Utility of 96 well Caco-2 cell system for increased throughput of P-gp screening in drug discovery

• Published in: European journal of pharmaceutics and biopharmaceutics Vol. 58; no. 1; pp. 99 - 105
• Main Authors: Balimane, Praveen V, Patel, Karishma, Marino, Anthony, Chong, Saeho
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01-07-2004
• Subjects: ATP-Binding Cassette, Sub-Family B, Member 1 - metabolism; Caco-2; Caco-2 Cells; Cell Membrane Permeability; Drug Design; Drug Evaluation, Preclinical - instrumentation; Drug Evaluation, Preclinical - methods; Efflux transporter; High throughput; Humans; Inhibitor; P-glycoprotein; Permeability; Pharmaceutical Preparations - metabolism; Substrate; TEER, trans-epithelial electrical resistance; P-gp, P-glycoprotein; A, to B, apical to basolateral; B, to A, basolateral to apical; HEPES, N-2-hydroxyethylpiperazine- N′-2-ethanesulfonic acid; NCE, new chemical entity; Permeability; Efflux transporter; P c, permeability co-efficient; High throughput; P-glycoprotein; Substrate; MRP, multi-drug resistance protein; BCRP, breast cancer resistance protein; ADME, absorption, distribution, metabolism, elimination; Caco-2; HBSS, Hank's balanced salt solution; Inhibitor; LRP, lung cancer resistance protein
• ISSN: 0939-6411; 1873-3441
• DOI: 10.1016/j.ejpb.2004.02.014

The use of Caco-2 cells for screening of discovery compounds for their permeability characteristics and P-glycoprotein interactions is well established and used routinely in pharmaceutical industries world-wide. The screening model involves growing cells on 12 or 24 well transwell format. In this manuscript, we report the use of Caco-2 cells grown on 96 well transwell plates for screening compounds for their potential to interact with P-gp. Bi-directionality studies were performed with known P-gp substrates such as saquinavir, indinavir, vinblastine, vincristine, verapamil, digoxin and taxol. P-gp inhibition studies were also conducted using radiolabeled digoxin as the probe. The results demonstrated that P-gp substrates had efflux ratios (
P
c (B to A)/
P
c (A to B)) in the 96 well format that were comparable to the ratios seen in 12 and 24 well format. Inhibition of digoxin efflux transport in presence of the test compounds (P-gp substrates) demonstrated that 96 well cells express adequate amounts of efflux transporters and perform as well as the 12 and 24 well Caco-2 cells. Thus, the 96 well Caco-2 cell set-up presents a higher throughput permeability model capable of identifying compounds that interact with P-gp and has the potential to significantly increase the efficiency of P-gp screening in early drug discovery.