Differential Recognition of β-Lactam Antibiotics by Intestinal and Renal Peptide Transporters, PEPT 1 and PEPT 2

Abstract

This study was initiated to determine if there are differences in the recognition of ␤-lactam antibiotics as substrates between intestinal and renal peptide transporters, PEPT 1 and PEPT 2. Reverse transcription-coupled polymerase chain reaction and/or Northern blot analysis have established that the human intestinal cell line Caco-2 expresses PEPT 1 but not PEPT 2, whereas the rat proximal tubule cell line SKPT expresses PEPT 2 but not PEPT 1. Detailed kinetic analysis has provided unequivocal evidence for participation of PEPT 2 in SKPT cells in the transport of the dipeptide glycylsarcosine and the aminocephalosporin cephalexin. The substrate recognition pattern of PEPT 1 and PEPT 2 was studied with cefadroxil (a cephalosporin) and cyclacillin (a penicillin) as model substrates for the peptide transporters constitutively expressed in Caco-2 cells (PEPT 1) and SKPT cells (PEPT 2). Cyclacillin was 9-fold more potent than cefadroxil in competing with glycylsarcosine for uptake via PEPT 1. In contrast, cefadroxil was 13-fold more potent than cyclacillin in competing with the dipeptide for uptake via PEPT 2. The substrate recognition pattern of PEPT 1 and PEPT 2 was also investigated using cloned human peptide transporters functionally expressed in HeLa cells. Expression of PEPT 1 or PEPT 2 in HeLa cells was found to induce H ؉ -coupled cephalexin uptake in these cells. As was the case with Caco-2 cells and SKPT cells, the uptake of glycylsarcosine induced in HeLa cells by PEPT 1 cDNA and PEPT 2 cDNA was inhibitable by cyclacillin and cefadroxil. Again, the PEPT 1 cDNA-induced dipeptide uptake was inhibited more potently by cyclacillin than by cefadroxil, and the PEPT 2 cDNA-induced dipeptide uptake was inhibited more potently by cefadroxil than by cyclacillin. It is concluded that there are marked differences between the intestinal and renal peptide transporters in the recognition of ␤-lactam antibiotics as substrates.

Transport of amino acid aryl amides by the intestinal H+/peptide cotransport system, PEPT1

“…Under these conditions, cells reached confluence the next day. Uptake of [ 14 C]Gly-Sar was measured at room temperature on the seventh to eighth day after seeding as described earlier [12,13]. The uptake buffer contained 25 mM Mes/Tris, pH 6.0, 140 mM NaCl, 5.4 mM KCl, 1.8 mM CaCl 2 , 0.8 mM MgSO 4 , 5 mM glucose.…”

“…Caco-2 cells (passages 35Ϫ53) were routinely cultured as described previously [12,13] in 75-cm 2 culture flasks with minimum essential medium supplemented with 10% fetal bovine serum, penicillin (100 units/ml), streptomycin (100 µg/ml), and 1% non-essential amino acid solution. Subconfluent cultures were treated for 5 min with Dulbecco's phosphate-buffered saline without Ca ϩϩ and Mg ϩϩ , pH 7.2 followed by a 1-min incubation with 0.05% trypsin/EDTA solution.…”

Screening of genetic variations of SLC15A2, SLC22A1, SLC22A2 and SLC22A6 genes

“…Notably, the SLC15A2 haplotype block in a Japanese cohort was more similar to African-American and European-American populations (Figures 1 and 2a) 8 were in absolute linkage disequilibrium (r 2 ¼1) (Supplementary Figure 1a). SLC15A2 has been shown to recognize a wide spectrum of pharmacologically active compounds, including b-lactam antibiotics, 22 inhibitors of the angiotensin converting enzyme 23,24 and non-peptidic drugs. 24 Genetic variations in SLC15A2 (F350L, S409P and K509R) displayed different K m values for glycyl-sarcosine.…”

Impact of transporter-mediated drug absorption, distribution, elimination and drug interactions in antimicrobial chemotherapy

“…However, only a few cephalosporins (cephalexin and cefadroxil) have been reported to be substrates of PEPT2. 34 Therapeutic use of cephaloridine in humans has been found to cause nephrotoxicity.…”