Abstract
The feasibility of using adenovirus-mediated human oligopeptide transporter (hPEPT1) gene transfer to achieve peptide drug delivery to the brain across the blood-brain barrier was tested by examining the accumulation of model peptides in a rat brain endothelial cell line (RBEC1) and rat brain after transduction with a recombinant adenovirus encoding hPEPT1-enhanced yellow fluorescent protein fusion gene (AdhPEPT1-EYFP). In vitro uptake of [3H]GlySar was determined in RBEC1 transduced with AdhPEPT1-EYFP. In vivo, the accumulation of cefadroxil in rat brain was evaluated after transduction of AdhPEPT1-EYFP. At pH 6.0, the uptake of [3H]GlySar by RBEC1 transduced with AdhPEPT1-EYFP was increased 4-fold compared with that of nontransduced cells. At pH 7.4, uptake of [3H]GlySar in AdhPEPT1-EYFP transduced RBEC1 was 1.5 times higher than that of nontransduced cells. Unlabeled glycylsarcosine (10 mM) reduced the uptake of [3H]GlySar to a level comparable with that of nontransduced cells. At 30 min after intravenous administration of cefadroxil to rats transduced with AdhPEPT1-EYFP at 3.2 × 109 plaque-forming units/rat by an in situ brain perfusion method, the brain-to-plasma concentration ratio (Kp) of cefadroxil was increased about 2 times compared with that of nontransduced or AdGFP (control vector)-transduced rats, although this was not statistically significant. In contrast, Kp of [14C]inulin, a marker for extracellular fluid space, remained unchanged after adenoviral transduction. In conclusion, our results suggest that adenovirus-mediated heterologous expression of hPEPT1 in vivo could be a useful approach to deliver oligopeptides to the brain.
Footnotes
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This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science, and Technology and by grants from the Uehara Memorial Foundation and Takeda Science Foundation.
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DOI: 10.1124/jpet.102.046243
- Abbreviations:
- CNS
- central nervous system
- BBB
- blood-brain barrier
- hPEPT1
- human H+ peptide cotransporter
- RBEC
- rat brain endothelial cell
- CMV
- cytomegalovirus
- PFU
- plaque-forming unit
- MOI
- multiplicity of infection
- MES
- 2-(N-morpholino)ethanesulfonic acid
- RT-PCR
- reverse transcription-polymerase chain reaction
- PBS
- phosphate-buffered saline
- EYFP
- enhanced yellow fluorescent protein
- HPLC
- high-performance liquid chromatography
- Kp
- tissue-to-plasma concentration ratio
- GFP
- green fluorescent protein
- Received October 26, 2002.
- Accepted December 31, 2002.
- The American Society for Pharmacology and Experimental Therapeutics
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