PGLU-L-DOPA-PRO - A TRIPEPTIDE PRODRUG TARGETING THE INTESTINAL PEPTIDE TRANSPORTER FOR ABSORPTION AND TISSUE ENZYMES FOR CONVERSION

Purpose. The purpose of this study is to investigate the characteristi cs of pGlu-L-Dopa-Pro as a prodrug of L-Dopa. Methods. pGlu-L-Dopa-Pro and L-Dopa-Pro were synthesized using the standard procedures of pept ide synthesis. The conversion of pGlu-L-Dopa-Pro to L-Dopa was studied using pyroglutamyl aminopeptidase I and prolidase. With rats as the a nimal model, the stability of pGlu-L-Dopa-Pro in intestinal homogenate s was determined, then the transport characteristics of pGlu-L-Dopa-Pr o were studied using in-situ perfusion and Ussing chambers. Results. p Glu-L-Dopa-Pro, relatively stable in intestinal homogenates and intest inal fluid, had a dimensionless permeability of 1.8 at 0.04 mM. Its in testinal permeability was significantly inhibited by 20 mM captopril, by a mixture of dipeptides, 80 mM Gly-Gly and 5 mM Gly-Pro, and by 2 m M cephradine. Further, in Ussing chambers, its mucosal to serosal perm eability decreased dramatically with concentration. Conversion studies showed that pGlu-L-Dopa-Pro was degraded by pyroglutamyl aminopeptida se I, an enzyme releasing the N-terminal pyroglutamic acid, with Vmax and Km of 0.6 mu mole/min/g protein and 21 mM, respectively, and that L-Dopa-Pro was degraded by prolidase with Vmax and Km of 44 mu mole/mi n/g protein and 0.48 mM, respectively. Conclusions. This tripeptide, a potential prodrug of L-Dopa, is absorbed by the intestinal peptide tr ansporter, is relatively stable in the gut wall, and is converted to L -Dopa by peptidases with the cleavage by pyroglutamyl aminopeptidase I to L-Dopa-Pro as the rate limiting step.