The mechanism of Daunorubicin-induced inhibition of prolidase activity in human skin fibroblasts and its implication to impaired collagen biosynthesis

One of the recognized side effects accompanying antineoplastic anthracyclin es administration is poor wound healing, resulting from impairement of coll agen biosynthesis. However, the precise mechanism of anthracyclines-induced inhibition of collagen synthesis has not been established. We have suggest ed that prolidase, an enzyme involved in collagen metabolism may be one of the targets for anthracyclines-induced inhibition of synthesis of this prot ein. Prolidase [E.C. 3.4.13.9] cleaves imidodipeptides containing C-termina l proline, providing large amount of proline for collagen synthesis. We hav e found that daunorubicin (DNR) induced coordinately inhibition of prolidas e activity (IC50 = 0.3 mu M) and collagen biosynthesis (IC50 = 1 mu M) in c ultured human skin fibroblasts. The decrease in prolidase activity due to t he treatment of confluent cells with DNR was not accompanied by any differe nces in the amount of the enzyme protein recovered from these cells as show n by western immunoblot analysis. Since prolidase is metaloprotease, requir ing manganese for catalytic activity and anthracyclines are known as a chel ators of divalent cations we considered that the chelating ability of anthr acyclines may be an underlying mechanism for daunorubicin-induced inhibitio n of prolidase activity. In order to determine the ability of DNR to form c omplex with manganese (II), potentiometric method was employed based on the measurement of protonation constant by pH-metric titrated assay. We have f ound that DNR forms stable complex with manganese (II) and the composition of the complex of DNR with Mn (II) was calculated as 3 : 1. The constant st ability value for the investigated complex was calculated as beta(av) = (1. 74 +/- 0.01) 10(23). The strong ability of DNR to chelate manganese may explain the potential me chanism for inhibition of prolidase activity, subsequently collagen biosynt hesis and poor wound healing in patients administered DNR.