Abstract
Inhibition of the sodium-proton exchanger (NHE) plays an important role in reducing tissue damage during ischemic reperfusion injury; however, pharmacological inhibitors of NHE have restricted access to acutely ischemic tissues because of severely compromised tissue perfusion. We describe the syntheses, characterization, and NHE inhibitory activities of a novel class of amiloride derivatives where peptides are conjugated to the amiloride C(5) amino group. These new peptide-C(5)-amiloride conjugates are inactive; however, peptide residues were chosen such that selective cleavage by neutral endopeptidase 24.11 (enkephalinase) liberates an amino acid-C(5)-amiloride conjugate that inhibits NHE in a glial cell line. These results confirm the feasibility of using peptide-amiloride conjugates as NHE inhibitor prodrugs. We envision the design of analogous peptide-amiloride prodrugs that can be administered prior to ischemic events and subsequently activated by endopeptidases selectively expressed by ischemic tissues.
Footnotes
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↵1 Relative molar absorptivity constants determined at 380-nm excitation and 510-nm emission for amiloride, EIPA, and Gly-C(5)-Am (4a) are 1, 2, and 13, respectively.
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↵2 cLogP values for amiloride (–2.22) versus 4a (–6.23, calculated) indicate a substantial difference in hydrophilicity.
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This work was supported by funding from National Institutes of Health Grants NS40489 and P50 CA097257-03 and by the University of California Cancer Research Coordinating Committee. F.A.G. and M.H.N. contributed equally to this work.
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doi:10.1124/jpet.104.076984.
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ABBREVIATIONS: NHE, sodium proton exchanger; NCX, sodium-calcium exchanger; I/R, ischemia-reperfusion; EIPA, ethylisopropylamiloride; SAR, structure-activity relationship; Am, amiloride; LC, liquid chromatography; MS, mass spectrometry; BCECF, 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein.
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↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
- Received August 31, 2004.
- Accepted October 20, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
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