Drug absorption is the movement of a drug into the bloodstream. Therefore, membrane permeability is an important property to consider during drug design, because is an important factor affecting oral absorption, blood-brain barrier permeation, and transport of drugs into tissues and across cell membranes. The permeability of a drug across a membrane is dependent on the passive permeability as well as the susceptibility of the drug to efflux or uptake by transporter proteins.
AMSbiopharma offers three different intestinal in vitro models to access drug permeability and predict drug absorption:
• Caco-2 permeability model.
• MDCK permeability model.
• Parallel artificial membrane permeability model (PAMPA).
Caco-2 permeability model
Human epithelial Caco-2 cell monolayers were used for the first time to model human intestinal absorption in the late 1980s. Since then, this model has become a standard tool for the prediction of intestinal drug absorption in humans and for mechanistic studies of drug transport. The Caco-2 cell monolayer permeability assay is reliable, easy to carry out, and requires only small quantities of compounds. Originally isolated from a human colon adenocarcinoma, Caco-2 cells undergo spontaneous enterocytic differentiation in culture to resemble epithelial cells of the small intestine. When grown to confluence, cell polarity and tight junctions are established in the Caco-2 cell monolayers, and several active transport systems are expressed as in the walls of the human small intestine. Because of that, the Caco-2 cell monolayer assay has become a standard in vitro model for assessing the intestinal permeability and transport of drug candidates and lead compounds.
The apical and basolateral chambers represent the luminal and blood/mesenteric lymph sides of the gastrointestinal tract, respectively.
In the Caco-2 intestinal epithelial cell assay for intestinal absorption, a solution of a test compound is placed on the apical side of a Caco-2 cell monolayer, and the change of the concentration of the test compound in both chambers is measured by HPLC-MS/MS. Alternatively, the test compound may be added to the basolateral side in order to test for the presence of active transport or efflux in one direction or the other across the Caco-2 cell monolayer.
MDCK permeability model
Madin-Darby Canine Kidney (MDCK) cell line, originally derived from dog kidney epithelial cells, is one such alternative in vitro cell culture model. MDCK cells, like Caco-2 cells, form polarize monolayers with tight junctions when cultured on semiporous membrane. Also, the MDCK cells have lower transephitelial electrical resistance (TEER) and shorter culture times than Caco-2 cells.
The permeability assay is also conducted in a double chamber diffusion apparatus with cells grown on semi-porous filter to represent passage of drug from intestinal lumen into the blood represented by the apical and basolateral chambers, respectively. The permeability of drug candidates can be measured in the AP-to BL (absorption) or BL-to-AP (secretion or efflux) directions.
In addition to intestinal permeability, MDCK permeability has also been found to be a useful predictor of blood brain barrier (BBB) permeability allowing the study of the suitability of your drug candidate for use as central nervous system (CNS) therapeutic.
Use AMSbiopharma MDCK permeability assay to identify intestinal or CNS permeability to find out if your drug candidate is suitable for oral administration or for use as a CNS therapeutic.
Parallel artificial membrane permeability model (PAMPA)
A new high-throughput permeability assay, called the “parallel artificial membrane permeability assay” (PAMPA), was first introduced to investigate passive absorption processes by Kansy in 1998. The PAMPA assay uses two aqueous buffer solution wells separated by an artificial membrane. The artificial membrane consists of a lipid or hexadecane membrane supported by a porous hydrophobic filter plate matrix. At the beginning of the experiment, the test compound is diluted in buffer and placed in the donor well. The compounds are transferred only by passive diffusion from the donor well into the acceptor well through the artificial membrane. Permeation determined with PAMPA using a lipidic or hexadecane artificial membrane provided significant correlations with gastrointestinal absorption in humans. Also, the PAMPA assay can be used to predict passive transport through the blood-brain barrier using a solution of porcine polar brain lipids as the artificial membrane.