Synthesis and cytotoxic activities of some 2-arylnaphtho[2,3-d]oxazole-4,9-dione derivatives on androgen-dependent (LNCaP) and androgen-independent (PC3) human prostate cancer cell lines.

The synthesis of five 2-arylnaphtho[2,3-d]oxazole-4,9-dione derivatives was accomplished by refluxing 2-amino-3-bromo-1,4-naphthoquinone with appropriate benzoyl chloride analogs at elevated temperatures. In vitro anticancer evaluation of these compounds was performed on androgen-dependent, LNCaP, and androgen-independent, PC3, human prostate cancer cell lines. In general, these compounds displayed slightly stronger cytotoxicity on the androgen-dependent LNCaP than on the androgen-independent PC3 prostate cancer cell lines. The meta-substituted 2-(3-Chloro-phenyl)-naphtho[2,3-d]oxazole-4,9-dione (10) appear to display the best cytotoxicity on both cell lines with an IC(50) of 0.03 µM on LNCaP and 0.08 µM on PC3 after 5 days of exposure.

Paramagnetic intermediates of (E)-4-hydroxy-3-methylbut-2-enyl diphosphate synthase (GcpE/IspG) under steady-state and pre-steady-state conditions.

(E)-4-Hydroxy-3-methylbut-2-enyl diphosphate synthase (GcpE/IspG) converts 2-C-methyl-D-erythritol-2,4-cyclodiphosphate (MEcPP) into (E)-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) in the penultimate step of the methyl-erythritol phosphate (MEP) pathway for isoprene biosynthesis. MEcPP is a cyclic compound and the reaction involves the opening of the ring and removal of the C3 hydroxyl group consuming a total of two electrons. The enzyme contains a single [4Fe-4S] cluster in its active site. Several paramagnetic species are observed in steady-state and pre-steady-state kinetic studies. The first signal detected is from a transient species that displays a rhombic electron paramagnetic resonance (EPR) signal with g(xyz) = 2.000, 2.019, and 2.087 (FeS(A)). A second set of EPR signals (FeS(B)) accumulated during the reaction. Labeling studies with (57)Fe showed that all species observed are iron-sulfur-based. (31)P-ENDOR measurements on the FeS(A) species showed a weak (31)P coupling which is in line with binding of the substrate to the enzyme in close proximity of the active-site cluster. On the basis of the EPR/ENDOR measurements, we propose a direct binding of the substrate to the [4Fe-4S] cluster during the reaction, and therefore that the iron-sulfur cluster is directly involved in a reductive elimination of a hydroxyl group. The FeS(B) signal also showed (31)P coupling; in this case, however, it could be shown that the signal is due to the binding of the reaction product HMBPP to the active site cluster.

Possible direct involvement of the active-site [4Fe-4S] cluster of the GcpE enzyme from Thermus thermophilus in the conversion of MEcPP.

The GcpE enzyme converts 2-C-methyl-D-erythritol-2,4-cyclodiphosphate (MEcPP) into (E)-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) in the penultimate step of the DOXP pathway for isoprene biosynthesis. Purification of the enzyme under exclusion of air leads to a preparation that contains solely [4Fe-4S] clusters. Kinetic studies showed that in the presence of the artificial reductant dithionite and MEcPP a new transient iron-sulfur-based signal is detected in electron paramagnetic resonance (EPR) spectroscopy. Similarity of this EPR signal to that detected in ferredoxin:thioredoxin reductase indicates that during the reaction an intermediate is directly bound to the active-site cluster.