AAPS 2011 Meeting and Exposition
Skin-PAMPA: A New Method for Fast Prediction of Skin Penetration
Purpose. Investigation of the transdermal drug delivery route has intensified recently for both novel and existing drug products. The aim of this work was to adopt the parallel artificial membrane permeability assay (PAMPA) for predicting skin penetration by employing an artificial membrane mimicking the stratum corneum layer of human skin.
Conclusions. The Skin-PAMPA model predicts human skin permeability reasonably well, and because of its standardization potential and high-throughput nature it can be a valuable cost effective alternative to Franz cell studies for early skin penetration prediction.
Dynamic Biorelevant Media Change in Miniaturinzed High Throughput Solubility Assay
Purpose. This study applies to standard 96-well microtitre plate format solubility measurements in fasted and/or fed state simulated intestinal fluids (FaSSIF/FeSSIF). It also introduces a novel automation friendly method of in situ dynamic conversion from simulated gastric fluid (SGF) to FaSSIF while measuring solubility in both media at up to 3 time points.
Conclusion. The high throughput UV-based µSOL method is suitable for accurate solubility measurements in FaSSIF and FeSSIF media in early drug discovery stage and provides better insight into potential in vivo absorption behavior of the drug candidates. The dynamic assay medium adjustment method allows combination of solubility measurements in SGF and FaSSIF in a single setup thereby minimizing sample consumption. This method provides valuable information about the kinetic of precipitation for the basic compounds and the dissolution of acid compounds relevant under in vivo conditions.
Preparation, Physical Characterization and Solubility-Dissolution Testing of Loratidine Hot Melt Extrudates Prepared on a Laboratory Mixing Extruder
Purpose. To prepare and test hot melt extrudates of Loratidine with Copovidone at 30% and 40% drug loading using a bench-top Dynisco laboratory mixing extruder (LME) at various temperatures. Dissolution behavior and solubility enhancement was tested using a small-volume in situ UV dissolution apparatus μDISS Profiler to collect concentration-time profiles in biorelevant media (e.g., FaSSIF and FeSSIF).
Conclusion. Temperature was an important process parameter in preparing amorphous solid dispersions of Loratidine with Copovidone at 30% or 40% API loading on the Dynisco LME. The μDISS Profiler allowed real-time concentration monitoring in small volumes of media thus providing a valuable tool for studying meta-stable solubility of amorphous form of API.
AAPS 2011 Emerging Oral Delivery Strategies and Technologies Workshop
High Throughput Solubility Assay in Simulated Intestinal Fluids
Purpose. It has been demonstrated that solubility of practically insoluble compounds can increase several orders of magnitude in more physiologically relevant media like fasted and/or fed simulated intestinal fluids (FaSSIF/FeSSIF) thus providing better understanding of in vivo absorption behavior. It was argued that some drugs could be re-classificatied from BCS Class II into BCS Class I based on their solubility in FaSSIF and FeSSIF.
This study adapts solubility measurements in FaSSIF/FeSSIF media to 96-well microtitre plate format enabling screening much earlier in drug discovery/development process.
Conclusions. Most of the compounds drastically increased their solubility in FaSSIF and FeSSIF comparing to the corresponding values in FaSSIFblk and FeSSIFblk. For example, solubility of tolfenamic acid increased more than 800 times in FeSSIF (71 µg/mL) compared to its solubility in FeSSIFblk (0.09 µg/mL) while for danazol the ratio between solubility values in FeSSIF (68 µg/mL) and FeSSIFblk (0.23 µg/mL) was around 300. Tamoxifen was fully dissolved in FeSSIF up to a concentration of 147 µg/mL, despite being very poorly soluble in FeSSIFblk (2.3 µg/mL). Solubility enhancement in FaSSIF ranged from ~ 1 (astemizole) to ~ 95 (danazol).
AAPS 2010
Evaluating Drug Salt Dissolution and Precipitation Processes for Rational Formulation Strategies
Purpose. The salt form of an API (active pharmaceutical ingredient) may dissolve, partially or completely, to transient concentrations above equilibrium solubility levels. The supersaturated solution may then precipitate as the free acid/base, sometimes coating the remaining input API or formulation, thus significantly modifying the subsequent dissolution rate. The timeframe of supersaturation and potential precipitation events may have profound effects on bioavailability in vivo. The aim of this study was to develop a practical and predictive in vitro powder and formulation dissolution method to monitor dissolution and concomitant precipitation processes in biorelevant media.
Conclusions. This novel in situ monitoring approach to in vitro powder and formulation dissolution/precipitation can provide a fundamental understanding of processes that may be relevant in vivo providing rational approach to formulation design and development.
Evaluation of Optimized Caco-2 Permeability Measurements
Purpose. To optimize pH conditions for Caco-2 assay that allow de-convoluting and understanding various contributions to passive transport of drugs through the cell membrane. The pH and concentration effects on carrier-mediated/active transport were also studied.
Conclusion. A novel approach of optimizing Caco-2 conditions based on predicted results enables the interpretation of difficult measurements with greater level of confidence. The developed approach saves costs, by avoiding poorly-designed assay protocols, and in some cases, avoiding the need for costly measurements.
Novel Device and Method for in situ UV Dissolution and Precipitation Monitoring in 96-well Plate
Purpose. A newly developed device was applied to assess the feasibility of performing in situ UV concentration monitoring experiments in 96-well microtitre plate format.
Conclusion. The study confirmed the feasibility of performing in situ dissolution and solubility monitoring in 96-well plate that was not possible before. In addition to high throughput solubility determination, other critical studies such as salt dissolution screening, dissolution/solubility determination in biorelevant media, solubility-excipient screening and others can be performed much earlier in the drug discovery saving costs by dramatically reducing required amount of compounds and expensive reagents.
Human Jejunal Permeability Predicted from Caco-2 Assay - A Biophysical Model Applied to Fluoroquinolone Antimicrobials
Purpose. The objective of the study was to predict human jejunal permeability, Peff, and absorption, %Fa, for a series of fluoroquinolones, using a biophysical model based on measured Caco-2 permeability. Values of Peff have not been reported for these compounds, and not all of the compounds have reported human absorption values.
Conclusion. Estimates of Peff can be predicted by the biophysical model. From these values, the human absorption may be calculated. Where absorption comparisons were possible, the agreement was acceptably good.
In Vitro-In Vivo Correlation (IVIVC): Prediction of pK a at 37 C, From the Measured Value at 25 C']);">To Improve In Vitro-In Vivo Correlation (IVIVC): Prediction of pKa at 37 ºC, From the Measured Value at 25 ºC
Purpose. Most of the drug pKa values have been determined at 25°C, instead of physiologically-relevant 37°C. The aim of this study is to develop a simple model to predict drug pKa values at 37oC from their corresponding 25oC values. This is expected to improve the in vitro-in vivo correlation (IVIVC) in some cases, and get an improved understanding of the drug absorption process.
Conclusion. The model we developed is simple to apply to investigative dissolution rate studies done at 37oC, to mechanistic cellular studies, such as Caco-2, as well as to other physiological condition-based measurements.
Development of PAMPA Model for Skin Penetration of Drugs
Purpose. Transdermal drug delivery has extensively grown in the past decades. Efforts in drug research have been devoted to find a useful model for predicting the skin penetration of new molecules. The developed in vitro methods either apply real human skin (for example: Franz Diffusion Cell; Saarbrücken Penetration Model) or use artificial model of human skin. The in vivo methods use animals like rats, pigs or cattle for studying the skin absorption. Neither of these methods are high throughput techniques that is a disadvantage for the drug discovery. Our aim was to extend application of parallel artificial membrane permeability assay (PAMPA) for prediction of skin penetration.
Conclusion. PAMPA assay, as a passive permeation screen method, has been shown to be a good high throughput alternative to in vivo and cell-based in vitro permeability screening for predicting human intestinal absorption and the rate of penetration through the blood-brain barrier. Developed skin PAMPA model is believed to be able to replace the expensive and labour-intensive animal or human skin measurements in early stage of drug discovery.
Very Thin PAMPA Membranes Indicate Higher Antipyrene Permeability but are Fragile and Contain Leaky Water Pores
Purpose. The aim of the study was to assess how thin PAMPA membrane barriers could be made before they become too fragile or leaky through water pores.
Conclusion. It was shown that antipyrine can have an elevated PAMPA permeability value (from 1 to 5 x10<sup>-6</sup> cm/s) in the plates with the thinnest barrier (BD), but it was clear that the plates were too leaky through water pores. This would limit the application to lipophilic molecules.
GRC 2010
In Vitro & In Situ Brain Perfusion Permeability Models of the BBB - Rate and Extent of Brain Penetration
Purpose. The persistent difficulty of delivering therapeutic molecules across the blood-brain barrier (BBB) to achieve optimal CNS exposure continues to be a formidable challenge in the neuropharmaceutical industry. During drug discovery, costly in vivo measurements of brain penetration are impractical, given the large number of molecules tested. This necessitates an ongoing search for simple and cost-effective in vitro and in silico models to predict both the rate and extent of BBB permeation. The goal of this study was to advance the science, to refine and improve on what is currently available.
Conclusion. This new PAMPA-BBB model can mimic the physicochemical microenvironment of the BBB governing passive permeability of basic drugs, with SC = 0.96, using rodent in situ brain perfusion data as a benchmark. The in combo PAMPA-BBB technique improved the general performance of all classes of compounds (n=185); it also predicted the intrinsic BBB permeability of 97 test set BBB data. This is a practical, low-cost, and rapid, quantitative method suitable for early passive BBB permeability screening. It is also useful for structure modification to improve the performance of test compounds in CNS discovery.
AAPS 2009
High Throughput Assay for Pre-Selecting Excipients Enhancing the Solubility of Low Soluble Compounds
Purpose. A newly developed assay was applied for fast in vitro screening of the effect of various solubilizing vehicles (e.g., 2-hydroxypropyl-b-cyclodextrin (HP-b-CD), polyethylene glycol 400 (PEG400), hydroxypropyl-methylcellulose (HPMC), etc.) and their mixtures on apparent solubility and absorption potential of sparingly soluble compounds.
Conclusion. A new in vitro method facilitates studying of the complex role excipients play in absorption. This low cost high throughput assay enables much more efficient pharmacokinetic studies by pre-selecting most promising formulation vehicles.
Prediction of in situ Blood-Brain Barrier Permeability using BBB PAMPA Model
Purpose. This study was conducted to assess the permeability of the blood-brain barrier (BBB) to drug-like compounds with a wide range of molecular properties, using the in situ brain perfusion technique in P-glycoprotein (Pgp) deficient [mdr1a(-/-)] mice, and to predict this permeability using an in combo PAMPA (in silico supported parallel artificial membrane permeability assay) model.
Conclusion. The in situ brain perfusion measurements of drug-like molecules, in Pgp deficient mice, can be predicted by an in combo PAMPA method to a satisfactory degree, based on a training set of in situ BBB measurements. The speed of the PAMPA method, and its substantial lower cost, compared to in vivo measurements, make it an attractive first-pass screening method for BBB passive permeability.
Miniaturization of Powder Dissolution Measurement and Estimation of Particle Size
Purpose. This research investigated the applicability and limitations of a novel approach for measuring intrinsic dissolution rates (IDR) of very small quantities of compounds introduced as powders to buffered solutions and compare these results to disk IDR obtained using the traditional Wood’s apparatus1.
Conclusion. Powder IDR measurements can be used in the early stages of drug development, as they use up to 1000 times less API than the traditional Wood’s rotating disk apparatus and are much faster, too. Application of the powder dissolution method to compounds with solubility >1 mg/mL requires additional study.
Impact of Biorelevant Media on Apparent Solubility and Biopharmaceutical Classification of Poorly Soluble Compounds
Purpose. The goal of this study was to measure the apparent solubility of a diverse series of poorly soluble compounds in four different biorelevant dissolution media (BDM, eg. FeSSIF, FaSSIF, and blanks for each) by simulating intestinal conditions to investigate the result in potential BCS classification. The µDISS Profiler™ allowed simultaneous measurement of dissolution rate and solubility in small volumes of BDM.
Conclusion. All seven compounds displayed higher solubility in FeSSIF than in the corresponding blank buffer. Increased solubility in the BDM resulted in an improved rating in the BCS for cinnarizine, felodipine, indomethacin, terfenadine from class II to class I. Danazol and glibenclamide remained unchanged.
AAPS 2008
Progress in BBB Prediction Using Artificial Membrane Models
Purpose. This study was conducted to investigate the predictability of a specially designed in combo BBB PAMPA model (in silico supported parallel artificial membrane permeability assay) on a set of marketed central nervous system (CNS) drugs with data recently published on rat in situ brain permeability and the fraction unbound to brain tissue (Summerfield et al., J. Pharmacol. Exp. Ther., 2007, 322, 205 – 213).
Conclusions. The in situ brain perfusion measurements of drug-like molecules in Pgp deficient mice can be predicted by an in combo BBB PAMPA method to a satisfactory degree, based on a training set of in situ BBB measurements. The speed of the PAMPA method, and its substantial lower cost compared to in vivo measurements, make it an attractive first-pass screening method for BBB passive permeability.
Powder Intrinsic Dissolution Rate Measurement: When is it Possible?
Purpose. This research investigated the applicability and limitations of a novel approach for measuring intrinsic dissolution rates (IDR) of very small quantities of compounds introduced as powders to buffered solutions and compare these results to disk IDR obtained using the traditional Wood’s apparatus.
Conclusion. Powder IDR measurements can be used in the early stages of drug development, as they use up to 1000 times less API than the traditional Wood’s rotating disk apparatus and are much faster, too. Application of the powder dissolution method to compounds with solubility >1 mg/mL requires additional study.
Progress in pKa Determination Technology: Case Studies in Low and High Soluble Flavonoids
Purpose. This study aimed to investigate the accuracy of software to predict the ionization constants, pKa for a variety of flavonoids.
Conclusion. Flavonoids exhibit several challenges to accurate pKa measurement, including low solubility and potential oxidative degradation. Cosolvents ensured that the compounds were completely in solution during the titration. While the assignments made herein are tentative, they suggest that the beta hydroxy group may undergo internal hydrogen bonding that may be incorporated into some commercially available software programs, thus providing better agreement between measured and predicted values. Future work will focus on investigating some of the underlying chemistry further.
Novel in vitro PK (non-animal PK) Assay for Excipient/Vehicle Formulation Studies
Purpose. A newly developed assay was applied for fast in vitro screening of the effect of various vehicles (e.g., 2-hydroxypropyl-b-cyclodextrin (HP-b-CD), polyethylene glycol 400 (PEG400, etc.) on solubility and absorption of sparingly soluble compounds.
Conclusion. A new in vitro PK method facilitates studying of the complex role excipients play in absorption. Sometimes, individual measurements of permeability and solubility, a task that is especially tedious for sparingly soluble compounds, can be avoided. This practical, low-cost, and relatively high-throughput UV-based assay incorporates the advances achieved in Double-Sink™ PAMPA and is demonstrated to be useful in early lead-investigation screening for formulation effects on absorption.
Comparison of Miniaturized Intrinsic Dissolution Rate Measurement to Traditional Wood's Apparatus
Purpose. The objective was to investigate the feasibility of using a miniaturized disk intrinsic dissolution rate (IDR) apparatus to determine the Biopharmaceutics Classification System (BCS) solubility class while significantly reducing compound consumption comparing to traditional Wood’s apparatus. Additional goal was to develop an approach where IDR measurements performed in media of different buffer capacity could be compared.
Conclusion. The results demonstrate that using 100-fold less drug does not sacrifice the quality of the measurement, and lends support to an earlier study1 that the disk IDR measurement may possibly serve as a surrogate for the BCS solubility classification. API-sparing miniaturized IDR measurements can be done much earlier in the drug development cycle.
Prediction of Caco-2 pH-Dependent Permeability based on High Quality in vitro Training Set
Purpose. The goal of the study is to predict the pH-dependent permeability profiles of drugs in a Caco-2 assay, based on an in combo procedure using measured PAMPA permeability values and calculated Abraham (alpha, beta, pi, R, Vx) descriptors.
Conclusion. The ability to predict with confidence how Caco-2 permeability will depend on the physicochemical properties of a drug is important in both planning measurements of practically-insoluble molecules and interpreting the results of difficult measurements. The method we developed appears to be useful in saving cost, by avoiding poorly-designed assay protocols, and in some cases, altogether avoiding the need for the costly measurements.
Determination of the Intrinsic Dissolution Rate (IDR) from a Powder and using a Miniaturized Wood's Apparatus
Purpose. The objective of the study was to investigate the properties of several of the polymorphs of sulfathiazole using a new miniaturized powder intrinsic dissolution rate (IDR) apparatus, and to relate the IDR values to solubility.
Conclusion. This study successfully demonstrated the application of the new method to simultaneously determine two important properties of the polymorphs of sulfathiazole: (1) intrinsic dissolution rate, and (2) pH-dependent solubility. Powder IDR measurements can be used in the early stages of drug development since they use up to 1000 times less API than traditional Wood’s rotating disk apparatus and are much faster.
Dissolution Rate and Apparent Solubility of Poorly Soluble Compounds in Biorelevant Fluids
Purpose: To experimentally determine dissolution rate and solubility of a number of poorly soluble drugs with a new miniaturized method.
Conclusions: The miniaturized dissolution method is suitable to use also when measuring truly poorly soluble compounds and using biorelevant media. The IDR showed excellent correlation with the apparent solubility values, indicating that the IDR can be used as a substitute for measurement of solubility. Hence, the method allows rapid assessment of dissolution rate and solubility using a small amount of compound only.
