Inactivation of dihydropteridine reductase (human brain) by platinum (II) complexes

Abstract

Potassium tetrachloroplatinate (K2PtC14) inactivates dihydropteridine reductase from human brain in a timedependent and irreversible manner. The inactivation has been followed by measuring enzyme activity and fluorescence changes. The enzyme is completely protected from inactivation by NADH, the pterin cofactor [quinonoid 6-methyl-7,8-dihydro(6H)pterin] and dithiothreitol. Evidence is presented that K2PtC14 reacts at the active site and that (a) thiol group(s) is involved in, or is masked by, this reaction.K2PtC14 is a stronger inhibitor of human brain dihydropteridine reductase than cis-and transdiaminodichloroplatinum, cis-dichloro[ethylenediamine]platinum and K,Fe(CN),, whereas H2PtC1, is considerably weaker and (Ph3P)3RhCl is inactive.Studies of the anti-tumour activities of platinum coordination compounds have lead to the use of cis-platin in the chemotherapy of cancer [l, 21. Cis-platin owes its anticancer activity to its reactivity with DNA [3]. Platinum(I1) compounds have also been shown to form complexes with amino acids and proteins, and were very useful when they crystallised with proteins because they facilitated X-ray crystallographic studies of those proteins [4]. Certain enzymes are inactivated by platinum(I1) complexes [5], and in a study of seven enzymes it was found that those enzymes which possessed essential thiol groups at the active site were inhibited significantly by chloroplatinate(I1) complexes. Enzymes that do not have essential thiol groups at the active site were not significantly affected under similar conditions [6]. This may not be surprising in view of the fact that platinum(I1) compounds react strongly with sulphur atoms to form square planar Pt-S complexes [7]. Because of the possibility that cisplatin may inhibit important enzymes during its use in the chemotherapy of cancers, Dhondt and Bellahsene [8] examined its effects on the activity of dihydropteridine reductase from rat tissues. They observed that the enzyme activity was 10% and 28% irreversibly inhibited at concentrations of 25 mg/l and 50 mg/l of cis-platin respectively. When the enzyme was pre-incubated for 5 min with cis-platin (100 mg/l) 60% inhibition was obtained and the enzyme was protected to a large extent (17% inhibition) when it was preincubated with potassium chloride (50 mM) before additionCorrespondence to

Mechanistic analysis of iron accumulation by endothelial cells of the BBB

“…We used Pt II , an inhibitor of pyridine nucleotide-dependent reductases (Armarego and Ohnishi 1987) including cell-surface ferrireductases (Eide et al 1992), to assess the relative contributions of ferrireductases in mediating the reduction of TBI and NTBI. Dose-response experiments indicated that 100 μM Pt II exihibited minimal cytotoxicity in hBMVEC monolayers (Supplementary Table 1) while displaying maximal effectiveness in inhibition of ferrireduction by hBMVEC (data not shown).…”

“…The Kaplan lab demonstrated that Fre1, a cell-surface ferrireductase found in Saccharomyces cerevisiae , is inhibited by Pt II (Eide et al 1992): Pt II had previously been demonstrated to inhibit pyridine-nucleotide reductases likely by reaction with protein cysteine thiol(s) (Armarego and Ohnishi 1987). We extended these observations regarding Pt II to our mammalian cell culture system and demonstrated that Pt II effectively inhibited cell-surface reduction of TBI (Figure 3a).…”