Abstract
The permeation characteristics of a model opioid peptide, H-Tyr-d-Ala-Gly-Phe-d-Leu-OH (DADLE), and its cyclic prodrugs [acyloxyalkoxy-based cyclic prodrug of DADLE (AOA-DADLE), coumarinic acid-based cyclic prodrug of DADLE (CA-DALE), and oxymethyl-modified coumarinic acid-based cyclic prodrug of DADLE (OMCA-DADLE)] across the blood-brain barrier (BBB) were determined using an in situ perfused rat brain model. The rat brains were perfused with Krebs-bicarbonate buffer containing test compounds in the absence or presence of a specific P-glycoprotein inhibitor (GF-120918). Brain samples were collected after perfusion and processed by a capillary depletion method. After liquid phase extraction with acetonitrile, samples were analyzed using high-performance liquid chromatography with tandem mass spectrometric detection. Linear uptake kinetics of DADLE and its cyclic prodrugs was observed within the range of 60 to 240 s of perfusion. The apparent permeability coefficient (Papp) of DADLE across the BBB was very low (<10−7 cm/s), probably due to its unfavorable physicochemical properties (e.g., charge, hydrophilicity, and high hydrogen-bonding potential). All three cyclic prodrugs, however, also exhibited low membrane permeation (Papp <10−7 cm/s) in spite of their more favorable physicochemical properties (e.g., no charge, high hydrophobicity, and low hydrogen-bonding potential). Inclusion of GF-120918 (10 μM) in the perfusates fully inhibited the P-gp activity in the BBB and dramatically increased the Pappvalues of AOA-DADLE, CA-DADLE, and OMCA-DADLE by approximately 50-, 460-, and 170-fold, respectively. In contrast, GF-120918 had no effect on the Papp value of DADLE. In addition, the observed bioconversions of the prodrugs to DADLE in the rat brains after 240-s perfusion were very low (5.1% from AOA-DADLE, 0.6% from CA-DADLE, and 0.2% from OMCA-DADLE), which was consistent with the in vitro bioconversion rates determined previously in rat brain homogenates.
Footnotes
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This research was supported by Grant DA09315 from the U.S. Public Health Service.
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DOI: 10.1124/jpet.102.037143
- Abbreviations:
- BBB
- blood-brain barrier
- DADLE
- [d-Ala2,d-Leu5]-enkephalin
- AOA-DADLE
- acyloxyalkoxy-based cyclic prodrug of [d-Ala2,d-Leu5]-enkephalin
- CA-DADLE
- coumarinic acid-based cyclic prodrug of [d-Ala2,d-Leu5]-enkephalin
- OMCA-DADLE
- oxymethyl-modified coumarinic acid-based cyclic prodrug of [d-Ala2,d-Leu5]-enkephalin
- P-gp
- P-glycoprotein
- LCCA
- left common carotid artery
- LC/MS/MS
- high-performance liquid chromatography with tandem mass spectrometric detection
- HPLC
- high-performance liquid chromatography
- Received April 9, 2002.
- Accepted August 2, 2002.
- The American Society for Pharmacology and Experimental Therapeutics
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