Caco-2 permeability filter

Karlsson, J. P. Artursson, P., A method for the determination of cellular permeability coefficients and aqueous boundary layer thickness in monolayers of intestinal epithelial (Caco-2) cells grown in permeable filter chambers, Int. J. Pharm. 7, 55-64 (1991). 

The trend in the industry has been to automate the Caco-2 permeability assay using semi- or fully automated procedures. With such a system it is possible to obtain a throughput in order of approximately 400-500 compounds per week. Automated Caco-2 assay systems are commercially available through Tecan/BD Bioscience and Bohdan Mettler Toledo. In addition, automated systems for maintenance of cell cultures are commercially available, while totally automated systems for both maintenance and culturing of cells grown on permeable filter supports are under development, e.g., by companies such as The Automation Partnership. 

Figure 7.8 Permeation of drugs through oil-soaked microfilters comparisons to Caco-2 permeabilities (dashed curves) [546] (a) oil-free (untreated hydrophilic) filters (b) unstirred water layer permeability versus log MW (c) octanol-soaked (impregnated) filters (d) isopropylmyristate-soaked filters.

Table 13 Influence of Stirring Flow Rate on the Permeability of [14C] Testosterone Across Caco-2 Cell Monolayer/Filter Support in 02/C02 Gas Lift Side-by-Side Diffusion at 37°C...

The identification and characterization of cell culture systems (e.g., Caco-2-cells) that mimic in vivo biological barriers (e.g., intestinal mucosa) have afforded pharmaceutical scientists the opportunity to rapidly and efficiently assess the permeability of drugs through these barriers in vitro. The results generated from these types of in vitro studies are generally expressed as effective permeability coefficients (Pe). If Pe is properly corrected to account for the barrier effects of the filter (PF) and the aqueous boundary layer (PAbl) as previously described in Section II.C, the results provide the permeability coefficient for the cell monolayer... 

Different filter supports have been tested nitrocellulose (NC), polycarbonate (PC), aluminium oxide (AO), polyethylenterephtalate (PET). NC filters have shown reduced non-specific binding compared to AO filters but seem to interact with marker PEG and steroids (Nicklin et al. 1992). AO filters displayed only half of permeability of PC for taurocholic acid. Reproducibility of binding and transport experiments was improved with PC filters (Hidalgo et al. 1989). Pore size is an important factor which needs to be considered and tested CACO-2 cells migrate through pores > 1 xm (Tucker et al. 1992 Hilgers et al. 1990). PET filters are translucent and allow microscopic observation as well as staining procedures. 

Permeability-molecular surface area-in vitro-in silico model The permeability values obtained from the Caco-2 cell monolayers have been traditionally used to devise in silico models for the prediction of drug absorption. In this paper, the use of molecular surface areas as descriptors of permeability and solubility will be reviewed. Moreover, a virtual filter for the prediction of oral drug developability based on the successful combination of in vitro and in silico models of drug permeability and aqueous drug solubility will be discussed. 

Small Intestinal Cell Lines. The most notable, and certainly best characterized, in vitro tool for studying absorption and, to a less extent, intestinal metabolism utilizes Caco-2 cells grown in a confluent monolayer on porous membrane filters and, for the experiments, mounted in diffusion chambers. Under these growing conditions they differentiate spontaneously into polarized enterocyte-like cells possessing an apical brush border and tight Junctions between adjacent cells, thereby retaining many characteristics of the intestinal brush border. The permeability of a compound is determined by the rate of its appearance in the basolateral compartment. 

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