Peptide carriers

H Yuasa, D Fleisher, GL Amidon. Noncompetitive inhibition of cephradine uptake by enalapril in rabbit intestinal brush-border membrane vesicles An enalapril specific inhibitory binding site on the peptide carrier. J Pharmacol Exp Ther 269 1107-1111, 1994. 

Figure 8 A peptide carrier mediated transport to improve oral absorption.

Enalaprilat and SQ27,519 are angiotensin-converting enzyme (ACE) inhibitors with poor oral absorption. Enalapril and fosinopril are dipeptide and amino acid derivatives of enalaprilat and SQ27,519, respectively [51] (Fig. 10). Both prodrugs are converted via deesterification to the active drug by hepatic biotransformation. In situ rat perfusion of enalapril indicated a nonpassive absorption mechanism via the small peptide carrier-mediated transport system. In contrast to the active parent, enalapril renders enalaprilat more peptide-like, with higher apparent affinity for the peptide carrier. The absorption of fosinopril was predominantly passive. Carrier-mediated transport was not demonstrated, but neither was its existence ruled out. 

Hu Ming, HI Mosberg, GL Amidon. Use of the peptide carrier system to improve the intestinal absorption of L-alpha-methyldopa Carrier kinetics, intestinal permeabilities, and in vitro hydrolysis of dipeptidyl derivatives of L-alpha-methyldopa. Pharm Res 6(1) 66—69, 1989. 

M Hu. Investigation into drug and drug analogs transported by the peptide carrier system Intestinal absorption of captopril and of peptidyl derivatives of methyldopa. PhD Thesis, College of Pharmacy, University of Michigan, 1988. 

PJ Sinko, GL Amidon. Characterization of the oral absorption of P-lactam antibiotics. II. Competitive absorption and peptide carrier specificity. J Pharm Sci 78 723-727, 1989. 

E. E., Tukker, J. J., Mapping the binding site of the small intestinal peptide carrier (PepTl) using comparative molecular field analysis, Receptors Channels 1998, 6, 189-200. 

Molecular determinants of recognition for the intestinal peptide carrier, J. Pharm. Sci. 1997, 86, 596-602. 

Swaan, P. W., Stenhouwer, M. C., Tukker, J., Molecular mechanism for the relative binding affinity to the intestinal peptide carrier. Comparison of three ACE-inhibitors enalapril, enalaprilat, and lisinopril, Biochim. Biophys. Acta 1995, 3236, 31-38. 

Wang TT, Fellows PF, Leighton TJ et al (2004) Induction of opsonic antibodies to the gamma-D-glutamic acid capsule of Bacillus anthracis by immunization with a synthetic peptide-carrier protein conjugate. FEMS Immunol Med Microbiol 40 231-237... 

Add peptide hapten to the carrier solution to obtain a concentration of about 2mg/ml. Alternatively, determine the molar ratio of peptide to carrier. Ratios of 20 1 to 40 1 (peptide carrier) usually result in good immunogens. 

Add fresh glutaraldehyde to the peptide/carrier solution to obtain a 1 percent final concentration. Mix well. 

On the other hand, several probe substrates of carrier-mediated transport systems in the small intestine have been reported to be not absorbed by carrier-mediated mechanism in the rat colon in situ. Those include L-carnitine [23], methotrexate [18], cephradine [18], and 5-fluorouracil [18], as substrates of the L-carnitine carrier, folate carrier, peptide carrier, and pyrimidine carrier, respectively (Table 3.3). It is based on the nonsaturable nature of their transport. Particularly, the apparent membrane permeabilities of L-carnitine and methotrexate are negligibly low, suggesting that these compounds are practically unabsorbable from the colon. In the case of 5-fluorouracil, Na+-independence of transport was observed in situ [18] and also in everted sacs in vitro in the colon [21], while its carrier in the small intestine is known to be Na+-dependent. Furthermore, for ascorbate and nicotinate, as described in everted sacs in vitro [21], and L-dopa, as described in situ [24], carrier-mediated transport cannot be observed in the rat colon. 

There have been sustained efforts in recent years to use the carrier systems of the brush-border membrane of intestinal mucosa to increase absorption of orally administered drugs [29] [30]. One system of particular interest is the intestinal peptide carrier (hPEPTl) whose physiological function is the absorption of di- and tripeptides and whose xenobiotic substrates include /3-lactam antibiotics, renin inhibitors, and angiotensin-converting enzyme (ACE) inhibitors [31]. 

A series of peptide prodrugs ofh-a-methyldopa were prepared and shown to exhibit high affinity for the peptide carrier system [32], In an in situ intestinal perfusion model, the prodrugs Phe-L-a-methyldopa (6.10) and l-a-methyldopa-Phe (6.11) showed permeabilities that were 10- and 20-times higher, respectively, than that of L-a-methyldopa. The other derivatives examined (Gly- and Pro-L-a-methyldopa, L-a-methyldopa-Pro) also had better permeabilities. These and other results indicate that the peptide transport system has a relatively low substrate specificity and can indeed be targeted by peptide prodrugs to improve absorption [33],... 

P. W. Swaan, M. C. Stehouwer, J. J. Tukker, Molecular Mechanism for the Relative Binding Affinity to the Intestinal Peptide Carrier. Comparison of Three ACE-Inhibitors Enalapril, Enalaprilat and Lisinopril , Biochim. Biophys. Acta 1995, 1236, 31-38. 

M. Hu, P. Subramanian, H. I. Mosberg, G. L. Amidon, Use of the Peptide Carrier System to Improve the Intestinal Absorption of L-a-Methyldopa Carrier Kinetics, Intestinal Permeabilities, and in vitro Hydrolysis of Dipeptidyl Derivatives of L-a-Methyldopa , Pharm. Res. 1989, 6, 66-70. 

Coupling reagent Peptide Carrier protein Binding to ... 

Dissolve about 20 mg EDAC hydrochloride (Mr 191.7) in 200 pi Soln. A immediately before use and mix vigorously with the peptide-carrier solution. Shake at RT for 2 h. 

Add 500 (xl of the peptide solution to 200 xl of carrier protein. For greater reaction volumes, keep the ratio of peptide/carrier addition the same and proportionally scale up the amount of EDC added in the next step. If the peptide is... 

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