Drug Metab. Pharmacokin. 18 (1): 1–15 (2003). Review Intestinal Absorption of Drugs Mediated by Drug Transporters: Mechanisms and Regulation Toshiya KATSURA and Ken-ichi INUI Department of Pharmacy, Kyoto University Hospital, Faculty of Medicine, Kyoto University, Kyoto, Japan Summary: The absorption of drugs from the gastrointestinal tract is one of the important determinants for oral bioavailability. Development of in vitro experimental techniques such as isolated membrane vesi- cles and cell culture systems has allowed us to elucidate the transport mechanisms of various drugs across the plasma membrane. Recent introduction of molecular biological techniques resulted in the suc- cessful identiˆcation of drug transporters responsible for the intestinal absorption of a wide variety of drugs. Each transporter exhibits its own substrate speciˆcity, though it usually shows broad substrate speciˆcity. In this review, we ˆrst summarize the recent advances in the characterization of drug trans- porters in the small intestine, classiˆed into peptide transporters, organic cation transporters and organic anion transporters. In particular, peptide transporter (PEPT1) is the best-characterized drug transporter in the small intestine, and therefore its utilization to improve the oral absorption of poorly absorbed drugs is brie‰y described. In addition, regulation of the activity and expression levels of drug transporters seems to be an important aspect, because alterations in the functional characteristics andWor expression levels of drug transporters in the small intestine could be responsible for the intra- and interindividual variability of oral bioavailability of drugs. As an example, regulation of the activity and expression of PEPT1 is summarized. Key words: intestinal absorption; transporter; brush-border membrane; basolateral membrane; Caco-2 transepithelial transport properties of various com- Introduction pounds. This cell line forms con‰uent monolayers of The absorption of drugs from the gastrointestinal well-diŠerentiated enterocyte-like cells with functional tract is one of the important determinants for oral properties of transporting epithelia4) and has extensively bioavailability. It has long been considered that intesti- been used to characterize intestinal transport mechan- nal absorption of drugs after oral administration is isms of various drugs.5–8) Using isolated intestinal brush- mediated by a simple diŠusion process, which depends border membrane vesicles or Caco-2 cells, transport on physicochemical properties of drugs such as characteristics of various drugs have been investigated, hydrophobicity and ionizing state. However, there have and the following criteria were usually used to deˆne the been numerous drugs exhibiting higher absorption rates transporter-mediated permeation across the plasma after oral administration than expected from their phys- membrane: saturability, temperature-dependence, icochemical properties. In the 1980's, development of in energy-dependence, cis-inhibition and trans-stimulation vitro experimental techniques such as isolated mem- eŠects by related compounds. Modifying reagents for brane vesicles and cell culture systems has allowed us to amino acid residues of the membrane proteins were elucidate the transport mechanisms of various drugs as sometimes used to see whether transport process was well as nutrients across the plasma membrane of small mediated by transport proteins (transporters). Extensive intestine and renal proximal tubules. The development surveys concerning the intestinal absorption mechan- of methods isolating the brush-border and basolateral isms for various ionic drugs revealed that drug trans- membrane vesicles from these epithelial cells permitted porters were mainly classiˆed into three systems; organ- to characterize the detailed mechanisms involved in the ic cation transport systems, organic anion transport transport of various compounds across each mem- systems and peptide transport systems. However, except brane.1–3) In addition, the human colon adenocarcinoma for a few drugs, less attention had been paid to trans- cell line Caco-2 has been introduced to characterize the porter-mediated drug absorption in drug discovery and Received; January 9, 2003, Accepted; March 6, 2003 To whom correspondence should be addressed: Prof. Ken-ichi INUI,Ph.D.,Department of Pharmacy, Kyoto University Hospital, Sakyo-ku, Kyoto 606-8507, Japan. Tel. ＋81-75-751-3577, Fax. ＋81-75-751-4207, E-mail: inui＠kuhp.kyoto-u.ac.jp 1 2ToshiyaK2 ATSURA and Ken-ichi INUI Table 1. Major transporter families involved in drug absorption and disposition Transporter Family HGNCa) Typical Substrates ABC transporter MDR family (MDR1 etc.) ABCB hydrophobic compounds, anticancer agents, digoxin, immunosuppressants MRP family (MRP2, MRP3 etc.) ABCC anionic conjugates, anticancer agents, methotrexate, pravastatin Peptide transporter PEPT family (PEPT1, PEPT2) SLC15 diWtripeptides, b-lactam antibiotics, bestatin, valacyclovir Monocarboxylic acid transporter MCT family (MCT1 etc.) SLC16 lactic acid, salicylic acid Organic anion transporter OATPWoatp family (OATP-C etc.) SLC21 taurocholic acid, estradiol 17b-glucuronide, sulfobromophthalein, thyroxin, pravastatin Organic ion transporter SLC22 OAT family (OAT1, OAT3 etc.) p-aminohippuric acid, b-lactam antibiotics, estrone-3-sulfate, methotrexate, cimetidine OCT family (OCT1, OCT2 etc.) tetraethylammonium, choline, dopamine, 1-methyl-4-phenylpyridinium, cimetidine OCTN family (OCTN1, OCTN2 etc.) L-carnitine, tetraethylammonium Nucleoside transporter CNT family (CNT1, CNT2 etc.) SLC28 purineWprimidine nucleoside, nucleoside derivatives ENT family (ENT1, ENT2) SLC29 purineWprimidine nucleoside, nucleoside derivatives a) Gene family nomenclature classiˆed by the Human Gene Nomenclature Committee (HGCN). See its homepage (http://www.gene.ucl.ac.ukW nomenclatureWgenefamily.shtml) for details. development stage and in clinical situations such as disposition. drug-drug interactions. In this review, we describe the recent studies concern- During the last decade, molecular biological tech- ing the intestinal absorption of drugs mediated by drug niques have been employed to identify drug transporters transporters. Although the fundamental characteristics responsible for drug absorption from the intestinal of each transporter and the classiˆcation of each trans- lumen. In 1987, Hediger et al.9) ˆrst succeeded in clon- porter family (nomenclature) are not described in detail, ing of the Na＋ Wglucose cotransporter (SGLT1) from several excellent reviews covering such information have rabbit small intestine by means of a functional expres- been published in the last few years.14–26) sion cloning strategy using Xenopus laevis oocytes.10) Transporters Involved in Drug Absorption Thereafter, various transporters for nutrients, neu- rotransmitters and other endogenous compounds have Peptide Transporters: Peptide transporters mediate been cloned using this technique, and PCR analyses H＋-coupled active transport of di- or tripeptides across allowed us to identify homologous genes. Since 1994, the brush-border membranes of the small intestine and when drug transporters such as peptide transporter the renal proximal tubules. The acidic luminal pH PEPT1,11) organic cation transporter OCT112) and or- generated by the Na＋ WH＋ exchanger (NHE3) expressed ganic anion transporting polypeptide oatp113) were ˆrst in the brush-border membrane serves as the driving isolated, many drug transporters have been cloned and force for the transport of small peptides (Fig. 1). Using characterized. It has been demonstrated that drug trans- intact small intestinal preparations, isolated intestinal porters are selectively expressed in pharmacokinetically brush-border membrane vesicles and Caco-2 cells, it has important tissues such as small intestine, liver, kidney been demonstrated that intestinal absorption of peptide- and brain capillary endothelial cells. Therefore, it is like drugs such as orally active b-lactam antibiotics is now well accepted that drug transporters play an im- mediated by the H＋ Wpeptide cotransport system. In portant role in drug absorption and disposition. Table 1 1994, intestinal peptide transporter was ˆrst cloned summarizes the major transporter families considered from rabbit small intestine and designated PEPT1.11) to be important determinants for drug absorption and Thereafter, PEPT1 was cloned and characterized from Intestinal Absorption of Drugs by Drug Transporters 3 L-dopa and its metabolite, dopamine, on the basolateral side was much greater than after addition of a parental compound, L-dopa. In addition, peptidic prodrugs of a novel aminomethyl tetrahydrofuranyl-1 b-methylcar- bapenem (CL 191,121) with L-amino acids such as alanine, valine, isoleucine and phenylalanine have been shown to improve the e‹cacies after oral administra- tion, though PEPT1-mediated transport was not directly demonstrated.44) Another possible approach is to conjugate a dipeptide to parental drugs, thus produc- ing peptidyl prodrugs. Ezra et al. synthesized dipep- tidyl-bisphosphonates, Pro-Phe-pamidronate and Pro- Phe-alendronate, and successfully demonstrated the improvement of oral absorption of these dipeptidyl- bisphosphonates.45) After oral administration, intestinal absorption of Pro-Phe-pamidronate and Pro-Phe- alendronate was increased 3-fold. In addition, the apical-to-basolateral transport of these prodrugs across Fig. 1. Peptide transporters in the small intestine. DiWtripeptides