A major obstacle in the development of many new pharmaceuticals, particularly proteins and peptides, is their absorption across the small intestine mucosal barrier. We have developed a cell culture screen to discover novel compounds that increase the intestinal absorption of poorly transported molecules. The absorption screen uses Caco-2 cells, a human-derived intestinal epithelial cell line, grown as monolayer cultures on a permeable membrane which separates apical (luminal) and basal (serosal) chambers. The baseline absorption of the Caco-2 cell system was measured by adding marker molecules to the apical chamber and measuring their appearance in the basal chamber. The amount of these marker molecules in the basal chamber after a 1 h(37°C) incubation was used as a measure of baseline absorption (< 1% of the amount administered). Known absorption enhancers such as polyoxyethylene-9-lauryl ether (POE 9) increased the transport of both the radiolabeled and fluorescent markers. POE 9 enhanced the transport of mannitol to a greater extent than the fluorescent markers SR 101 (MW 625) and the dextran FD10 (MW 10,000). Transport of the higher molecular weight marker (FD10) was enhanced the least. In the same assay, cytotoxicity of the absorption enhancer was quantitated by measuring the release of cellular lactate dehydrogenase (LDH) into the apical solution. Neither the fluorochromes used for monitoring absorption nor the enhancers themselves interfered with the LDH detection, therefore the effect of agents on absorption and cytotoxicity could be measured simultaneously. The change in the flux of the fluorochromes across the Caco-2 cell monolayer, as well as the cytotoxic effects, were measured for five compounds that have been reported to increase epithelial absorption. This assay has been used as a screen to identify a number of potential absorption enhancers. A modification of the assay was used to assess the reversibility of the increased absorption following removal of the putative absorption enhancer. The cell culture screen should allow the identification of novel nontoxic agents which can be used to facilitate the oral absorption of pharmaceuticals. © 1992.
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