Inhibition of the growth of the human ovarian cancer cell line A2780 by organometallic ruthenium(II) complexes of the type [(eta(6)-arene)Ru(X)(Y)(Z)], where arene is benzene or substituted benzene, X, Y, and Z are halide, acetonitrile, or isonicotinamide, or X,Y is ethylenediamine (en) or N-ethylethylenediamine, has been investigated. The X-ray crystal structures of the complexes [(eta(6)-p-cymene)Ru(en)Cl]PF(6) (5), [(eta(6)-p-cymene)RuCl(2)(isonicotinamide)] (7), and [(eta(6)-biphenyl)Ru(en)Cl]PF(6) (9) are reported. They have "piano stool" geometries with eta(6) coordination of the arene ligand. Complexes with X,Y as a chelated en ligand and Z as a monofunctional leaving group had the highest activity. Complexes 5, 6 (the iodo analogue of 5), 9, and 10 (ethylethylenediamine analogue of 9) were as active as carboplatin. Hydrolysis of the reactive Ru-Cl bond in complex 5 was detected by HPLC but was suppressed by the addition of chloride ions. Complex 5 binds strongly and selectively to G bases on DNA oligonucleotides to form monofunctional adducts. No inhibition of topoisomerase I or II by complexes 5, 6, or 9 was detected. These chelated Ru(II) arene complexes have potential as novel metal-based anticancer agents with a mechanism of action different from that of the Ru(III) complex currently on clinical trial.

Reactions of the anticancer drug carboplatin ("Paraplatin") with a variety of sulfur-containing amino acids have been investigated by (1)H and (15)N NMR spectroscopy and by HPLC. Thiols react very slowly and sulfur-bridged species containing four-membered Pt(2)S(2) rings are the predominant products. In contrast, reactions with thioether ligands are much more rapid, and kinetics for the initial stages of the reaction with L-methionine have been determined (k = 2.7 x 10(-)(3) M(-)(1) s(-)(1)). Surprisingly, very stable ring-opened species are formed such as cis-[Pt(CBDCA-O)(NH(3))(2)(L-HMet-S)] which has a half-life for Met-S,N ring-closure of 28 h at 310 K. A study of the formation of the analogous product for N-acetyl-L-methionine and its subsequent ring closure is reported. Reactions such as these may play a role in the biological activity of carboplatin.

NMR investigations of the kinetics and thermodynamics of the competitive binding of L-methionine (Met), L-histidine (His), and 5'-monophosphates of guanosine (5'-GMP), adenosine (5'-AMP), thymidine (5'-TMP) and cytidine (5'-CMP) to [Pt(dien)CI]+ (dien = 1,5-diamino-3-azapentane) in aqueous solution show that 5'-GMP selectively displaces S-bound Met, a finding which has implications for DNA platination by anticancer drugs in vivo.

The Ru(II) organometallic antitumor complex [(eta(6)-biphenyl)RuCl(en)][PF(6)] (1) reacts slowly with the amino acid L-cysteine (L-CysH(2)) in aqueous solution at 310 K. Reactions were followed over periods of up to 48 h using HPLC, electronic absorption spectroscopy, LC-ESI-MS, and 1D or 2D (1)H and (15)N NMR spectroscopy. Reactions at a 1 mM/2 mM (Ru/L-CysH(2)) ratio were multiphasic in acidic solutions (pH 5.1) and appeared to involve aquation as the first step. Initially, 1:1 adducts involving substitution of Cl by S-bound or O-bound L-CysH(2), [(eta(6)-biphenyl)Ru(S-L-CysH)(en)](+) (4a) and [(eta(6)-biphenyl)Ru(O-L-CysH(2))(en)](2+) (4b) formed, followed by the cystine adduct [(eta(6)-biphenyl)Ru(O-Cys(2)H(2))(en)](2+) (3), and two dinuclear complexes from which half or all of the chelated ethylenediamine had been displaced, [(eta(6)-biphenyl)Ru(H(2)O)(microS,N-L-Cys)Ru(eta(6)-biphenyl)(en)](2+) (5) containing one bridging cysteine, and [(eta(6)-biphenyl)Ru(O,N-L-Cys-S)(S-L-Cys-N)Ru(eta(6)-biphenyl)(H(2)O)] (6) containing two bridging cysteines. The unusual cluster species [(biphenyl)Ru](8) (7a) was also detected by MS and was more prevalent in reactions at higher L-CysH(2) concentrations. Complex 5 was the dominant product at pH 2-5, but overall, only ca. 50% of 1 reacted with L-CysH(2) in these conditions. The reaction between 1 and L-CysH(2) was suppressed in 50 mM triethylammonium acetate solution at pH > 5 or in 100 mM NaCl. Only 27% of complex 1 reacted with L-methionine (L-MetH) at an initial pH of 5.7 after 48 h at 310 K and gave rise to only one adduct [(eta(6)-biphenyl)Ru(S-L-MetH)(en)](2+) (8).

Detailed studies of the kinetics of platination of the single‐stranded 14‐base DNA oligonucleotide d(ATACATGGTACATA) and the corresponding duplex by cis‐[Pt(NH3)2(H2O)2]2+ show that HPLC and NMR are complementary methods which provide similar results. The 5'‐G and 3'‐G monofunctional intermediates were trapped, separated and characterized by NMR (via 15NH3 labeling) and enzymatic digestion followed by mass spectrometry. The kinetic data are compared with those for the corresponding reactions of cis‐[PtCl2(NH3)2] (cisplatin) and its monohydrolysed analogue. For both single and double strands of the oligonucleotide, the aqua complex shows little selectivity for the 5'‐G or the 3'‐G in the initial platination step, whereas the chloro‐complex preferentially platinates the 3'‐G. The base on the 3' side of the GG sequence appears to play an important role in controlling this selectivity; replacement of T by C increases the selectivity of duplex platination by the diaqua complex by a factor of about 6, and the selectivity of chelation of the 3'‐G monofunctional adduct by a factor of about 3. In general the reactivity of the 5'‐G in a GG sequence appears to be enhanced in a duplex compared with a single‐strand. For both the aqua‐monoadduct and chloro‐monoadduct, cis‐[Pt(NH3)2(NG)(H2O or Cl)], the 5'‐G monoadduct is much longer lived (t½≈ 4 h at 288 K for aqua, 80 h at 298 K for chloro) than the 3'‐G monoadduct (t½≤ 45 min at 288 K for aqua, 6 h at 298 K for chloro). Inspection of molecular mechanics models of the end states of various monofunctional adducts provided insight into H‐bonding and destacking interactions in these adducts and the sequence selectivity observed in their formation. Such adducts may play an important role in the mechanism of action of platinum anticancer drugs.

L-Methionine (L-HMet) increased the rate of reaction of the anticancer drug cisplatin, cis- [PtCI,(NH,),], with guanosine 5'-monophosphate (5'-GMP) at pH 7. The course of the reaction has been elucidated by 'H and ['H, 15N] N M R spectroscopy. Novel intermediates detected and characterized include cis-[ Pt(Y-GMP-W) ( L-H Met-S) ( NH,),I2+ and [Pt( L-Met-S,N) (5'-GMP-W ) ( NH,)] + (charges on 5'-GMP ignored), the formation of which involves ammine release. Monodentate S-bound L-H Met can co-ordinate reversibly, whereas S,N-chelated L-Met is much less reactive. Thus methionine residues in peptides and proteins could play a role in the transfer of Pt onto DNA. Comparative reactions of [Pt(en)CI,] (en = 1.2-diaminoethane) have also been

Photosystem I reduction by the soluble metalloproteins cytochrome c(6) and plastocyanin, which are alternatively synthesized by some photosynthetic organisms depending on the relative availability of copper and iron, has been investigated in cyanobacteria, green algae and plants. The reaction mechanism is classified in three different types on the basis of the affinity of the membrane complex towards its electron donor protein. The role of electrostatic interactions in forming an intermediate transient complex, as well as the structural and functional similarities of cytochrome c(6) and plastocyanin are analysed from an evolutionary point of view. The proposal made is that the heme protein was first "discovered" by nature, when iron was much more abundant on the Earth's surface, and replaced by plastocyanin when copper became available because of the oxidizing conditions of the new atmosphere.

Oxidation of the soluble, truncated form of cytochrome f by wild-type and mutant species of plastocyanin has been analyzed by laser flash absorption spectroscopy in the cyanobacterium Nostoc (formerly, Anabaena) sp. PCC 7119. At low ionic strengths, the apparent electron transfer rate constant of cytochrome f oxidation by wild-type plastocyanin is 1.34 x 10(4) s(-)(1), a value much larger than those determined for the same proteins from other organisms. Upon site-directed mutagenesis of specific residues at the plastocyanin interaction area, the rate constant decreases in all cases yet to varying extents. The only exception is the D54K variant, which exhibits a higher reactivity toward cytochrome f. In most cases, the reaction rate constant decreases monotonically with an increase in ionic strength. The observed changes in the reaction mechanism and rate constants are in agreement with the location of the mutated residues at the interface area, as well as with the peculiar orientation of the two partners within the Nostoc plastocyanin-cytochrome f transient complex, whose NMR structure has been determined recently. Furthermore, the experimental data herein reported match well the kinetic behavior exhibited by the same set of plastocyanin mutants when acting as donors of electrons to photosystem I [Molina-Heredia, F. P., et al. (2001) J. Biol. Chem. 276, 601-605], thus indicating that the copper protein uses the same surface areas-one hydrophobic and the other electrostatic-to interact with both cytochrome f and photosystem I.