Comparison of free software platforms for the calculation of the 90% confidence interval of f2 similarity factor by bootstrap analysis.

INTRODUCTION: The calculation of the 90% confidence interval of f2 based on the bootstrap methodology has been proposed and accepted by the main regulatory authorities when the dissolution data shows excessive variability. Different free software platforms allow the calculation of the 90% CI of f2 by means of bootstrapping. Their use in regulatory submissions is growing, but divergent results have been observed between the available software platforms. Therefore, the objective of this work is to analyze the characteristics of these software platforms and evaluate their results. METHODS AND MATERIALS: Highly variable in vitro dissolution data from two products were selected. Three different similarity factors, f2, E(f2) and bc-f2, and their corresponding 90% confidence intervals were calculated with three free software platforms, Pheq_bootstrap, Bootf2bca and DDSolver, and computed by four different approaches, normal approximation, bootstrap-t-CI, percentile CI, and bias corrected and accelerated CI. RESULTS: All three platforms report the same f2 value, 49.66 upon comparison of products 1 and 3 and 54.87 for products 2 and 3 (no truncation rule). Bootf2bca and Pheq_bootstrap provided the same f2 and E(f2) also under other truncation rules (EMA or FDA), which are not implemented in DDSolver. Pheq_bootstrap allowed the calculation of bc-f2. The conclusion of similarity is based on a bootstrap percentile CI of E(f2) and f2 in Pheq_bootstrap and DDSolver, respectively. Bootf2bca provides all four 90% CI. DISCUSSION: Bootf2bca or Pheq_bootstrap should be considered for the estimation of the 90% CI of the f2 similarity factor when dissolution profiles show high variability.

A multilevel object-oriented modelling methodology for physiologically-based pharmacokinetics (PBPK): Evaluation with a semi-mechanistic pharmacokinetic model.

BACKGROUND AND OBJECTIVE: The aims of this study are (i) to assess the predictive reliability of the physiologically based software PhysPK versus the well-known population approach software NONMEM for the cited semi-mechanistic PK model, (ii) to determine whether these modelling approaches are interchangeable and (iii) to compare acausal with causal modelling approaches in the framework of semi-mechanistic PK models. METHODS: A semi-mechanistic model was proposed, which assumed oral administration of a solid dosage form with a peripheral compartment and two active metabolites. The model incorporates intestinal transit, dissolution limited by solubility, variable efflux transporter expression along the gut and linear and non-linear metabolism in the gut and liver. Four different approximations to the theoretical model were developed in order to validate both the new software and modelling methodology. RESULTS: Plasmatic concentrations correlation plots as well as relative errors in AUC0-48 and Cmax predictions revealed the accuracy of PhysPK in the prediction of these exposition parameters. Physiological and acausal object oriented version systematically under-estimated AUC0-48 and Cmax of the parent drug, whereas metabolites were over-estimated when taking the semi-mechanistic and extraction-based metabolism version as the reference. CONCLUSIONS: PhysPK has been properly validated, where differences are related to numerical precision of integrators and solvers. A systematic bias for the parent drug and active metabolites was predicted when a semi-mechanistic approach including extraction-based metabolism was compared to the physiologic and acausal approach, showing that interchangeability might be possible when intrinsic-clearance metabolism is implemented in the semi-mechanistic approach. The acausal and object-oriented methodology allows for defining the semi-mechanistic model through its local mechanisms and relationships among entities, without the need to build the final set of Ordinary Differential Equations.

Semi-mechanistic Pharmacokinetic/Pharmacodynamic model of three pegylated rHuEPO and ior®EPOCIM in New Zealand rabbits.

Marketed formulations of erythropoietin (EPO) ior®EPOCIM, MIRCERA® and two newly developed pegylated-EPO analogues (PEG-EPO 32 and 40 kDa) formulations were intravenously administered to New Zealand rabbits. A semi-mechanistic Pharmacokinetic/Pharmacodynamic (PK/PD) model describing in a simultaneous and integrated form the time course of reticulocytes, red blood cells and hemoglobin was built to account for the time course of hematopoiesis stimulation after erythropoietin administration. Data analysis was performed based on the population approach with the software NONMEM version 7.3. Erythropoietin disposition of each of the administered formulations was best described with a two compartment model and linear elimination. Different formulations show different clearance and apparent volume of distribution of the central compartment but share estimates of inter-compartmental clearance and apparent peripheral volume of distribution. A semi-mechanistic model including cell proliferation, maturation, and homeostatic regulation provided a good description of the data regardless the type of erythropoietin formulation administered. The system-, and drug-related parameters showed consistency and differed across formulations, respectively. A single IV administration of PEG-EPO 32 and 40 kDa formulations in New Zealand rabbits achieves a median change of 27% and 22% on RET levels, and of 47% and 63% on RBC and HGB levels, respectively compared to MIRCERA®. The administration of new branched PEG-chains formulations improves PK and PD properties of EPO, in terms of increasing elimination half-lives and pharmacological activity on RET, RBC and HGB compared to commercially available formulations (ior®EPOCIM and MIRCERA®).

Defining level A IVIVC dissolution specifications based on individual in vitro dissolution profiles of a controlled release formulation.

Regulatory guidelines recommend that, when a level A IVIVC is established, dissolution specification should be established using averaged data and the maximum difference between AUC and Cmax between the reference and test formulations cannot be greater than 20%. However, averaging data assumes a loss of information and may reflect a bias in the results. The objective of the current work is to present a new approach to establish dissolution specifications using a new methodology (individual approach) instead of average data (classical approach). Different scenarios were established based on the relationship between in vitro-in vivo dissolution rate coefficient using a level A IVIVC of a controlled release formulation. Then, in order to compare this new approach with the classical one, six additional batches were simulated. For each batch, 1000 simulations of a dissolution assay were run. Cmax ratios between the reference formulation and each batch were calculated showing that the individual approach was more sensitive and able to detect differences between the reference and the batch formulation compared to the classical approach. Additionally, the new methodology displays wider dissolution specification limits than the classical approach, ensuring that any tablet from the new batch would generate in vivo profiles which its AUC or Cmax ratio will be out of the 0.8-1.25 range, taking into account the in vitro and in vivo variability of the new batches developed.

IVIVC approach based on carbamazepine bioequivalence studies combination.

The aim of the present study was to explore the feasibility of obtaining an IVIVC by combination of data from two bioequivalence (BE) studies of carbamazepine (CBZ) in order to assess if the previously published dissolution media and conditions could be applicable to any other oral immediate release (IR) CBZ products with conventional excipients. Twenty-four healthy male subjects from two BE study received one IR dose of the test (test 1 or 2) or the reference formulation (Tegretol, 400 mg). Dissolution studies of the IR CBZ tablets were performed in two different laboratories. In order to develop IVIVC, individual or average data analysis were considered. A level C, level B and level A correlation have been successfully developed by combining data from different BE studies of CBZ immediate release drug products. A level A IVIVC was developed with all four datasets with a good R2 for all the combinations of in vivo and in vitro data. A dissolution medium containing 1% SLS has demonstrated its suitability as the universal biopredictive dissolution medium, even if different batches and in vivo/in vitro studies were combined.

Agitation Rate and Time for Complete Dissolution in BCS Biowaivers Based on Investigation of a BCS Biowaiver for Dexketoprofen Tablets.

The objective of the present work is to investigate the validity of the existing requirements for BCS biowaivers of immediate release products containing a class I drug in relation to the agitation rate (50 or 75 rpm in the paddle apparatus) and the time limit for complete dissolution (30 min) in the current biowaivers in vitro dissolution tests. Further, the possibility of extensions will be examined since it has been proposed that the time limit for complete dissolution should be revised to 60 min, and also, if cone formation occurs with apparatus 2 at 50 rpm, then a higher agitation rate is acceptable to eliminate it. The development of four generic dexketoprofen immediate release tablets is described. Dexketoprofen is the eutomer of ketoprofen. According to the BCS, dexketoprofen is a class I drug. Three out of the four products failed to show bioequivalence for Cmax in the initial bioequivalence study conducted with the product despite similar but nonrapid dissolution profiles at 50 rpm in the paddle apparatus, or similar and very rapid dissolution profiles at 75 rpm. In conclusion, these data indicate that BCS biowaivers for class I drugs should be granted only when dissolution with the paddle apparatus is complete in 30 min at 50 rpm. The time limit for complete dissolution should not be extended to 60 min. Furthermore, the agitation rate should not be increased to 75 rpm, even in the case of a coning effect.

Tubulin acetylation promoting potency and absorption efficacy of deacetylase inhibitors.

BACKGROUND AND PURPOSE: Histone deacetylase 6 (HDAC6) and silent information regulator 2 (SIRT2) control the dynamics of the microtubule network via their deacetylase activities. Tubulin polymerization promoting protein (TPPP/p25) enhances microtubule acetylation by its direct binding to HDAC6. Our objective was to characterize the multiple interactions of the deacetylases and to establish the inhibitory potency and the pharmacokinetic features of the deacetylase inhibitors, trichostatin A (TSA) and AGK2. EXPERIMENTAL APPROACH: The interactions of deacetylases with tubulin and TPPP/p25 were quantified by elisa using human recombinant proteins. The effect of inhibitors on the tubulin acetylation was established in HeLa cells transfected with pTPPP and CG-4 cells expressing TPPP/p25 endogenously by celisa (elisa on cells), Western blot and immunofluorescence microscopy. The pharmacokinetic features of the inhibitors were evaluated by in situ kinetic modelling of their intestinal transport in rats. KEY RESULTS: Deacetylases interact with both tubulin and TPPP/p25, notwithstanding piggy-back binding of HDAC6 or SIRT2 to the TPPP/p25-associated tubulin was established. Much higher inhibitory potency for TSA than for AGK2 was detected in both HeLa and CG-4 cells. Pioneer pharmacokinetic studies revealed passive diffusion and diffusion coupled with secretion for TSA and AGK2 respectively. Both inhibitors exhibited greater permeability than some other well-established drugs. CONCLUSIONS AND IMPLICATIONS: TPPP/p25-directed deacetylase inhibition provides mechanisms for the fine control of the dynamics and stability of the microtubule network. Deacetylase inhibitors with chemical structures similar to TSA and AGK2 appear to be excellent candidates for oral drug absorption.

Ion-pair strategy for enabling amifostine oral absorption: rat in situ and in vivo experiments.

This study shows the effect of ion pair formation on intestinal absorption and oral bioavailability of amifostine. Amifostine is a prodrug used as a highly potent and selective radiotherapy and chemotherapy protectant but due to its low lipophilicity and charge at physiological pH range, its trans epithelial transport and its potential for oral drug delivery is very low. Ion pair formation with negatively charged counter ions was evaluated by in situ rat perfusion studies as a possible strategy to enhance intestinal absorption of amifostine. Succinic acid, phthalic acid and benzoic acid were used as counter ions. Rat intestinal perfusion studies confirmed a statistically significant increase in amifostine permeability in the presence of the counter ions in the order of succinic>phthalic>benzoic. Rat pharmacokinetic studies in vivo were performed to calculate oral absolute bioavailability of amifostine alone and with ion pairs in order to confirm the in situ perfusion results and the applicability of the ion pair approach. Intravenous and intraduodenal administrations were done in rats using a permanent jugular vein cannulation technique and a duodenal cannulation method to avoid drug degradation in stomach. In vivo oral bioavailability studies demonstrated a 20-30-fold increase in amifostine bioavailability with succinic acid depending on counter ion ratio and 10-fold increase with phthalic acid as ion pair. In summary ion pair strategy with succinic acid could enable amifostine oral administration on enteric coated formulations.

Mathematical modeling of oral absorption and bioavailability of a fluoroquinolone after its precipitation in the gastrointestinal tract.

The objective was to characterize the in vivo absorption and bioavailability (BA) of a low solubility, high permeability fluoroquinolone (CNV97101) that precipitates in the gastrointestinal (GI) tract by mathematical modeling approach. In situ rat intestinal perfusion studies were performed to characterize the absorption mechanism. The oral fraction absorbed in vivo was lower than the predicted based on the in situ intestinal permeability. Two additional routes of administration, intraduodenal (ID) and intraperitoneal (IP) were investigated to explore if precipitation in stomach and subsequent partial re-dissolution were the causes of the lower in vivo BA. Ex vivo precipitation studies with the stomach content of fasted rats were also carried out. Fitting procedures were performed with NONMEM VII 1.2. The in situ experiments confirmed simultaneous passive and carrier-mediated absorption processes. The ex vivo experiments confirmed precipitation in stomach lowering in vivo the oral fraction absorbed compared with the IP and ID administrations. Due to the almost complete availability of CNV97101 following IP administration, a first hepatic pass could be excluded. The ex vivo assay results and the pharmacokinetic modeling of in vivo data supported the hypothesis of precipitation in the stomach and partial re-dissolution. Nevertheless, other factors such as residence time in the GI may reduce the fraction absorbed even for low oral doses for which re-dissolution was almost complete in vivo.

In vitro-in situ permeability and dissolution of fexofenadine with kinetic modeling in the presence of sodium dodecyl sulfate.

The purpose of this study was to estimate passive permeability and efflux transport parameters of fexofenadine (FEX) from in vitro cell culture and in situ rat experiments and determine the dissolution profile of FEX in the absence and presence of sodium dodecyl sulfate (SDS). The dissolution rate of FEX was investigated at different pH values and in the presence of SDS. The permeability of FEX was determined in situ in small intestine of Wistar rats and in vitro in Caco-2 cell monolayer. Permeability of FEX was determined at different donor concentrations, and the effect of SDS at two concentration levels (10 and 50 µM) on FEX permeability was evaluated. The transepithelial electrical resistance values of the in vitro technique were measured to assess monolayer integrity before and after the permeability studies. The FEX permeability parameters in the absence and presence of SDS were estimated using Phoenix WinNonlin software program and compared with other laboratory results for both in vitro and in situ studies. The results showed that FEX has a low permeability in the paracellular permeability as well a potential inhibition of secretion mediated by P-glycoprotein (P(gp)), and its permeability increased with presence of SDS. The dissolution of FEX was pH-dependent and significantly enhanced, especially in the presence of 50 mg SDS.

Drug penetration across the blood-brain barrier: an overview.

The brain is one of the most protected organs in the body. There are two key barriers that control the access of endogenous substances and xenobiotics (drugs or toxins) to the CNS. These physiological structures are the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier. The BBB represents the main determinant of the effective delivery of drugs to the CNS. Good access through the BBB is essential if the target site is located within the CNS or, in contrast, can be a disadvantage if adverse reactions occur at central level. The development of new drugs targeted to the CNS requires a better knowledge of the factors affecting BBB permeation as well as in vitro and in silico predictive tools to optimize screening, and to reduce the attrition rate at later stages of drug development. This review discusses the particular characteristics of the biology and physiology of the BBB with respect to the permeation and distribution of drugs into the brain. The factors affecting rate, extent and distribution into the brain are discussed and a brief description of the in silico, in vitro, in situ and in vivo methods used to measure BBB transport are presented. Finally, the lastest proposals and strategies to enhance transport across the BBB of new CNS drugs are summarized.