Analysis of drug transporter expression in human intestinal Caco‐2 cells by real‐time PCR

Abstract

Expression of drug transporters corresponds to a crucial parameter in intestinal Caco-2 cells widely used for investigating drug absorption. In order to characterize it in an accurate, reproducible and comparative manner, we analyzed mRNA levels of 19 influx and efflux drug transporters through real-time quantitative polymerase chain reaction assays combined with the use of a total RNA reference standard. Profiles of transporter expression were found to be significantly correlated in two independent Caco-2 cell clones and in human small intestine, which may support the use of Caco-2 cells for investigating intestinal drug transport. Several transporters were nevertheless quantitatively expressed at higher (MRP2, MRP3, MRP4, MRP5, MRP6, OATP-A, OATP-B, OCT1 and MCT1) or lower (BCRP) levels in Caco-2 cells comparatively to small intestine. Moreover, MDR1, MRP2, OATP-A and PEPT1 mRNA relative expression were found to differ in the two analyzed Caco-2 cell clones by at least a twofold factor, highlighting that some variations in transporter expression may occur in Caco-2 cells depending on cell origin, and therefore underlining the interest of carefully characterizing transporter levels in any Caco-2 cell clone before its use for drug transport assays.

Intestinal transporters for endogenic and pharmaceutical organic anions: the challenges of deriving in-vitro kinetic parameters for the prediction of clinically relevant drug–drug interactions

“…MRP5 ABCC5 N.R. Jejunum, ileum > duodenum [28] Jejunum = colon [14] Jejunum [20] Small intestine [21,32] Colon [28,32] N.R.…”

“…However, it is generally accepted that the absorptive transporters ASBT, MRP3, MCT1, OATP2B1 and OSTa/b have moderate to high intestinal expression levels. [12,[14][15][16]18,20,21,29,32,33,36,39,40] The three known intestinal exsorptive transporters for organic anions (BCRP, MRP2 and OSTa/b) all have relatively high expression levels -especially BCRP has a high intestinal expression compared with other tissues. [14,16,18,20,21,23,39] A high transporter expression level is likely indicative of an important role in the absorption or elimination of endogenous and exogenous substances.…”

“…The intestinal expression of the absorptive transporter OATP1A2 is less evident. Two scientific papers have suggested that OATP1A2 is expressed at mRNA levels in the small intestine, and this transporter has also been [12,13] Jejunum > colon [14] Ileum [15] Ileum > duodenum [13] BCRP ABCG2 Ex Ileum > duodenum, colon [16] Duodenum > ileum > colon [17] Jejunum > colon [14,18] Duodenum [19] Jejunum [20] Small intestine [21] Duodenum = ileum = colon [22] Small intestine and colon [23] MCT1 SLC16A1 Ab Colon > duodenum > ileum [16] Colon > duodenum [24] Colon > jejunum [18] Colon > ileum [25] Jejunum = colon [15] Small intestine [21,26] Colon > ileum > jejunum [24] Colon [25] MCT3 SLC16A8 N.R. N.R.…”

“…Jejunum > colon [14] Colon > ileum, not in jejunum [24] MRP1 ABCC1 Ab Jejunum = ileum = colon [27] Jejunum, ileum > duodenum [28] Jejunum = colon [14] Duodenum [19] Jejunum [20] Small intestine [21,29] Colon [29,30] Small intestine and colon [31] Too low expression in jejunum, ileum and colon for protein determination [29] MRP2 ABCC2 Ex Jejunum > ileum > > colon [27] Duodenum > ileum > > colon [32] Duodenum = ileum > colon [16] Jejunum, ileum > duodenum [28] Duodenum > > colon [33] Jejunum > colon [18] Duodenum [19] Jejunum [16,20] Small intestine [21] Jejunum > ileum = colon [27] MRP3 ABCC3 Ab Colon > Ileum = duodenum [16] Jejunum, ileum > duodenum [28] Colon > jejunum [18] Duodenum [19] Jejunum [16,20] Small intestine [21,32] Colon [28,32] Small intestine and colon [29] MRP4 ABCC4 N.R. Jejunum, ileum > duodenum [28] Jejunum = colon [14] Jejunum [20] Small intestine [21,32] Colon [28,32] N.R. MRP5 ABCC5 N.R.…”

“…Jejunum, ileum > duodenum [28] Jejunum [20] Small intestine [21,34] Colon [34] Small intestine and colon [34] OATP1A2 (OATP-A) SLCO1A2 Ab Duodenum [19] Small intestine [21] NOT in duodenum, ileum or colon [12] NOT in jejunum or colon [14] NOT in small intestine [35] Duodenum [19] OATP2B1 (OATP-B) SLCO2B1 Ab Duodenum = ileum = colon [12,16] Jejunum = colon [14] Jejunum > colon [18] Duodenum [19] Jejunum [36] Small intestine [21,35] Small intestine [37] OATP3A1_v1 (OATP-D) SLCO3A1 N.R. Jejunum [14] Small intestine and colon [35,36] NOT in colon [14] N.R.…”