(620ca) A Method to Quantify Drug Uptake and Transport in 3D Multicellular Spheroids

Authors: 
McCalla, S., Montana State University
Tripathi, A., Brown University

P-glycoprotein (Pgp) is a regulator of the absorption, distribution, and excretion of clinically important drugs. In diseased tissues, such as the cells of a solid tumor, Pgp is often upregulated and increases the resistance to anti-cancer chemotherapeutics. Numerous inhibitors of Pgp have been identified using 2D monolayers of cells, but there has been poor clinical translation due to a lack of efficacy or unwanted side effects in vivo. In this study, we used three dimensional (3D) multi-layer tumor spheroids which mimic physiological barriers to drug transport in conjunction with a new algorithm to quantitatively study uptake and inward diffusion of fluorescent calcein.  This analysis was applied to study the effect of Pgp efflux inhibitors (verapamil, loperamide, and cyclosporin A) in the 3D environment. While overall uptake of calcein was increased, a positional analysis revealed that samples treated with verapamil and loperamide only increased calcein concentration in the outermost layers of cells, and only samples treated with cylcosporin A had an increased calcein concentration at the spheroid core. This study shows that in addition to blocking drug efflux, Pgp inhibitors influence the transport of drug between cells and to the core of the spheroids.  This method is amenable to many different cell types and may serve as a quantitative 3D model that more accurately replicates in vivo barriers to drug uptake and diffusion.