Characterization of intestinal transporters in human ileal spheroid-derived differentiated cells for the prediction of intestinal drug absorption.

This study aimed to characterize the functional properties of human spheroid-derived differentiated cells for absorption, distribution, metabolism, and excretion studies. Three-dimensional human intestinal spheroids were successfully established from crypts isolated from fresh surgical specimens of the terminal ileum. The mRNA expression of major intestinal transporters and drug-metabolizing enzymes was tested. Region-specific functional expression of proton-coupled folate transporter/solute carrier (SLC) SLC46A1 and apical sodium-dependent bile acid transporter (ASBT)/SLC10A2 was confirmed in freshly isolated human proximal jejunal and terminal ileal tissue sections mounted onto Ussing chambers. Proximal jejunal and terminal ileal spheroid-derived differentiated cell monolayers showed H+- and Na+-coupled uptake of methotrexate and [3H]-taurocholic acid, respectively. The functional expression of CYP3A, CYP2C9, UGT1A, P-glycoprotein, and breast cancer resistance protein (BCRP) was confirmed based on the formation of metabolites (1'-hydroxy midazolam, 4'-hydroxy diclofenac, raloxifene-4'-glucuronide, and raloxifene-6-glucuronide) and the efflux ratio of typical substrates (digoxin, sulfasalazine, rosuvastatin, dantrolene, and furosemide). Terminal ileum-derived differentiated cell monolayers showed extensive taurocholic acid-d5 transport in the apical-to-basal direction, which was markedly inhibited by the ASBT inhibitor elobixibat. One of the terminal ileal spheroids was identified as homozygous for the mutant allele of ABCG2 (rs2231142). The transport activity of BCRP in the differentiated cells, when dantrolene was used as a probe, might reflect this genetic variation. In conclusion, the functional expression of region-specific uptake transporters (proton-coupled folate transporter and ASBT) was successfully reproduced in human intestinal spheroid-derived differentiated cells, which also maintain the activities of P-glycoprotein, BCRP, and metabolic enzymes. Human intestinal spheroid-derived differentiated cells would be useful for investigating region-specific functions in drug transport and metabolism. SIGNIFICANCE STATEMENT: The expression levels of some transporters are not homogeneous along the longitudinal axis of the intestine. To date, there is no in vitro model that can reproduce regional differences in the transporter-mediated transport of nutrients and drugs in the intestine. This study successfully demonstrated that the functional expression of region-specific transporters, the proton-coupled folate transporter and the apical sodium-dependent bile acid transporter, was maintained in human proximal jejunal and terminal ileal spheroid-derived differentiated cell monolayers.