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Abstract
The objective of this study was to develop a population pharmacokinetic-pharmacodynamic model of subcutaneously administered bupivacaine in a novel extended release microparticle formulation for postoperative pain management. Bupivacaine was administered subcutaneously in the lower leg to 28 healthy male subjects in doses from 150 to 600 mg in a phase 1 randomized, placebo-controlled, double-blind, dose-ascending study with two different compositions of microparticle formulations called LIQ865. Population pharmacokinetic-pharmacodynamic models were fitted to plasma concentration-effect-time data using non-linear mixed-effects modeling. The pharmacokinetics were best described by a two-compartment model with biphasic absorption as two parallel absorption processes: a fast, zero-order process and a slower, first-order process with two transit compartments. The slow absorption process was found to be dose-dependent and rate-limiting for bupivacaine clearance at higher doses. Bupivacaine clearance and the transit rate constant describing the slow absorption process both decreased with increasing doses following a power function with a shared covariate effect of dose on the two parameters. The pharmacokinetic-pharmacodynamic relationship between plasma concentrations and effect was best described by a linear function. This model gives new insight into the pharmacokinetics and pharmacodynamics of microparticle formulations of bupivacaine, and the biphasic absorption seen for several local anesthetics.
Glossary of Terms
PK pharmacokinetic
popPK population pharmacokinetic
PD pharmacodynamic
PLGA poly-lactic-co-glycolic acid
WDT warmth detection threshold
k tr transit rate constant
CL/F apparent clearance
OFV objective function value
GOF goodness-of-fit
C max maximum plasma concentration
IIV inter-individual variability
CV coefficient of variance
Q/F apparent inter-compartmental clearance
V/F apparent distribution volume of the central compartment
V2/F apparent distribution volume of the peripheral compartment
Introduction and Background
Persistent postoperative pain following surgical procedures is still common and leads to poor outcomes and prolonged hospitalization for patients [1]. As the number of surgical procedures is increasing globally, there is a growing need to improve postoperative pain management [1]. The many well-known drawbacks of using opioids for postoperative pain treatment make opioid-sparing approaches preferable. The vast majority of local anesthetics for postoperative interventional pain management have an efficacy of less than 24 hours [2,3]. In recent years, several new prolonged-release formulations of the local anesthetic bupivacaine have been developed or are in active development, and two have been approved by the FDA and/or by EMA [3]. These formulations have a reported duration of pain relief between 48 and 72 hours [3], but their impact on clinical outcomes after surgery appears to be limited [4,5]. This study uses data from a phase 1 randomized, placebo-controlled, double-blind, dose-ascending study, which explores two different compositions of a novel microparticle formulation LIQ865 containing bupivacaine for extended release using particle replication in non-wetting templates (PRINT®) technology [6].
Previous studies have found biphasic absorption profiles and effect trajectories of extended duration anesthetics such as bupivacaine and lidocaine in humans and dogs [7-12]. A commonality of this biphasic absorption of bupivacaine is a fast initial absorption process followed by (or parallel to) a slower absorption of the remaining drug [2,7-12]. There is large variability between individuals in the dose fractions absorbed by the fast and the slow absorption processes, respectively [8,9], which a simple average would tend to hide. Population pharmacokinetic (popPK) modeling is, therefore, an ideal tool to describe the PK of locally administered bupivacaine, as the inter-individual variability can be accounted for and quantified. Previously developed popPK and/or pharmacodynamic (PD) models of bupivacaine were based on central neuraxial blocks [7,10-12]. The models developed by Doherty, Simon, et al. (2004)[10] and Olofsen, et al. (2008)[11] all described biphasic absorption by two parallel processes but implemented in different manners to various degrees of success in producing the biphasic concentration-time profile. Therefore, this study seeks to develop a popPK/PD model of bupivacaine after subcutaneous administration based on data from 28 volunteers in a first-in-human study of the novel microparticle bupivacaine-containing formulation LIQ865. The purpose of the model is to describe the biphasic absorption and effect profiles and to investigate the influence of increasing doses on the pharmacokinetic parameters.
Materials and Methods
Pharmacokinetic and Pharmacodynamic Data
The data was collected in a phase 1 randomized, placebo-controlled, double-blind, dose-ascending clinical study (dose safety and pharmacodynamics is described in a separate paper submitted to Basic & Clinical Pharmacology & Toxicology: ‘Bupivacaine in a novel extended-release micro-particle formulation (LIQ865). A phase 1, randomized, double-blind, placebo-controlled, dose-ascending study in male volunteers: Dose safety and pharmaco-dynamics’ by Jensen EK, Bøgevig S, Balchen T, Springborg AH, Royal MA, Storgaard IK, Lund TM, Møller K, Werner MU). The study protocol with updated amendments was approved by the Research Ethical Committee for the Capital Region of Denmark (H-16040444) and the Danish Medicines Agency (2016092814) and was registered in EudraCT (2016-002420-88) and ClinicalTrials.gov (NCT02982889, principal investigator: Mads U. Werner, date of registration: 06-DEC-2016) prior to study start. The study was conducted in accordance with the Basic & Clinical Pharmacology & Toxicology policy for experimental and clinical studies [13]. Written informed consent was obtained from all subjects included in the study.
Twenty-eight healthy, non-smoking male subjects (required age 18-45 years; BMI 18.5-30.0 kg/m2; weight> 60 kg) were divided into five cohorts receiving subcutaneous injections of bupivacaine (doses 150, 225, 300, 450, or 600 mg) as either formulation LIQ865A or LIQ865B. The extended-release study drugs were made using the PRINT-technology, producing hexagonal microparticles[6]. LIQ865A (formulation A) consisted of a mix of the PRINT bupivacaine base and poly-lactic-co-glycolic acid (PLGA) in a ratio of 55%/45%, while LIQ865B (formulation B) consisted of 100% PRINT bupivacaine base. Subcutaneous injections were performed from two injection points with fan-like needle trajectories in a testing area (2.5 x 5.0 cm2) on the lower legs, with bupivacaine formulation on one side (active drug) and a corresponding volume of diluent on the other (control). Diluent control was used as the baseline for PD. Aqueous 0.5% bupivacaine was scheduled for use as an active control in cohort 5. However, as this cohort was discontinued due to the first patient reporting symptoms of possible systemic side effects, a historical control utilizing data from a similarly designed study was used instead [14]. Due to the study being first-in-man for this formulation, group sizes were kept small.
Blood samples (129 mL in total) were taken during Day 0 at times 0, 0.5, 1.0, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 8, 12, and 24 hours, and once daily on Day 2-5 post-injection. Locally, blood samples were centrifuged, separated, frozen, and stored at -80°C. The frozen samples were then shipped on dry ice to the analysis facility (AIT Bioscience, Indianapolis, IN), where they were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay with a lower limit of detection of 2.00 µg/L.
Warmth detection threshold (WDT) measurement was used as a surrogate PD endpoint in this study, and the PD assessments were made at baseline and 1, 2, 4, 6, 8, 12, 24 hours, and at Days 2-5 post-injection. The assessments were made in triplicate at each time point on both legs. The temperature was increased from a baseline of 32°C using a ramp rate of 1°C/s until a sensation of warmth was perceived. The stimulus was then terminated by the subject using a handheld stop button device. The cut-off limit was 50°C, set to avoid skin damage. Assessment values exceeding 50°C were assigned the value of 51°C. A higher WDT thus indicated lower sensibility and, therefore, a higher effect.
Population Model Development, Software, and Statistics
Data management was performed in Phoenix NLME version 8.3.295 (Certara, USA). Models were fitted to the concentration-time data using non-linear mixed effects modeling[15-18] in Phoenix. The first-order conditional estimation-extended least squares (FOCE-ELS) algorithm was used during the development of the structural and stochastic model, and the quasi-random parametric expectation maximization (QRPEM) algorithm was used for further modeling of covariates and PK/PD model as the models became more complex. Models were parameterized in terms of clearances and volumes.
One-, two-, and three-compartment PK models were fitted to the concentration-time data. Different absorption models were applied; first order absorption, zero-order absorption, absorption with lag time, parallel absorption from the same or two different dose points, transit compartment models with and without an absorption compartment, and combinations of these. Additive, proportional, and combined additive and proportional error models were fitted, and the best was chosen. Different combinations of random effects (inter-individual variability) applied on fixed effects were investigated. The influences of formulation (A or B) and dose described either as cohort (1-5), high or low dose group, or dose as a continuous covariate (relative to the lowest dose) were tested as covariates on relevant population parameters. A shared covariate effect of dose on the transit rate constant (k tr) and clearance (CL) was also tested and compared to a model with separate covariate effects of dose onk tr and CL.
After the PK model was developed, PD data was linked to plasma concentrations through a number of different PK/PD models without freezing PK parameters (open model) [19]. Due to the upper value of 51°C for WDT, it was explored whether Beal’s M3 or M4 methods [20] could be used to account for the censored data above the threshold.
Criteria considered for model evaluation were objective function value (OFV) (p < 0.05 for the structural model and p< 0.001 for covariate inclusion), relative standard errors for fixed effects (preferably < 30%), and goodness-of-fit (GOF) plots generated in Phoenix. Shrinkage values for random effects reflect the reliability of individual (post hoc) parameter estimates (with lower values indicating better reliability) and should ideally be below 25%.
Results
Pharmacokinetic and Pharmacodynamic Data
The final dataset for PK/PD modeling contained a total of 453 data points for PK and 335 data points for PD from 28 healthy adult male subjects. Data points covered measurement times from 0 (pre-dose) to 120 hours after dosing. Cohorts 1 (150 mg), 2 (225 mg), and 3 (300 mg) each had 6 subjects, 3 given each formulation A and B. Cohort 4 (450 mg) had 9 subjects, 6 given formulation A and 3 given formulation B. Cohort 5 (600 mg) had a single subject given formulation A. Cohort 5 was discontinued after subject #504 reported symptoms indicating local anesthetic systemic toxicity after injection of 600 mg LIQ865A. Three other subjects (#501, #502, and #503) already included in cohort 5 were therefore reassigned to cohort 4, expanding this cohort. Other adverse effects were minor and local to the injection sites. These included reversible indurations lasting 4-13 weeks experienced by 5 subjects receiving LIQ865A. The mean of the control WDT measurements in the leg injected with diluent was 36.5°C (CV = 4.5%).
Initial evaluations of plots of plasma concentration vs. time for individual subjects indicated the presence of a bi-phasic systemic absorption with two peaks apparent in the majority of plasma concentration-time profiles. Supplementary Digital Content S1 shows plasma concentration-time profiles and effect-time (with WDT as effect measure) profiles for cohort 4 as an example.
The first peak occurred within the first hours after administration, while the second peak was broader and peaked around 25-50 hours after administration. There was high variability between subjects regarding the presence, magnitude, and dominance of each absorption process. Clearance appeared to slow down with increasing doses, indicating either saturation of the elimination processes or dose-dependent absorption rates resulting in flip-flop kinetics at higher doses. The latter option is supported by the fact that there are no previous reports of saturation of clearance in clinically used doses of bupivacaine. Large inter-individual variability in bupivacaine plasma concentration profiles was observed, as subjects given the same dose had considerably varying magnitudes of maximum plasma concentrations (C max). An example of this is shown in the individual profiles for subjects #109 and #112 in Figure 1, whereC max of subject #112 is approximately 2.5 times that of subject #109, though both were given the lowest dose of 150 mg. In addition, subject #202 did not reach the sameC max as subject #112 despite having received the higher dose of 225 mg.
Pharmacokinetic Model with Biphasic Absorption
For the structural model, the PK of bupivacaine was best described using a 2-compartment model compared to a 1-compartment model (p< 0.001). Absorption models with lag time, fixed numbers of transit compartments, variable numbers of transit compartments (Stirling approximation), zero- and first-order absorption models, and variations and combinations thereof were evaluated but were not able to describe the bi-phasic absorption profile well. Finally, the following two parallel absorption phases were implemented with separate dosing points: the fast absorption process described by zero-order kinetics with the duration τ0, and the slow absorption process described by first-order kinetics with two transit compartments and Erlang type absorption (where the transit rate constant k trequals the absorption rate constant k a). The fraction of the dose absorbed through the fast process was defined as the parameter Fr, while the fraction of the dose absorbed through the slow process was defined as (1-Fr). Fr was then limited to a value between 0 and 1. This model was able to describe the bi-phasic absorption well and was the overall best fit as evaluated on goodness-of-fit plots and OFV. A schematic representation of the population PK model is shown in Figure 2.
A proportional error model was the best fit for the data in terms of GOF plots and standard deviations. Regarding individual parameter estimates, random effects (inter-individual variability, IIV) were applied on the parameters CL, τ0, Fr, and k tr, with shrinkages below 10%, indicating good reliability on individual estimates for these parameters.
Significant covariate relationships included dose (scaled to the lowest dose of 150 mg, since this cohort appeared to have the apparent clearance closest to clearance after an i.v. dose based on literature [2] and initial evaluations of data) as a covariate on CL andk tr. Formulation (A or B) did not show any significant covariate effect on any parameters. CL only appeared to slow down once k tr decreased to a value below the elimination rate constant, and both parameters followed approximately the same rate of decline. This pointed to flip-flop kinetics as the cause of the decrease in CL rather than a saturation of elimination processes. As the observed decrease in CL followed the dose-dependent decrease in k tr, a model with shared covariate effect for dose on CL and k tr was applied. The implementation of this covariate removed the trends in the eta vs. covariate plots as seen for CL and k tr (Figure 3). This model was chosen as the final PK model based on GOF plots and OFV. With this model, it is possible to estimate population values of CL and k tr for any chosen dose of bupivacaine (see Table 1 and Figure 4).
Population Pharmacokinetic-pharmacodynamic Model
The following relations between plasma concentration and effect were tested: linear, log-linear, Emax model, sigmoidal Emax model, the addition of an effect compartment (indirect models), and effect relating to the plasma concentration of the peripheral compartment. The linear correlation between plasma concentration and effect was the best fit for the PK/PD model. Although methods of letting the model estimate effects at values above the ceiling of 51°C were explored, these did not improve the model overall, and the linear relation parameters between plasma concentration and effect were the same. A random effect parameter (IIV) was included on the slope of the linear relation. The inclusion of a random effect parameter on the baseline of the relation was tested. However, this resulted in a high shrinkage of 49% and a small estimated value (CV% < 2%) and, therefore, this parameter was not included. Parameter values, relative standard errors, IIV, and shrinkage for the final PK/PD model are presented in Table 1. The population-predicted profiles of plasma concentration and effect (WDT) over time are presented in Figure 5. For the full model code of the final popPK/PD model, see Supplemental Digital Content S2. For selected GOF plots of the final popPK/PD model, see Supplemental Digital Content S3.
Discussion
The PK profiles of individual subjects showed large variability in maximum plasma concentrations and relative amounts absorbed through the fast and slower absorption routes, even within the same dosing cohorts (see Figure 1). This is reflected in the IIV on the parameters τ0 (51.9%) and Fr (57.4%), describing the duration of and the fraction absorbed through 0-order absorption, respectively. The model is able to capture this variability relatively well in a descriptive manner, but it complicates the prediction of the magnitude of plasma concentrations and effects (and, therefore, side effects). This issue could be illuminated further by a study with larger cohort sizes than what was ethically reasonable in this first-in-human trial. Reasons for the large variability in C max could be the varying degree of inflammation observed in the subjects, as inflammation increases the blood flow in the injection area. This, in turn, could increase both Fr and the transit rate constantk tr. Another reason could be the method of preparation of the drug suspension for injection, as the homogeneity and concentration of drug throughout the suspension could vary.
From the observed PK profiles, CL appeared to be dose-dependent with possible flip-flop kinetics. Covariate box plots of eta onk tr and CL faceted on cohorts showed a trend of decreased k tr and CL with higher doses (Figure 3). Therefore, it appeared more likely that k trwas the dose-dependent parameter while CL was dependent onk tr at higher doses. Ask tr decreases and approaches the elimination rate constant k e, flip-flop kinetics occur, where the rate of elimination is limited by the rate of absorption. Covariate models with different covariate effects for dose onk tr and CL were investigated, along with models where a shared covariate effect was applied, as chosen for the final PK model. The dose-dependence of the absorption rate could be due to saturable absorption processes or properties of the formulation, e.g., longer diffusion distances through the formulation before absorption is possible. With this covariate model, it is possible to estimate CL andk tr for a given dose using the values in Table 1. However, as the present study was first-in-human, the subjects were all male and of a limited weight range, and extrapolating the model to real-world patients should be done with caution.
The absorption of bupivacaine was best described by two simultaneous absorption processes: a 0-order absorption process, functionally similar to an infusion (the fast phase), and a first-order absorption with two transit compartments with Erlang type absorption (the slower phase). This combined absorption model was able to describe the individual PK profiles well. A possible explanation for the simultaneous phases may be found in the microparticle suspension formulation. A fraction of drug molecules may be readily available for systemic absorption upon administration, while the remainder must diffuse through the formulation and diluent first. The type of formulation, A or B, did not show a significant covariate effect on any parameters, although increased signs of local inflammation were observed in subjects given the formulation with PLGA.
Subject #504 was the only subject given a dose of 600 mg, as the dosing group was discontinued after this subject experienced suspected systemic side effects including dizziness, light-headedness, pricking paresthesia, and numbness of the tongue. These were treated as suspected unexpected serious adverse reactions suggestive of local anesthetic systemic toxicity. These side effects followed a biphasic trajectory, as was seen for the PK profiles and effects in general. Several subjects experienced a period with some recovery of sensory function between two periods of complete local anesthesia. This breakthrough sensitivity should be considered when using extended-release local formulations of local anesthetics like the ones in this study.
Intravenous (i.v.) PK data were not obtained in this study, but a review by Heppolette, et al. (2020)[2] found bupivacaine clearance after i.v. dosing to be between 27.9 and 39 L/h. The apparent clearance found in this study of 35.4 L/h (CV = 6.3%, IIV = 24.5%) for the low dose group is in the upper end of this interval, indicating high bioavailability. Therefore, it seems likely that the CL found for the low dose group is either not affected or only affected to a negligible degree by flip-flop kinetics. The apparent volume of distribution of the central compartment (V/F) of 347 L found in this study is relatively high, as literature values for distribution volumes are generally below 200 L [2]. Distribution volumes in literature are found following i.v., epidural, peripheral nerve block (femoral or sciatic) administration, and not subcutaneous administration as in this study [2]. This study found a 2-compartment model to be the best fit for the data; Olofsen, et al. (2008)[11] likewise describe a model with distribution to peripheral compartments. Overall, the model is structurally similar to previously developed models of epidurally administered bupivacaine, with some exceptions, particularly the structure of the absorption model, which is due to the different administration routes and formulations of bupivacaine.
In the PK/PD model, a linear relationship between plasma concentration and effect (measured as WDT) best described the data. There was a ceiling effect for the PD data, as applying higher temperatures than the maximum of 50°C would have carried the risk of skin damage. The ceiling effect influences the quality of effect predictions using the linear relation, and the variability of the linear relationship is also reflected in the relatively high IIV on the slope (56.7%). The intercept (baseline) corresponded to the mean of the PD control measurements. Schnider, et al. (1996)[12] found an Emax type model to be the best fit for PD data, but this model type did not provide a better fit in this study. Interestingly, the local anesthetic effect seems to follow systemic plasma concentrations. This indicates that the plasma concentration proportionally is proportional to the drug concentration at the effect site, with no observable lag in the distribution between plasma and effect site.
In conclusion, we developed a population PK/PD model to describe the biphasic systemic absorption of subcutaneously administrated bupivacaine in a microparticle formulation by two parallel absorption processes: a fast process with 0-order absorption and a slow process with two transit compartments and first-order, Erlang-type absorption. The model had two compartments with first-order elimination and a linear relation between plasma concentration and effect measured as warmth detection threshold. The highest inter-individual variability (>50%) was seen for the duration of the fast absorption process, the fraction of dose absorbed by the fast process, and the slope of the linear relation between plasma concentration and effect. The slow absorption process was dose-dependent and limited apparent clearance (CL/F) at larger doses (flip-flop kinetics), as seen by implementing dose relative to the lowest dose of 150 mg as a covariate on the transit rate constant (k tr) and CL/F with shared covariate effect. With this model, k tr and CL/F can be estimated for any dose by multiplying the population estimates of 0.117 h-1 and 35.4 L/h, respectively, with the factor (Dose/150 mg)-0.43. This model may be used for further development of extended-release local anesthetics with caution given to the large variability on certain parameters and the homogeneity of the subject population.
Acknowledgements