolution volumes were integrated into SimCYP
`according to the methods previously described by Guimaraes
`(2002) (40). In brief, a single-stage fasted intestinal profile
`(FaSSIF dissolution conditions as an intestinal profile) was
`integrated as the “intestinal profile.” Using this approach,
`the software considers the same dissolution profile for the
`stomach and intestinal compartments (FaSSIF dissolution
`conditions). A second scenario was also included where
`both the single-stage fasted gastric and intestinal profiles
`(input of the CBZ single-stage dissolution profile in fasted
`state simulated gastric fluids (FaSSGF) entered as “stomach
`profile” and single-stage dissolution profile in fasted state
`simulated intestinal fluids entered as “intestinal profile” thus
`incorporating both FaSSGF + FaSSIF dissolution conditions.
`
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`

`The AAPS Journal (2023) 25:67
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`Page 5 of 18 67
`
`Model prediction was validated using published clinical
`PK studies with CBZ (Tegretol© IR 100 or 200 mg) con-
`ducted in an adult population (41–44). Note that, here, the
`term validation is used as defined by Kuemmel 2019, “Process
`of determining the degree to which a model or simulation is
`an accurate representation of the real world.” In other terms,
`“validation relates to model (model form, model inputs),
`comparator (test samples, test conditions), and assessment
`(equivalency of input parameters, output comparisons)” (45).
`SimCyp Pediatric Version 21 (Release 1) was used as
`the pediatric PBPK (ped-PBPK) modeling platform. CBZ
`specific properties (including its metabolic clearance)
`used in the adult PBPK model (Supplementary Table S4)
`were transferred into the pediatric model (Supplementary
`Table S5). The metabolic clearance of CBZ is mediated by
`CYP3A4, CYP 3A5, and CYP2C8; hence, the ontogeny of
`CBZ-metabolizing enzymes in pediatric populations was
`expressed according to the following sigmoidal function
`(Eq. 3) which is a default equation used in SimCyp (46, 47).
`
`+ (F
`Fraction of adult = F
`− F
`max
`Birth
`Birth
`× Agen∕(Age50
`n + Agen)
`
`)
`
`(3)
`
`where Fmax is the maximum adult relative expression, Fbirth
`is the relative expression of enzyme at birth, Age50 is the age
`at which the fractional expression (/activity) is in the middle
`of the birth and adult values, and n is the exponent related to
`the sigmoidicity of the developmental curve or analogous to
`the Hill coefficient.
`In order to incorporate the age-dependent expression pat-
`tern of CYP3A4, CYP3A5, and CYP2C8 in the pediatric
`PBPK model, the time to reach 50% maturation (Age50) for
`each enzyme was used. The Age50 of hepatic and intestinal
`CYP3A4 maturation was 0.64 and 2.36 years, respectively
`(47–49). For CYP3A5, since no age-dependent change in
`hepatic expression was noted (50), no ontogeny for hepatic
`CYP3A5 was assumed in this study but for intestinal ontog-
`eny, similar Age50 value (i.e., 2.36) of CYP3A4 was consid-
`ered. For CYP2C8 ontogeny, Age50 of hepatic maturation
`was 0.366 years (51). A zero value was used for intestinal
`CYP2C8 ontogeny due to lack of abundance data (52).
`Generally, young children have lower volume of distri-
`bution (Vss) for lipophilic drugs due to reduced body fat
`compared to adults. Due to lack of a volume of distribution
`(Vd) values in children, a Vd of 0.3 L/kg in young children
`(compared to 0.78–1.9 L/kg in adults) was estimated by our
`visual trial and error simulations in order to fit the pediatric
`PBPK model to the observed data (53). Moreover, the vol-
`ume of fluid intake with CBZ tablet was changed to 120 mL
`for pediatric populations compared to 240 mL for adult pop-
`ulations in line with reported clinical protocols (13).
`The CBZ model was validated in adults by calculating the
`prediction error percentage (PE%) fold error (FE), average
`
`fold error (AFE), and absolute average fold error (AAFE) for
`Cmax and AUC. The adult model was subsequently updated
`and applied to children (6–15 years) and further verified and
`validated using clinical PK data conducted in children on
`20 mg/kg dose (n = 12) by Hartley (1991) (54) and 9.3 mg/
`kg dose (n = 6) by Bano (1986) (55) (Table III).
`This PBPK model was also used to predict the plasma con-
`centration profile of CBZ in pediatric populations younger
`than 6 years to better understand the potential differences
`in CBZ exposure between children younger and older than
`6 years old when CBZ was administered as an IR suspension
`(prepared from crushed Tegretol© tablets) (56). CBZ doses
`of 17 and 19 mg/kg suspension were tested in 20.9 days new-
`born (n = 7; 3.2 kg; 25 mL water) and 5.1-year-old children
`(1.25–8 years; n = 5; 120 mL water), respectively (56). Based
`on the fraction absorbed (66%) from the IR suspension (42),
`the 66% was assumed to be dissolved for an IR suspension to
`run the simulations in younger (< 6 years) pediatric popula-
`tion. This assumption would be a limitation of model simula-
`tions, and any future dissolution work would help to further
`improve the model prediction.
`Due to higher clearance values in younger children (57), a
`clearance value of 3.18 L/h (42) was used to run the model;
`the volume of distribution (Vss) was fitted (by visual obser-
`vation of the superimposed predicted and observed data-
`sets) (2.6 L/kg for 20.9 days new-born and or 2.850 L/kg
`for 5.1 years child) to capture the observed clinical datasets
`published in Rey (1979) study (56).
`
`Virtual Bioequivalence (VBE) Studies
`
`Preliminary exposure simulations were performed using the
`dissolution data generated in Ad-FaSSGF and Ad-FaSSIF
`500 mL media for Tegretol© 100 mg IR tablets and the pre-
`dicted PK profile was validated using literature data (Kohl-
`man study (42); 200 mg IR tablet). Although the dissolution
`data was generated using 100 mg tablet, previous literature
`has reported that the dissolution of these IR tablets is similar
`to the 200 mg product used in the clinical study (54). So, the
`assumption of similar dissolution profiles for both 100 mg
`and 200 mg tablets was used for the simulation purposes.
`VBE simulations were performed in a fully replicated, two
`treatment, multiple trials (N = 10) in a cross-over design with
`a sample size of n = 12,16, 24, 36, and 48 in healthy adults
`(18–45 years) (58). The number trials (N = 10) were fixed as
`per the SimCyp VBE protocol (23, 58). A single dose of CBZ
`IR tablets 100 mg was administered with 240 mL of fluids.
`For VBE simulation, the dissolution profiles for Tegretol©
`100 mg and generic product 100 mg (Medreich) generated
`in Ad-FaSSGF and FaSSIF 500 mL were incorporated into
`the adult PBPK model. The default coefficients of variation
`(%CV) for accounting the inter-subject variability of the
`physiological parameters provided in the SimCyp database
`
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` 67 Page 6 of 18
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`The AAPS Journal (2023) 25:67
`
`for the North European Caucasian healthy adult volunteers
`population were applied for each parameter.
`For pediatrics, VBE simulations were performed with
`a sample size of 12, 16, 24, 36, and 48 healthy subjects
`(6.5–15 years, to match existing published clinical data sets)
`and a single dose of CBZ IR tablets 100 mg administered
`with 120 mL of fluids. Furthermore, the pediatric simulation
`outputs were validated using literature data (Hartley (1991);
`20 mg/kg/day given as 100 or 200 mg tablets twice daily and
`Bano (1986); 9.3 mg/kg per day).
`The input dissolution profile from the full suite of
`in vitro dissolution tests (Ad-FaSSGF and FaSSIF at
`500/200/100/50 mL; Ped-FaSSGF and FaSSIF-14 BS at
`200 mL or Ped-FaSSGF and FaSSIF-50% 14 BS at 200 mL or
`Ped-FaSSGF and FaSSIF-Na TCA at 200 mL) was integrated
`into the PBPK model to identify which profile gave the best
`match to the existing clinical data set from Hartley study (54).
`
`Evaluation of Predictability of the PBPK Model
`
`Results
`
`In Vitro Dissolution Data
`
`The dissolution profiles of CBZ 100 mg tablets obtained using
`the pH-adjusted USP media (30) were compared with those
`obtained using biorelevant adult FaSSGF and FaSSIF media
`with a dissolution volume of 900 mL (Supplementary Figure S2).
`When the dissolution profile from USP dissolution media
`1% SLS (pH 1.2 and 6.5) was compared to Adult FaSSGF/
`FaSSIF profiles, a decrease in % of release for Tegretol©
`100 mg tablets was observed. The major difference in these
`media is the surfactant type and concentration used; a 1%
`SLS media is a high surfactant concentration and perhaps it
`is unsurprising that such raid dissolution is observed com-
`pared to the biorelevant media. The results indicated that the
`pH of the media did not affect the overall dissolution.
`The impact of media volume on dissolution was evalu-
`ated using both Ad-FaSSGF and Ad- FaSSIF media and the
`results are shown in Fig. 1a and b, respectively.
`As expected for a poorly soluble drug like CBZ, reduc-
`ing the volume of dissolution media resulted in decreased
`% drug release from the tablet formulation. However, the
`decrease in % release was not significant according to the
`F1/F2 statistic when the volume was reduced from 900 to
`500 mL and from 100 to 50 mL. For both Ad-FaSSGF and
`Ad-FaSSIF media, the F1/F2 values are shown in Table I.
`The impact of media composition on the dissolution of
` Tegretol© 100 mg tablets was further assessed in a 200 mL
`volume of both simulated pediatric gastric and intestinal
`media to mimic the pediatric GI fluid volume (Fig. 2a and b).
`When 200 mL of media was used, based on F1 and F2
`comparison, dissimilarity was observed between Ad-FaS-
`SIF and Ped-FaSSIF 50% 14BS (Table II). This is likely
`resulting from the large differences in the BS concentra-
`tion (3000 vs 89 μM).
`
`In Vitro Dissolution Comparison Between Generic
`and RLD Product
`
`Adult FaSSGF/FaSSIF (500 mL)
`
`Dissolution of the RLD and generic product was compared
`in simulated adult GI media using a volume of 500 mL. The
`volume of 500 mL is consistent with in vivo studies where it
`is known that by ingesting 250 mL of water with the dosage
`form, plus the residual water in the adult GI tract that a maxi-
`mum total volume of about 300–500 mL will be available in the
`proximal small intestine (1, 63, 64). Based on the F1/F2 criteria,
`the dissolution profiles of the two products were similar in Ad-
`FaSSIF (F1 = 6.5, F2 = 64.2), but not in Ad-FaSSGF (F1 = 21.7,
`F2 = 43.1) (Fig. 3). A similar conclusion was reached when the
`test was repeated in 200 mL simulated pediatric media (Fig. 4).
`
`Prediction error for AUC and Cmax were evaluated by the
`following formula (14, 59, 60):
`|Predicted parameter − Observed parameter|
`
`% Prediction Error =
`
`× 100
`
`Observed parameter
`
`(4)
`In this study, we considered a PE value of < 20% confirm-
`ing the good predictability of the model. In addition, Fold
`error (FE) for AUC and Cmax were evaluated by the follow-
`ing equation (40, 59–61):
`
`Fold Error =
`
`Predicted
`Observed
`
`(5)
`
`Furthermore, the mean predicted plasma concentra-
`tion–time profiles were also assessed by the average fold
`error (AFE) and validated by the absolute average fold error
`(AAFE) (Eqs. 6 and 7 respectively) (40, 61).
`∑
`,log (Predicted)i
`(Observed)i
`
`1 n
`
`AFE = 10
`
`(6)
`
`(Observed)i
`
`∑|||,log (Predicted)i
`|||
`(7)
`AAFE = 10
`where n denotes the number of observed PK sampling
`points, Predictedi is the predicted concentration at time point
`i, Observedi is the observed concentration at time point i.
`The FE indicates the predictive accuracy of each data point
`as shown in equation (62). The AFE indicates whether the
`predicted profile underestimates (AFE < 1) or overestimates
`(AFE > 1) the observed values, as shown in Eq. 6, whereas the
`AAFE quantifies the absolute error from the observed values,
`as shown in Eq. 7. An AAFE value close to unity represents
`the precision of the simulations (62). When comparing the
`simulated data to clinical data, the populations were matched
`in terms of age range and sample size for each study.
`
`1 n
`
`1 3
`
`

`

`The AAPS Journal (2023) 25:67
`
`Fig. 1 Dissolution profiles of
`100 mg CBZ tablets (Tegretol©)
`in Ad-FaSSGF a and Ad-
`FaSSIF b media as a function of
`dissolution media volume. The
`data points show the mean of 6
`values and the error bars show
`the % CV
`
`Page 7 of 18 67
`
`Pediatric FaSSGF/FaSSIF (200 mL)
`
`Comparison of % of release between reference CBZ
` (Tegretol©) and generic (Medreich) 100 mg tablets in Ped-
`FaSSIF media containing 14 BS showed dissolution simi-
`larity between the two products (Fig. 4). At pH 1.2, com-
`parison of both the reference and generic profiles showed
`differences (F1 = 34.8 and F2 = 43.0). It appears that CBZ
`
`generic has higher % release at gastric pH as compared to
`the reference drug.
`
`Validation of Adult and Pediatric PBPK Model
`
`Simulations conducted using the adult PBPK model cap-
`tured the observed plasma time-concentration profiles for
`CBZ given as a single oral dose of 100 mg (41), 200 mg (41,
`
`Table I The F1/F2 Values for the Dissolution Profiles that Use an Alternative Volume of Media. Note that F1 Values ≤ 15 and F2 Values ≥ 50
`Indicate Similarity Between Profiles, Where Similarity Is Shown the Numbers Are Highlighted in Green
`Ad-FaSSGF
`Ad-FaSSIF
`500 mL
`500 mL
`9.1
`12.5
`58.8
`52.9
`
`900 mL
`
`500 mL
`
`200 mL
`
`100 mL
`
`200 mL
`42.4
`25
`36.6
`29.8
`
`100 mL
`79.7
`12.6
`77.7
`15.3
`64.8
`29.6
`
`50 mL
`87.1
`F1
`10.4
`F2
`85.8
`F1
`12.7
`F2
`77.6
`F1
`25.4
`F2
`45.1
`F1
`57.6
`F2
`Ad-FaSSGF adult-fasted state simulated gastric fluid, Ad-FaSSIF adult-fasted state simulated intestinal fluid
`
`200 mL
`45.2
`24.0
`37.3
`30.5
`
`100 mL
`78.6
`12.9
`75.5
`16.7
`61
`32.3
`
`50 mL
`87.7
`10.7
`85.9
`14.0
`77.5
`27.0
`42.4
`59.4
`
`1 3
`
`

`

`
`
` 67 Page 8 of 18
`
`Fig. 2 Comparison of the
`dissolution profiles of 100 mg
`CBZ tablets (Tegretol©) in
`200 mL simulated gastric fluid
`(a) and intestinal fluids (b). The
`data points show the mean of 6
`values and the error bars show
`the % CV
`
`The AAPS Journal (2023) 25:67
`
`42), and 400 mg (as 2 × 200 mg)