Article Figures & Data
Figures
- Fig. 1.
Relationship between Papp values and intestinal availability (Fa or FaFg) for 21 drugs. hiPSC-SIECs (A) or Caco-2 cells (B) were incubated with transport buffer containing each of the 21 different drugs for 90 minutes at 37°C. The regression curves between the Papp and Fa or FaFg of these 21 drugs were fitted to the following formula in accordance with previous reports (Takenaka et al., 2016; Amidon et al., 1988): Fa or FaFg = 1 − e−Psf×Papp. Compounds used were as follows: 1, piroxicam; 2, carbamazepine; 3, verapamil; 4, warfarin; 5, antipyrine; 6, cephalexin; 7, metoprolol; 8, propranolol; 9, acebutolol; 10, ribavirin; 11, metformin; 12, hydrochlorothiazide; 13, terbutaline; 14, cimetidine; 15, enalapril; 16, ranitidine; 17, atenolol; 18, sulpiride; 19, pravastatin; 20, methotrexate; and 21, raloxifene. Data are presented as means for hiPSC-SIECs (n = 2) and means ± S.D. for Caco-2 cells (n = 3).
- Fig. 2.
Relative expression levels of genes encoding UGT isoforms of hiPSC-SIECs and Caco-2 cells. Real-time reverse transcription polymerase chain reaction analysis of the expression levels of UGT isoform-related proteins. mRNA levels were normalized to those of glyceraldehyde-3-phosphate dehydrogenase mRNA gene expression levels in the hPECs, the levels of which were arbitrarily defined as 100. The gene expression levels of hPECs were measured in and averaged across two separate lots. Data are presented as means ± S.D. (n = 3) for hiPSC-SIECs and Caco-2 cells.
Tables
Gene name Forward primer Reverse primer UGT1A1 5′-AATAAAAAAGGACTCTGCTATGCT-3′ 5′-ACATCAAAGCTGCTTTCTGC-3′ UGT1A6 5′-CATGATTGTTATTGGCCTGTAC-3′ 5′-TCTGTGAAAAGAGCATCAAACT-3′ UGT1A8 5′-GAAAGCACAAGTACGAAGTTTG-3′ 5′-GGGAGGGAGAAATATTTGGC-3′ UGT1A9 5′-TGGAAAGCACAAGTACGAAGTATATA-3′ 5′-GGGAGGGAGAAATATTTGGC-3′ UGT1A10 5′-GAAAGCACAGGCACAAAGTATA-3′ 5′-GGGAGGGAGAAATATTTAGCAAC-3′ UGT2B7 5′-GGAGAATTTCATCATGCAACAGA-3′ 5′-CAGAACTTTCTAGTTATGTCACCAAATATTG-3′ UGT2B15 5′-CTTCTGAAAATTCTCGATAGATGGAT-3′ 5′-CATCTTTACAGAGCTTGTTACTGTAGTCAT-3′ Glyceraldehyde-3-phosphate dehydrogenase 5′-TCCACTGGCGTCTTCACC-3′ 5′-GGCAGAGATGATGACCCTTTT-3′ Compound Papp hiPSC-SIECs Caco-2 cells 10−6 cm/s 10−6 cm/s Daidzein 1.6 36.8 ± 1.6 Genistein 2.7 33.7 ± 1.0 Quercetin 0.46 9.49 ± 0.64 Curcumin 0.07 3.50 ± 0.81 Epicatechin 0.13 0.12 ± 0.08 Epigallocatechin 0.34 0.29 ± 0.06 Epigallocatechin gallate 0.16 0.12 ± 0.03 Caffeic acid 1.03 0.18 ± 0.05 Gallic acid 0.41 0.24 ± 0.05 Bisphenol A 4.3 29.7 ± 2.2 Bisphenol S 5.9 23.4 ± 0.7 MeIQx 1.36 7.59 ± 0.15 Acrylamide 23.3 32.3 ± 0.8 Fenitrothion 34.0 25.4 ± 1.3 Picloram 3.88 2.98 ± 0.15 The cells were incubated with transport buffer containing each of the food-derived compounds at 37°C for 90 minutes. The solution was collected from the basal chambers at every 30 minutes. Unchanged substrates were measured using ultraperformance liquid chromatography–tandem mass spectrometry. Data are presented as means for hiPSC-SIECs (n = 2) and means ± S.D. for Caco-2 cells (n = 3).
- TABLE 3
The amounts of unchanged compounds and metabolites after the membrane permeability assays in hiPSC-SIECs and Caco-2 cells.
Unchanged (basal)2.638.2 ± 5.2 Unchanged (basal)49504 ± 31 Unchanged (basal)167450 ± 14 Unchanged (basal)111325 ± 48
Compounds Amounts hiPSC-SIECs Caco-2 cells pmol/1.5 h pmol/1.5 h Raloxifene Glucuronide (apical) 27.0 7.3 ± 0.5 Glucuronide (basal) 68.6 7.5 ± 0.4 Daidzein Glucuronide (apical) 238 8.3 ± 0.6 Glucuronide (basal) 202 2.8 ± 0.1 Genistein Glucuronide (apical) 221 23.7 ± 1.3 Glucuronide (basal) 158 3.5 ± 0.2 Bisphenol A Glucuronide (apical) 65.1 16.4 ± 2.0 Glucuronide (basal) 879 26.4 ± 3.5 Sulfate (apical) 77.0 53.7 ± 2.5 Sulfate (basal) 21.7 10.2 ± 1.1 The cells were incubated with transport buffer containing each of the food-derived compounds at 37°C for 90 minutes. Unchanged substrates and metabolites were measured using ultraperformance liquid chromatography–tandem mass spectrometry. Data are presented as means for hiPSC-SIECs (n = 2) and means ± S.D. for Caco-2 cells (n = 3).
- TABLE 4
Comparison of metabolic rates of typical UGT substrates and food-derived compounds between hiPSC-SIECs, Caco-2 cells, and hPECs.
Compound, metabolite Metabolic rate hiPSC-SIECs Caco-2 cells hPECs pmol/2 h/mg protein pmol/2 h/mg protein pmol/2 h/mg protein 7-Hydroxycoumarin Glucuronide 47.2 39.9 ± 1.7 6.10 Sulfate 19.6 4.90 ± 0.40 5.10 Raloxifene 4′-Glucuronide 6.33 0.09 ± 0.01 6.63 6-Glucuronide 10.5 0.025 ± 0.001 0.843 Estradiol 3-Glucuronide 5.98 0.027 ± 0.005 1.59 17-Glucuronide 0.054 0.063 ± 0.015 0.070 Daidzein 7-Glucuronide 60.1 0.43 ± 0.05 8.4 4′-Glucuronide 0.54 0.032 ± 0.003 3.86 Genistein 7-Glucuronide 15.5 0.69 ± 0.02 5.8 4′-Glucuronide N.D. N.D. 0.99 Bisphenol A Glucuronide 27.7 0.70 ± 0.03 1.57 Sulfate 6.10 1.69 ± 0.18 6.08 These cells were incubated with the apical and basal transport buffer for hiPSC-SIECs and Caco-2 cells or incubation medium for hPECs containing 10 µM UGT substrates or food-derived compounds for 120 minutes at 37°C. Supernatants were collected, and metabolites were measured using ultraperformance liquid chromatography–tandem mass spectrometry. The metabolic rates of hPECs were measured in and averaged across two separate lots. Data are presented as means for hiPSC-SIECs (n = 2) and means ± S.D. for Caco-2 cells (n = 3). N.D.: Not detected.
Data Supplement
- Supplemental Table -
Pathway and transporter susceptibility during intestinal absorption and reported F a or F a F g values in humans of 21 tested drugs.
- Supplemental Table -
