METABOLISM OF [DES-GLY(10),D-TRP(6)]LHRH ETHYLAMIDE IN THE RABBIT CONJUNCTIVA

It was the objective of this study to determine whether [des-Gly(10), D-Trp(6)]LHRH ethylamide, an LHRH agonist known as deslorelin, is degr aded by the rabbit conjunctiva. Intact conjunctiva was incubated with deslorelin either alone or in the presence of 0.1 mM ouabain, 0.1% 2,4 -dinitrophenol, 0.1 mM phosphoramidon, 0.1 mM N-tosyl-L-phenylalanine chloromethylketone (TPCK), 2% EDTA, 1% ZnCl2, 0.1% dithiothreitol (DTT ), or 0.1% N-ethylmaleimide (NEM) at 37 degrees C. Furthermore, deslor elin alone was incubated with conjunctiva at 4 degrees C. All incubati on solutions were made isotonic, and the pH was adjusted to 5.0. A rev ersed-phase HPLC was used to analyze supernatants collected at the end of 90 min. Deslorelin metabolism was inhibited at low temperature, as suggested by the disappearance of metabolite peaks at low temperature . Both ouabain and dinitrophenol failed to alter the intact drug remai ning in the supernatant, indicating that energy-dependent cellular upt ake of deslorelin is unlikely in the conjunctiva. Phosphoramidon- and TPCK- sensitive endopeptidases did not contribute to the observed meta bolism, as suggested by the lack of effect of phosphoramidon and TPCK on deslorelin levels. DTT and NEM also failed to affect deslorelin lev els. On the other hand, EDTA and ZnCl2 significantly elevated the inta ct deslorelin levels by 61 and 53%, respectively, and almost completel y abolished the metabolite peaks, indicating a possible role for eithe r metal-dependent peptidases or metallo-peptidases in the conjunctival metabolism of deslorelin.