A bioassay using intestinal organoids to measure CFTR modulators in human plasma  R. Dekkers, L.A.W. Vijftigschild, A.M. Vonk, E. Kruisselbrink, K.M. de Winter-de Groot, H.M. Janssens, C.K. van der Ent, J.M. Beekman  Journal of Cystic Fibrosis  Volume 14, Issue 2, Pages 178-181 (March 2015) DOI: 10.1016/j.jcf.2014.10.007 Copyright © 2014 Terms and Conditions

Fig. 1 Functional activity of CFTR modulators in plasma. (A) Pooled plasma from ten healthy donors, VX-770 and forskolin (0.4μM) were combined to stimulate CFTR-S1251N expressing organoids. Swelling relative to t=0min was measured for 60min, and area under the curve (AUC) data were calculated and normalized to stimulation with 2μM of VX-770 in medium. Data were measured at three independent time points in triplicate and presented as mean (±SEM). (B) Normalized AUC measurements of organoid swelling after stimulation with serial dilutions of pooled plasmas containing VX-770 (0.5, 1.5 and 4μM in 100% plasma) using 0.4μM forskolin. Data were measured at three independent time points in triplicate and presented as mean (±SEM). (C) Representative examples of organoids from (B) at t=0 and t=60min for indicated conditions (scale bar indicates 250μm). (D) Swelling of R117H organoids after stimulation with forskolin (0.05μM) and serial dilution of plasma containing VX-770 (2μM in 100% plasma). Data are presented as mean (±SD). (E) Forskolin-induced (5μM) swelling of F508del organoids after 20h incubation with 10% plasma and VX-809 (0, 0.5, 1 or 2μM). Data were measured in triplicate in four independent experiments and presented as mean (±SEM). Journal of Cystic Fibrosis 2015 14, 178-181DOI: (10.1016/j.jcf.2014.10.007) Copyright © 2014 Terms and Conditions

Fig. 2 Variation in VX-770-dependent organoid swelling between plasma samples. (A) CFTR-S1251N expressing organoids were stimulated with 6% plasma from 9 different donors in the absence or presence of VX-770 (60 nM, representing 1μM in 100% plasma) and forskolin (0.5μM). Individual data points were normalized to the average VX-770 response of the 9 plasma samples. (B) The VX-770-dependent part of organoid swelling was calculated by subtraction of the plasma response from the plasma+VX-770 response per plasma in (A). Data were normalized to the average of all data points. All data were measured at three independent time points in triplicate and presented as mean (±SEM). Journal of Cystic Fibrosis 2015 14, 178-181DOI: (10.1016/j.jcf.2014.10.007) Copyright © 2014 Terms and Conditions