Cisplatin mediates selective downstream hydrolytic cleavage of Met-(Gly)(n)-His segments (n=1,2) in methionine- and histidine-containing peptides: the role of ammine loss trans to the initial Pt-S(Met) anchor in facilitating amide hydrolysis

The pH- and time-dependent reactions of the antitumor drug cisplatin, cis-[PtCl(2)(NH(3))(2)], with the methionine- and histidine-containing pentapeptides Ac-Met-Gly-His-Gly-Gly-OH, Ac-Met-Gly-Gly-His-Gly-OH and Ac-Gly-Met-Gly-His-Gly-OH (Gly=glycyl, Met=L-methionyl, His=L-histidyl) at 313K have been investigated by high performance liquid chromatography, mass spectrometry and nuclear magnetic resonance. Cisplatin mediates a rapid "downstream" hydrolytic cleavage of the Met-Gly amide bond in weakly acid solution (pH < or =5) for all three peptides, leading to release of H-Gly-His-Gly-Gly-OH, H-Gly-Gly-His-Gly-OH and H-Gly-His-Gly-OH, respectively, and formation of kappa(2)S,N(M) chelate complexes of the methionine-containing residuals Ac-Met-OH or Ac-Gly-Met-OH. An alternative reaction pathway affords tridentate kappa(3)S,N(M),N(imidazole) macrochelates of the original pentapeptide following ammine loss. The downstream cleavage pathway is competitive with the likewise cisplatin-mediated upstream cleavage of the Ac-Gly linkage in the pentapeptide Ac-Gly-Met-Gly-His-Gly-OH. This leads to formation of both the kappa(3)S,N(M),N(G1) complex of H-Gly-Met-Gly-His-Gly-OH due to upstream cleavage and the analogous tridentate complex for H-Gly-Met-OH due to initial downstream loss of H-Gly-His-Gly-OH followed by upstream loss of acetic acid. As downstream cleavage is not observed for Ac-(Gly)(2)-Met-(Gly)(2)-OH under similar conditions, it may be concluded that rapid histidine imidazole substitution of the ammine ligand in trans-position to an anchoring methionine S atom must assist hydrolytic cleavage of the Met-Gly amide bond.     

Interaction of cisplatin with methionine- and histidine-containing peptides: competition between backbone binding,macrochelation and peptide cleavage

The pH- and time-dependent reaction of cis-[PtCl2(NH3)2] with the methionine- and histidine-containing peptides H-Gly-Met-OH, H-Gly-Gly-Met-OH, Ac-His-Gly-Met-OH, and Ac-His-(Ala)3-Met-OH at 313 K has been investigated by ion-pairing reverse phase HPLC and NMR spectroscopy. For equimolar solutions (c=0.8 mM, pH approximately equals 3 or 8.8), initial formation of the kinetically favored S-bound complex is followed by relatively rapid metallation of the neighboring methionine amide nitrogen NM to afford a kappa2NM,S six-membered chelate. The strong trans effect of the methionine S then favors facile NH3 substitution, leading to generation of tridentate complexes such as [Pt(H-Gly-MetH(-1)-OH)-kappa3NG,NM,S)(NH3)]+ or [Pt(H-Ac-His-GlyH(-1)-MetH(-1)-OH-kappa3NG,NM,S)(NH3)]. In the case of H-Gly-Gly-Met-OH, this reaction is accompanied by loss of a second NH3 ligand in alkaline solution to generate the tetradentate kappa4NG1,NG2,NM,S species. In contrast, cleavage of the backbone C(O)-N bond to the second metallated amide nitrogen after t>100 h is common to the tridentate complexes of the tri- and pentapeptides at pH<5. Although an imidazole-coordinated kappa2N3H,S macrochelate is formed throughout the whole range 2.5 < or = pH < or = 10 for Ac-His-Gly-Met-OH, it slowly decays (t=10-1000 h) to the thermodynamically more stable tridentate kappa3NG,NM,S complex. All major final products were separated and fully characterized by NMR and MS.     

Reactions of cisplatin hydrolytes with methionine, cysteine, and plasma ultrafiltrate studied by a combination of HPLC and NMR techniques

The reactions of cis-[PtCl(NH3)2(H2O)]+ with L-methionine have been studied by 1D 195Pt and 15N NMR, and by 2D[1H, 15N] NMR. When the platinum complex is in excess, the initial product, cis-[PtCl(NH3)2(Hmet-S)]+ undergoes slow ring closure to [Pt(NH3)2(Hmet-N,S)]2+. Slow ammine loss then occurs to give the isomer of [PtCl(NH3)(Hmet-N,S)]+ with chloride trans to sulfur. When methionine is in excess, a reaction sequence is proposed in which trans-[PtCl(NH3)(Hmet-S)2]+ isomerises to the cis-isomer, with subsequent ring closure reactions leading to cis-[Pt(Hmet-N,S)2]2+. Near pH 7, methionine is unreactive toward cis-[PtCl(OH)(NH3)2]. By contrast, L-cysteine reacts readily with cis-[PtCl(OH)(NH3)2] at pH 7, but there were many reaction products, including bridged species. Cis-[PtCl(OH)(NH3)2] reacts with reduced thiols in ultrafiltered plasma but these are oxidized if the plasma is not fresh or appropriately stored. With very low concentrations of the platinum complexes (35.5 microM), HPLC experiments (UV detection at 305 nm) indicate that the thiolate (probably cysteine) reactions become simpler as bridging becomes less important.     

Comparison of the electronic properties,and thermodynamic and kinetic parameters of the aquation of selected platinum(II) derivatives with their anticancer IC<Subscript>50</Subscript> indexes

Three potential anticancer agents {trans-[PtCl(2)(NH(3))(thiazole)], cis-[PtCl(2)(NH(3))(piperidine)], and PtCl(2)(NH(3))(cyclohexylamine) (JM118)} were explored and compared with cisplatin and the inactive [PtCl(dien)](+) complex. Basic electronic properties, bonding and stabilization energies were determined, and thermodynamic and kinetic parameters for the aquation reaction were estimated at the B3LYP/6-311++G(2df,2pd) level of theory. Since the aquation process represents activation of these agents, the obtained rate constants were compared with the experimental IC(50) values for several tumor cells. Despite the fact that the processes in which these drugs are involved and the way in which they affect cells are very complex, some correlations can be deduced.     

The fate of platinum(II) and platinum(IV) anti-cancer agents in cancer cells and tumours

SRIXE mapping has been used to gain insight into the fate of platinum(II) and platinum(IV) complexes in cells and tumours treated with anticancer active complexes to facilitate the development of improved drugs. SRIXE maps were collected of thin sections of human ovarian (A2780) cancer cells treated with bromine containing platinum complexes, cis-[PtCl(2)(3-Brpyr)(NH(3))] (3-Brpyr=3-bromopyridine) and cis,trans,cis-[PtCl(2)(OAcBr)(2)(NH(3))(2)] (OAcBr=bromoacetate), or a platinum complex with an intercalator attached cis-[PtCl(2)(2-[(3-aminopropyl)amino]-9,10-anthracenedione)(NH(3))]. After 24h the complexes appear to be localised in the cell nucleus with a lower concentration in the surrounding cytoplasm. In cells treated with cis-[PtCl(2)(3-Brpyr)(NH(3))] the concentration of bromine was substantially higher than in control cells and the bromine was co-localised with the platinum consistent with the 3-bromopyridine ligand remaining bound to the platinum. The cells treated with cis,trans,cis-[PtCl(2)(OAcBr)(2)(NH(3))(2)] also showed an increased level of bromine, but to a much lesser extent than for those treated with cis-[PtCl(2)(3-Brpyr)(NH(3))] suggestive of substantial reduction of the platinum(IV) complex. Maps were also collected from thin sections of a 4T1.2 neo 1 mammary tumour xenograft removed from a mouse 3h after treatment with cis,trans,cis-[PtCl(2)(OH)(2)(NH(3))(2)] and revealed selective uptake of platinum by one cell.     

The influence of methionine-containing peptides on the reaction of carboplatin with 5'-guanosine monophosphate: a comparison with cisplatin

The pH- and time-dependent reaction of the anticancer drug carboplatin, [Pt(cbdca-kappa(2)O,O')(NH(3))(2)] (cbdca=cyclobutane-1,1-dicarboxylate), with the tripeptides H-glyglymet-OH (glycylglycyl-L-methionine) and Ac-glyglymet-OH at 313 K was investigated by high-performance liquid chromatography, NMR and mass spectrometry. The relative stability of the initial ring-opened kappaS complex [Pt(cbdca-kappaO)(Ac-glyglymet-OH-kappaS)(NH(3))(2)] leads to increased formation of the kinetically favoured kappaS:kappaS' bis-adduct [Pt(Ac-glyglymet-OH-kappaS)(2)(NH(3))(2)](2+) in comparison with cisplatin. As a result a second 1:2 reaction pathway kappaS-->kappaS:kappaS'-->kappa(2)N(M), S:kappaS'-->kappa(3)N(G2),N(M), S:kappaS', where N(M) and N(G2) represent, respectively, metallated methionine and glycine nitrogen atoms, competes with the 1:1 route kappaS-->kappa(2)N(M), S-->kappa(3)N(G2),N(M), S also observed for cisplatin. Cleavage of N-acetylglycine at the backbone C(O)-N bond to the second gly residue (G2) is observed after 100 h for the respective tridentate complexes [Pt(Ac-glyglyH(-1)metH(-1)-OH-kappa(3)N(G2),N(M), S) (Ac-glyglymet-OH-kappaS)] and [Pt(Ac-glyglyH(-1)metH(-1)-OH-kappa(3)N(G2),N(M), S)(NH(3))] at pH <5.2. The longevity of the initial kappaS complex leads to about an eight-fold increase in the rate of formation of the kappaN7:kappaN7' bis-adduct [Pt(5'-GMP-kappaN7)(2)(NH(3))(2)](2-) for the reaction of carboplatin with 5'-GMP(2-) at pH 7 in the presence of Ac-glyglymet-OH. A mixed-ligand kappaS:kappaN7 species [Pt(5'-GMP-kappaN7)(Ac-glyglymet-OH-kappaS)(NH(3))(2)] provides the major precursor for this 1:2 nucleotide complex and kappaN7 coordination of 5'-GMP(2-) is also observed in the kappa(2)N(M),S:kappaN7 complex [Pt(5'-GMP-kappaN7)(Ac-glyglymetH(-1)-OH-kappa(2)N(M),S)(NH(3))(2)](-) formed by substitution of the ammine ligand trans to the methionine sulphur. As the intermediate kappaS:kappaN7 species is formed rapidly within the first 10 h of reaction, these results suggest that the transfer reaction pathway kappaS-->kappaS:kappaN7-->kappaN7:kappaN7' involving kappaS platinated peptides could play an important role in accelerating the rate of DNA binding for carboplatin.     

The mechanism of action of platinum(IV) complexes in ovarian cancer cell lines

The reduction potentials, lipophilicities, cellular uptake and cytotoxicity have been examined for two series of platinum(IV) complexes that yield common platinum(II) complexes on reduction: cis-[PtCl(4)(NH(3))(2)], cis,trans,cis-[PtCl(2)(OAc)(2)(NH(3))(2)], cis,trans,cis-[PtCl(2)(OH)(2)(NH(3))(2)], [PtCl(4)(en)], cis,trans-[PtCl(2)(OAc)(2)(en)] and cis,trans-[PtCl(2)(OH)(2)(en)] (en=ethane-1,2-diamine, OAc=acetate). As previously reported, the reduction occurs most readily when the axial ligand is chloride and least readily when it is hydroxide. The en series of complexes are marginally more lipophilic than their ammine analogues. The presence of axial chloride or acetate ligands results in a slighter higher lipophilicity compared with the platinum(II) analogue whereas hydroxide ligands lead to a substantially lower lipophilicity. The cellular uptake is similar for the platinum(II) species and their analogous tetrachloro complexes, but is substantially lower for the acetato and hydroxo complexes, resulting in a correlation with the reduction potential. The activities are also correlated with the reduction potentials with the tetrachloro complexes being the most active of the platinum(IV) series and the hydroxo being the least active. These results are interpreted in terms of reduction, followed by aquation reducing the amount of efflux from the cells resulting in an increase in net uptake.     

Heavy metal-nucleotide interactions. 15. Reactions of calf thymus DNA with the electrophiles methylmercury(II) nitrate, cis-dichlorodiammineplatinum(II), and trans-dichlorodiammineplatinum(II) studied using Raman difference spectroscopy. Evidence for the formation of c-DNA upon metalation

This study details the reactions of the electrophiles CH3Hg(NO3), cis-[PtCl2(NH3)2] (cis-DDP) and trans-[PtCl2(NH3)2] (trans-DDP) with calf thymus DNA using Raman and Raman difference spectroscopy. The order of CH3Hg(II) binding to calf thymus DNA is G > T > C > A. The electrophilic attack of cis- and trans-DDP on calf thymus DNA produces different orders of binding: cis-DDP-G>C approximately AT, trans-DDP-G approximately C approximately AT. The reaction of CH3Hg(II) with DNA results in a decrease in the percentage of B-form DNA. whereas the reactions of cis- and trans-DDP with DNA decrease the percentage of B-DNA and cause the formation of C-DNA structure.     

Pyrazolyl derivatives as bifunctional chelators for labeling tumor-seeking peptides with the fac-[M(CO)3]+ moiety (M = 99mTc, Re): synthesis, characterization, and biological behavior

Radiolabeling of biologically active molecules with the [(99m)Tc(CO)(3)](+) unit has been of primary interest in recent years. With this in mind, we herein report symmetric (L(1)) and asymmetric (L(2)-L(5)) pyrazolyl-containing chelators that have been evaluated in radiochemical reactions with the synthon [(99m)Tc(H(2)O)(3)(CO)(3)](+) (1a). These reactions yielded the radioactive building blocks [(99m)Tc(CO)(3)(k(3)-L)](+) (L = L(1)-L(5), 2a-6a), which were identified by RP-HPLC. The corresponding Re surrogates (2-6) allowed for macroscopic identification of the radiochemical conjugates. Complexes 2a-6a, with log P(o/w) values ranging from -2.35 to 0.87, were obtained in yields of > or =90% using ligand concentrations in the 10(-5-)10(-4) M range. Challenge studies with cysteine and histidine revealed high stability for all of these radioactive complexes, and biodistribution studies in mice indicated a fast rate of blood clearance and high rate of total radioactivity excretion, occurring primarily through the renal-urinary pathway. Based on the framework of the asymmetric chelators, the novel bifunctional ligands 3,5-Me(2)-pz(CH(2))(2)N((CH(2))(3)COOH)(CH(2))(2)NH(2) (L(6)) and pz(CH(2))(2)N((CH(2))(3)COOH)(CH(2))(2)NH(2) (L(7)) have been synthesized and their coordination chemistry toward (NEt(4))(2)[ReBr(3)(CO)(3)] (1) has been explored. The resulting complexes, fac-[Re(CO)(3)(k(3)-L)]Br (L(6)(7), L(7)(8)), contain tridentate ancillary ligands that are coordinated to the metal center through the pyrazolyl and amine nitrogen atoms, as observed for the other related building blocks. L(6) and L(7) were coupled to a glycylglycine ethyl ester dipeptide, and the resulting functionalized ligands were used to prepare the model complexes fac-[Re(CO)(3)(kappa(3)-3,5-Me(2)-pz(CH(2))(2)N(glygly)(CH(2))(2)NH(2))](+) (9/9a) and fac-[Re(CO)(3)(kappa(3)-pz(CH(2))(2)N(CH(2))(3)(glygly)(CH(2))(2)NH(2))](+) (10/10a) (M = Re, (99m)Tc). These small conjugates have been fully characterized and are reported herein. On the basis of the in vitro/in vivo behavior of the model complexes (2a-6a, 9a, 10a), we chose to evaluate the in vitro/in vivo biological behavior of a new tumor-seeking Bombesin pyrazolyl conjugate, [(L(6))-G-G-G-Q-W-A-V-G-H-L-M-NH(2)], that has been labeled with the [(99m)Tc(CO)(3)](+) metal fragment. Stability, in vitro cell binding assays, and pharmacokinetics studies in normal mice are reported herein.     

Consequences of nucleic acid conformation on the binding of a trinuclear platinum drug.

BBR3464, a charged trinuclear platinum compound, is the first representative of a new class of anticancer drugs to enter phase I clinical trials. The structure of BBR3464 is characterized by two [trans-PtCl(NH(3))(2)] units linked by a tetraamine [trans-Pt(NH(3))(2)?H(2)N(CH(2))(6)NH(2)?(2)] unit. The +4 charge of BBR3464 and the separation of the platinating units indicate that the mode of DNA binding will be distinctly different from those of classical mononuclear drugs such as cisplatin, cis-[PtCl(2)(NH(3))(2)]. The reaction of BBR3464 with three different nucleic acid conformations was assessed by gel electrophoresis. Comparison of single-stranded DNA, RNA, and double-stranded DNA indicated that the reaction of BBR3464 with single-stranded DNA and RNA was faster than that with duplex DNA, and produced more drug-DNA and drug-RNA adducts. Electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry was used to further characterize the binding modes of BBR3464 with the DNA substrates. BBR3464 binding to different nucleic acid conformations raises the possibility that the adducts of single-stranded DNA and RNA may play a role in the different antitumor efficacies of this novel drug as compared with cisplatin.     

Reaction of cis- and trans-[PtCl2(NH3)2] with reduced glutathione inside human red blood cells, studied by 1H and 15N-[1H] DEPT NMR

Reactions of cis- and trans-[PtCl2(NH3)2] with glutathione (GSH) inside intact red blood cells have been studied by 1H spin-echo nuclear magnetic resonance (NMR). Upon addition of trans-[PtCl2(NH3)2] to a suspension of red cells, there was a gradual decrease in the intensity of the resonances for free GSH, and new peaks were observed that were assignable to coordinated GSH protons in trans-[Pt(SG)Cl(NH3)2], trans-[Pt(SG)2(NH3)2], and possibly the S-bridged complex trans-[[NH3)2PtCl)2SG]+. Formation of trans-[Pt(SG)2(NH3)2] inside the cell was confirmed from the 1H NMR spectrum of hemolyzed cells, which were ultrafiltered to remove large protein molecules; the ABM multiplet of the coordinated GSH cys-beta CH2 protons was resolved using selective-decoupling experiments. Seventy percent of the total intracellular GSH was retained by the ultrafiltration membrane, suggesting that the mixed complex trans-[Pt(SG)(S-hemoglobin)(NH3)2] also is a major metabolite of trans-[PtCl2(NH3)2] inside red cells. The reaction of cis-[PtCl2(NH3)2] with intracellular GSH was slower; only 35% of the GSH had been complexed after a 4-hr incubation compared to 70% for the trans isomer. There was a gradual decrease in the intensity of the GSH 1H spin-echo NMR resonances, but no new peaks were resolved. This was interpreted as formation of high-molecular weight Pt:GSH and mixed GS-Pt-S(hemoglobin) polymers. By using a 15N-[1H] DEPT pulse sequence, we were able to study the reaction of cis-[PtCl2(15NH3)2] with red cells at concentrations as low as 1 mM. 15NH3 ligands were released, and no resonances assignable to Pt-15NH3 species were observed after a 12-hr incubation.     

Kinetics and mechanism for platination of thione-containing nucleotides and oligonucleotides: evaluation of the salt dependence

Reactions of cis-[PtCl(NH(3))(CyNH(2))(OH(2))](+) (Cy=cyclohexyl) with thione-containing single-stranded oligonucleotides d(T(8)XT(8)) and d(XT(16)) (X=(s6)I or (s4)U) and the mononucleotides 4-thiouridine ((s4)UMP) and 6-mercaptoinosine ((s6)IMP) have been studied in aqueous solution at pH 4.1. The reaction kinetics was followed using HPLC methodology as a function of ionic strength in the interval 5.0 mM相似文献   

Steric control of stereoselective interactions between the platinum(II) complex [PtCl2(1,4-diazacycloheptane)] and DNA: comparison with cis-[PtCl2(NH3)2] and [PtCl2(ethane-1,2-diamine)] using DNA binding and molecular modeling studies

The rate and extent of binding of [PtCl2(hpip)] (hpip=homopiperazine-1,4-diazacycloheptane) and cis-[PtCl2(NH3)2] to calf thymus DNA was measured using atomic absorption spectroscopy and it was found that [PtCl2(hpip)] bound both more rapidly and to a greater extent than did cis-[PtCl2(NH3)2]. The binding of [PtCl2(hpip)] and [PtCl2(en)] (en=ethane-1,2-diamine) to salmon sperm DNA and to synthetic, self-complementary 10-base-pair and 52-base-pair oligonucleotides was studied using enzymatic digestion and HPLC analysis of the products. [PtCl2(hpip)] forms approximately two-fold fewer GpG and ApG intrastrand adducts and concomitantly more monofunctional adducts than does [PtCl2(en)]. In the case of [PtCl2(hpip)], two GpG adducts, corresponding to the different orientations of the hpip ligand with respect to the DNA, were observed in a 1:3.3 ratio. The minor product corresponds to the orientation in which the bulkier propylene chain of the hpip ligand is adjacent to, and makes close contacts with, the floor of the major groove. When the reaction was repeated with a synthetic oligonucleotide decamer duplex, the ratio of the two forms was approximately 1:1.9 and with the 52-mer duplex it was 1:2.4, revealing an apparent systematic dependence of stereoselectivity on nucleotide size. Computer modeling of the two adducts formed by [PtCl2(hpip)] and those formed by [PtCl2(en)] and cis-[PtCl2(NH3)2] revealed that non-bonded interactions between the hpip ligand and the DNA were probably responsible for both the decreased proportion of GpG adducts formed by [PtCl2(hpip)] and the stereoselectivity exhibited in the formation of these adducts. This is the first case in which the stereoselectivity can be ascribed to steric factors alone.     

S1 nuclease sensitivity to cis- and trans-diamminedichloroplatinum(II) modified DNAS: influence of (G+C) content and nucleotide sequence

The sensitivity of S1 nuclease to cis- and trans-(NH3)2PtCl2 modified DNAs is examined as a function of the level of cis- and trans-(NH3)2PtCl2 bound, the % (G+C) content in DNA from different sources and the sequence dependence in poly(dG).poly(dC) and poly(dG-dC).poly(dG-dC). The extent of DNA digested increases with increasing levels of either isomer and is inversely influenced by the % (G+C) content of the DNA. However, the difference in the extent of digestion between the cis-and trans-(NH3)2PtCl2 modified DNAs at equivalent levels of bound isomer follows the order, calf-thymus greater than M. lysodeikticus greater than poly(dG-dC).poly(dG-dC). While there is virtually no difference in the digestion profiles for poly(dG-dC).poly(dG-dC) modified with the two isomers, there is a striking difference in the extent of digestion between cis- and trans-(NH3)2PtCl2 modified poly(dG).poly(dC). These results are discussed in light of the possible modes of binding for cis-(NH3)2PtCl2, previously reported findings on modified DNA and possible implications for modifications in cellular chromatin.     

Multinuclear NMR spectroscopy and antitumor activity of novel platinum(II) complexes with 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines

Novel platinum(II) complexes with 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines have been synthesized and characterized by infrared and multinuclear magnetic resonance spectroscopic techniques (1H, 13C, 15N, 195Pt). The complexes are of two types: [PtCl2(L)2] and [PtCl2(NH3)(L)], where L=5,7-diphenyl-1,2,4-triazolo[1,5-a]pyrimidine (dptp) and 5,7-ditertbutyl-1,2,4-triazolo[1,5-a]pyrimidine (dbtp). Significant 15N NMR upfield shifts (92-95 ppm) were observed for N(3) atom indicating this nitrogen atom as a coordination site. The molecular structure suggest that Pt(II) ion has the square planar geometry with N(3) bonded 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines, N-bonded second ligand (NH3 for cis-[PtCl2(NH3)(L)] or, respectively, 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines for cis-[PtCl2L2]) and two cis chloride anions. The antiproliferative activity in vitro of complexes (1-4) have been tested against the cells of four human cell lines: SW707 rectal adenocarcinoma, A549 non-small cell lung carcinoma, T47D breast cancer and HCV29T bladder cancer. The results indicate a moderate antiproliferative activity of (4) against the cells of rectal, breast and bladder cancer and a marked and selective cytotoxic effect of (1-3) against the cells of all studied human cancer lines.     

5-Fluorouracil-cisplatin adducts with potential antitumor activity

Using 5-fluorouracil (5-FU) and cis-diamminedichloroplatinum(II) (cisplatin, CDDP) as starting compounds, 5-FU-cisplatin adducts cis-[Pt(NH(3))(2)(HFU)Cl] (1) and cis-[Pt(NH(3))(2)(HFU)(2)] (2) were prepared. The obtained complexes were characterized by IR, ES-MS and 1H NMR spectroscopy. Complex 1 reacted with guanosine-5'-monophosphate (5'-GMP) and gave rise to a stable mixed-ligand complex cis-[Pt(NH(3))(2)(HFU)(GMP)] (3), whereas 2 did not undergo a similar reaction. In vitro cell growth inhibition tests of complexes 1 and 2 exhibited moderate antitumor activities against the melanoma B16-BL6 cell line. This work provides the basis for a potential alternative for the combinational use of 5-FU and CDDP in cancer therapy.     

Preparation of platinum(IV) complexes with dipeptide and diimine. X-ray crystal structure and 195Pt NMR spectra

We prepared platinum(IV) complexes containing dipeptide and diimine or diamine, the [PtCl(dipeptide-N,N,O)(diimine or diamine)]Cl complex, where -N,N,O means dipeptide coordinated as a tridentate chelate, dipeptide=glycylglycine (NH(2)CH(2)CON(-)CH(2)COO(-), digly, where two protons of dipeptide are detached when the dipeptide coordinates to metal ion as a tridentate chelate), glycyl-L-alanine (NH(2)CH(2)CON(-)CHCH(3)COO(-), gly-L-ala), L-alanylglycine (NH(2)CH CH(3)CON(-)CH(2)COO(-), L-alagly), or L-alanyl-L-alanine (NH(2)CHCH(3)CON(-)CHCH(3)COO(-), dil-ala), and diimine or diamine=bipyridine (bpy), ethylenediamine (en), N-methylethylenediamine (N-Me-en), or N,N'-dimethylethylenediamine (N,N'-diMe-en). In the complexes containing gly-L-ala or dil-ala, two separate peaks of the (195)Pt NMR spectra of the [PtCl(dipeptide-N,N,O)(diimine or diamine)]Cl complexes appeared in, but in the complexes containing digly or L-alagly, one peak which contained two overlapped signals appeared. One of the two complexes containing gly-L-ala and bpy, [PtCl(gly-L-ala-N,N,O)(bpy)]NO(3), crystallized and was analyzed. This complex has the monoclinic space group P2(1)2(1)2(1) with unit cell dimensions of a=9.7906(3)A, b=11.1847(2)A, c=16.6796(2)A, Z=4. The crystal data revealed that this [PtCl(gly-L-ala-N,N,O)(bpy)]NO(3) complex has the near- (Cl, CH(3)) configuration of two possible isomers. Based on elemental analysis, the other complex must have the near- (Cl, CH(3))-[PtCl(gly-L-ala-N,N,O)(bpy)]NO(3) configuration. The (195)Pt NMR chemical shifts of the near- (Cl, CH(3))-[PtCl(gly-L-ala-N,N,O)(bpy)]NO(3) complex and the far- (Cl, CH(3))-[PtCl(gly-L-ala-N,N,O)(bpy)]NO(3) complex are 0 ppm and -19 ppm, respectively (0 ppm for the Na(2)[PtCl(6)] signal). The additive property of the (195)Pt NMR chemical shift is discussed. The (195)Pt NMR chemical shifts of [PtCl(dipeptide-N,N,O)(bpy)]Cl appeared at a higher field when the H attached to the dipeptide carbon atom was replaced with a methyl group. On the other hand, the (195)Pt NMR chemicals shifts of [PtCl(dipeptide-N,N,O)(diamine)] appeared at a lower field when the H attached to the diamine nitrogen atom was replaced with a methyl group, in the order of [PtCl(digly-N,N,O)(en)]Cl, [PtCl(digly-N,N,O)(N-Me-en)]Cl, and [PtCl(digly-N,N,O)(N,N'-diMe-en)]Cl.     

Effects of spectator ligands on the specific recognition of intrastrand platinum-DNA cross-links by high mobility group box and TATA-binding proteins

The results presented describe the effects of various spectator ligands, attached to a platinum 1,2-intrastand d(GpG) cross-link in duplex DNA, on the binding of high mobility group box (HMGB) domains and the TATA-binding protein (TBP). In addition to cisplatin-modified DNA, 15-base pair DNA probes modified by [Pt(1R,2R-diaminocyclohexane)](2+), cis-[Pt(NH(3))(cyclohexylamine)](2+), [Pt(ethylenediamine)](2+), cis-[Pt(NH(3))(cyclobutylamine)](2+), and cis-[Pt(NH(3))(2-picoline)](2+) were examined. Electrophoretic mobility shift assays show that both the A and B domains of HMGB1 as well as TBP discriminate between different platinum-DNA adducts. HMGB1 domain A is the most sensitive to the nature of the spectator ligands on platinum. The effect of the spectator ligands on protein binding also depends highly on the base pairs flanking the platinated d(GpG) site. Double-stranded oligonucleotides containing the AG*G*C sequence, where the asterisks denote the sites of platination, with different spectator ligands are only moderately discriminated by the HMGB proteins and TBP, but the recognition of dsTG*G*A is highly dependent on the ligands. The effects of HMGB1 overexpression in a BG-1 ovarian cancer cell line, induced by steroid hormones, on the sensitivity of cells treated with [Pt(1R,2R-diaminocyclohexane)Cl(2)] and cis-[Pt(NH(3))(cyclohexylamine)Cl(2)] were also examined. The results suggest that HMGB1 protein levels influence the cellular processing of cis-[Pt(NH(3))- (cyclohexylamine)](2+), but not [Pt((1R,2R)-diaminocyclohexane)](2+), DNA lesions. This result is consistent with the observed binding of HMGB1a to platinum-modified dsTG*G*A probes but not with the binding affinity of HMGB1a and HMGB1 to platinum-damaged dsAG*G*C oligonucleotides. These experiments reinforce the importance of sequence context in platinum-DNA lesion recognition by cellular proteins.     

Induction of Cell Cycle Arrest and Apoptosis by Ruthenium Complex cis-(Dichloro)tetramineruthenium(III) Chloride in Human Lung Carcinoma Cells A549

Lung cancer is one of the leading causes of death in the world, and non-small cell lung carcinoma (NSCLC) accounts for approximately 75-85% of all lung cancers. In the present work, we studied the cytotoxic activity, cell cycle arrest and induction apoptosis of the compound cis-(dichloro)tetramineruthenium(III) chloride {cis-[RuCl(2)(NH(3))(4)]Cl} in human lung carcinoma tumor cell line A549. The results of MTT and trypan blue assays showed that cis-[RuCl(2)(NH(3))(4)]Cl causes reduction in the viability of A549 cells when treating with 95 and 383 μM of the compound for 48 and 72 h. Lower concentrations of the compound (19, 3.8 and 0.38 μM), however, only slightly affected cell viability. The IC(50) value for the compound was about 383 μM. Survival analysis of the A549 cells after treatment with ruthenium(III) compound using long term clonogenic assay showed that it reduced colony formation ability at concentrations of 0.38 and 3.8 μM, and at concentrations of 95 and 383 μM no colonies were observed. Cell cycle analysis showed that compound ruthenium led to an accumulation of A549 cells in S phase and increased in the sub-G1 peak. In addition, cis-(dichloro)tetramineruthenium(III) chloride treatment induced apoptosis, as observed by the increased numbers of annexin V-positive cells and increased messenger RNA expression of caspase-3.