PT  - JOURNAL ARTICLE
AU  - Kesheng Zhao
AU  - Guoxiong Luo
AU  - Guo-Min Zhao
AU  - Peter W. Schiller
AU  - Hazel H. Szeto
TI  - Transcellular Transport of a Highly Polar 3+ Net Charge Opioid Tetrapeptide
AID  - 10.1124/jpet.102.040147
DP  - 2003 Jan 01
TA  - Journal of Pharmacology and Experimental Therapeutics
PG  - 425--432
VI  - 304
IP  - 1
4099  - http://jpet.aspetjournals.org/content/304/1/425.short
4100  - http://jpet.aspetjournals.org/content/304/1/425.full
SO  - J Pharmacol Exp Ther2003 Jan 01; 304
AB  - Oligopeptides are generally thought to have poor permeability across biological membranes. Recent studies, however, suggest significant distribution of [Dmt1]DALDA (Dmt-d-Arg-Phe-Lys-NH2; Dmt is 2′,6′-dimethyltyrosine), a 3+ net charge opioid peptide, to the brain and spinal cord after subcutaneous administration. Peptide transporters (PEPT1 and PEPT2) play a major role in the uptake of di- and tripeptides across cell membranes, but their ability to transport tetrapeptides is not clear. The purpose of this study was to determine whether [Dmt1]DALDA can translocate across Caco-2 cell monolayers and whether PEPT1 plays a role in the uptake process. Our results show that [3H][Dmt1]DALDA can readily translocate across Caco-2 cells, with a permeability coefficient estimated to be 1.24 × 10−5 cm/s. When incubated with Caco-2 cells, [3H][Dmt1]DALDA was detected in cell lysates by 5 min. The internalization of [Dmt1]DALDA was confirmed visually with a fluorescent [Dmt1]DALDA analog (H-Dmt-d-Arg-Phe-dnsDap-NH2; dnsDap is β-dansyl-l-α,β-diaminopropionic acid). The uptake of [3H][Dmt1]DALDA was concentration-dependent but temperature- and pH-independent. Treatment with diethylpyrocarbonate (DEPC) inhibited [14C]glycine-sarcosine uptake but increased [3H][Dmt1]DALDA uptake 34-fold. These findings suggest that PEPT1 is not involved in [Dmt1]DALDA internalization. [Dmt1]DALDA uptake was also observed in SH-SY5Y, human embryonic kidney 293, and CRFK cells, and was independent of whether the cells expressed opioid receptors. The efflux of [3H][Dmt1]DALDA from Caco-2 cells was temperature-dependent and was inhibited by DEPC, but was not affected by verapamil, an inhibitor of P-glycoprotein. These data show transcellular translocation of a highly polar 3+ charge tetrapeptide and suggest that [Dmt1]DALDA may not only distribute across the blood-brain barrier but also it may even have reasonable oral absorption. The American Society for Pharmacology and Experimental Therapeutics