Stereoselective uptake of β‐lactam antibiotics by the intestinal peptide transporter

Abstract

1 The stereoselective transport of fi-lactam antibiotics has been investigated in the human intestinal epithelial cell line, Caco-2, by use of D-and L-enantiomers of cephalexin and loracarbef as substrates. 2 The L-isomers of cephalexin, loracarbef and dipeptides displayed a higher affinity for the oligopeptide/H+-symporter in Caco-2 cells than the D-isomers. This was demonstrated by inhibition of the influx of the f0-lactam, [3H]-cefadroxil.3 By measurement of the substrate-induced intracellular acidification in Caco-2 cells loaded with the pH-sensitive fluorescent dye BCECF (2',7'-bis(2-carboxyethyl)-5-(6)-carboxy-fluorescein), it was demonstrated for the first time that L-isomers of f0-lactams not only bind to the peptide transporter with high affinity but are indeed transported.4 Efficient proton-coupled transport of L-fl-lactam antibiotics was also shown to occur in Xenopus laevis oocytes expressing the cloned peptide transporter PepTi from rabbit small intestine. 5 Both cell systems therefore express a stereoselective transport pathway for fi-lactam antibiotics with very similar characteristics and may prove useful for screening rapidly the oral availability of peptidederived drugs.Keywords: Proton-coupled transport; dipeptide; cephalosporin; Caco-2 cells; enterocytes; intracellular pH; f-lactam antibiotics Introduction f-Lactam antibiotics are bactericidal agents that act by inhibition of bacterial cell wall synthesis. They exert their effects by interrupting the transpeptidation and hence the linkage of individual peptidoglycan chains of the bacterial cell wall (Boyd, 1982;Neu, 1985). The fi-lactam group encompasses the penicillins, cephalosporins, carbapenems and monobactams (Donowitz & Mandell, 1988). The general structure of the cephalosporins resembles the backbone of a tripeptide with the second peptide bond incorporated-into a lactam ring. The excellent oral availability in particular of aminocephalosporins is explained by the fact that they serve as substrates for the intestinal peptide transporter(s). In the apical membrane of intestinal epithelial cells and the human colon carcinoma cell line, Caco-2, di-and tripeptides (Ganapathy et al., 1984;Thwaites et al., 1994b), peptide-mimetics, including selected filactam antibiotics (Tsuji et al., 1986;Dantzig et al., 1994a), immunostimulants (Saito & Inui, 1993; Takano et al., 1994) and angiotensin-converting-enzyme inhibitors (Boll et al., 1994;Thwaites et al., 1994a), are transported by a common electrogenic H+/peptide symport system. Although an interaction of L-isomers of peptides and f-lactam antibiotics with the substrate binding site of the intestinal peptide transporter(s) has been demonstrated by competition experiments in tissue preparations (Matthews & Adibi, 1976;Matthews, 1987;Tamai et al., 1988), brush border membrane vesicles (Rajendran et al., 1985;Tsuji et al., 1986) and Caco-2 cells (Dantzig & Bergin, 1990;Thwaites et al., 1994a) direct evidence for a stereoselective transport of these compounds was absent. In addition, studies wit...

The Extracellular pH Dependency of Transport Activity by Human Oligopeptide Transporter 1 (hPEPT1) Expressed Stably in Chinese Hamster Ovary (CHO) Cells: A Reason for the Bell-Shaped Activity versus pH

“…This activity is more than enough to cause the intracellular acidification due to the influx of the transporter which can be detected using pH-sensitive fluorescent dye. 5) This activity also opens an avenue to determine the influx of the substrate with an electrophysiological technique or a whole-cell patch clamp technique.…”

“…1) Therefore, a majority of protein digestion products are obligatorily absorbed via an intestine oligo-peptide transporter called PEPT1. Of interest, PEPT1 also mediates the absorption of a huge number of pharmacologically important compounds including peptidomimetic drugs, 2,3) such as oral b-lactam antibiotics, [4][5][6] ACE-inhibitors like captopril, 7) or the peptidase inhibitor bestatine 8) and even compounds without an obvious peptide bond or equivalent, 9) such as d-amino levulinic acid. 10) This surprisingly high tolerance for structural diversity of the substrate by PEPT1 has been expected to be used as a channel to increase the intestinal absorption of poorly absorbable drugs.…”

PepT1-mediated epithelial transport of dipeptides and cephalexin is enhanced by luminal leptin in the small intestine

“…This is particularly important because the PepT1 transport system can also be used to transfer a range of orally administered peptidomimetic drugs into the blood (25)(26)(27)(28).…”

Design, Synthesis and Preliminary Pharmacological Evaluation of New Metformin Derivatives

“…Several studies on di/tripeptide transport (PepT1) revealed that a largescale absorption of dipeptides takes place in the human and animal intestine with stereochemical specificity [11]. Natural amino acids are mainly of the L-form and peptides consisted of L-enantiomers also show the highest affinity to the transporter [12]. As a result of the wide substrate specificity range of PepT1, it is an interesting idea to modify a compound with a low intrinsic bioavailability with a promoiety so as to make it acceptable for the transport across the intestinal wall [10].…”