539.13 - The development of an intestinal microphysiological system to model oral drug disposition
Saturday, April 2, 2022
2:50 PM – 3:00 PM
Room: 113 C - Pennsylvania Convention Center
Introduction: Recent advances in the development of novel testing systems that recapitulate the human microenvironment and microanatomy have advanced drug and chemical screening. Using microphysiological systems, investigators are able to consider the influence of fluid flow, cell-cell communication, extracellular matrix, and 3-dimensional organization in organs-on-a-chip and tissue-engineered organ constructs and microtissues. Advancement of this technology includes the robust characterization of transporter expression and function, often in concert with evaluation of drug metabolizing enzymes and regulatory factors. This session will highlight examples of microfluidic systems and novel tissue cultures that recapitulate human transporter function across a number of organ systems. Speakers will review the potential application of these model systems for drug development and toxicity screening.
Christopher Arian (University of Washington), Edward Kelly (University of Washington), Kenneth Thummel (University of Washington)
The intestine has important gate-keeping functions that can profoundly impact the systemic blood exposure of orally administered drugs. Over the past decade, there has been growing effort to develop advanced in vitro tissue models that better predict in vivo DMET processes. One such novel model is the microphysiological system (MPS), which provides a more physiologically and anatomically relevant epithelial cell culture platform than traditional, static cell culture models. The MPS incorporates flow of media down luminal and abluminal channels, with accompanying shear forces that have been shown to be beneficial for multicellular viability and function. In this study, we cultured the immortalized cell line, LS180, in the Microfluidic ChipShop’s “Fluidic 480” MPS. LS180 cells have demonstrated high metabolic activity, particularly for CYP3A substrates, but display poor performance for monolayer permeability studies due to a lack of tight junction formation and uncertain transporter function. We assessed CYP3A metabolic activity, cell monolayer integrity, and global RNA expression for LS180 cells cultured in the MPS versus traditional TranswellTM culture. To measure CYP3A activity, we incubated LS180 cells cultured MPS and TranswellTM with the CYP3A probe substrate, midazolam, and monitored total product formation and parent midazolam appearance in the basolateral media of the two culturing platforms (with static or dynamic flow). LS180 cells cultured in MPS display a 2-3-fold greater midazolam extraction ratio (ER) compared to LS180 cells cultured in TranswellTM. We then compared cell monolayer integrity in the two culture platforms by measuring the apparent permeability (Papp) of the paracellular transport probe, atenolol. LS180 cells cultured in MPS display a 3-fold lower Papp for atenolol than LS180 cells cultured in TranswellTM, indicating improved barrier integrity and tight junction formation for LS180 cells cultured in MPS. Using RNA-seq, we compared global RNA expression for LS180 cells cultured in MPS versus TranswellTM. There were minimal expression differences for phase I and II metabolic enzymes between the two groups. Interestingly, transporter expression differed greatly between the two groups, with some transporters displaying greater expression in MPS compared to TranswellTM (e.g., P-gp, MRP-2) while others displayed a decrease in expression (e.g., OATP2B1, ENT1). Further testing is required to assess whether the changes in transporter RNA expression leads to changes in transporter protein abundance and localization. The sum of these results suggest that culturing LS180 cells in a MPS improves their performance and ability to predict the disposition of orally administered drugs.
Support or Funding Information
This work was supported in part by WA State funds through an Innovation Pilot Award from the UW Institute for Stem Cell and Regenerative Medicine, as well as a grant from the National Institutes of Health – 2U54 AT008909.
This work was supported in part by WA State funds through an Innovation Pilot Award from the UW Institute for Stem Cell and Regenerative Medicine, as well as a grant from the National Institutes of Health amp;ndash; 2U54 AT008909.
