Drug Transporters in Farm Animals

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Drug Transporters in Farm Animals Membrane transporters in the ATP Binding Cassette Family (ABC) play critical roles in cellular efflux of endogenous metabolites as well as xenobiotics including prescription drugs. A key ABC transporter gene that is present in mammals and other species is ABCB1, which encodes P-glycoprotein (P-gp). Highly expressed in environment-organism barrier tissues such as the intestine, P-gp represents a major impediment to drug absorption limiting the oral bioavailability of its substrates. Accordingly, candidate drugs are screened for their interactions with human P-gp and those with limited or no interactions with P-gp are generally selected for further development. However, P-gp from various mammalian species has been shown to have profound differences in its substrate and inhibitor specificities. Though human P-gp expressing cell lines are available for screening during drug development, currently, cell lines expressing farm animal P-gps are not available. Therefore, companies that are developing drugs to treat animals and in particular farm animals, must rely on cells expressing human P-gp to predict drugs to take forward in costly drug development efforts. The overall goals of this Phase I SBIR research proposal is to create and characterize cell lines stably expressing farm animal P-gps including those from chicken, pig, sheep and cattle. The cell lines will serve as a resource for drug developers to predict the absorption of candidate drugs. A secondary goal is to create and validate a transformed yeast strain expressing human P-gp for use in testing food additives and contaminants in animal feed as potential inhibitors of human P-gp. Two aims are proposed. In aim 1, we will create stable cell lines expressing P-gp from the various farm animals and characterize the interactions of canonical substrates and inhibitors of human P-gp with the various animal P-gps. In aim 2, we will transform yeast with human P-gp (ABCB1) and create a robust assay to use in testing feed contaminants and additives for their ability to inhibit human P-gp and cause potential toxicities to people ingesting animal products. Methods used in the proposed studies include new technologies for creation of cell lines stably expressing the transporter orthologs, fluorescent and isotopic uptake and flux assays in cells, as well as yeast genetics, transformation protocols and cytotoxicity assays. Overall these studies will lead to a set of new tools that can be used in the development and repurposing of drugs for the treatment of diseases of farm animals, and the assessment of ingredients or contaminants in feed for potentially toxic effects on human P-gp.