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.     

Uracilato and 5-halouracilato complexes of Cu(II), Zn(II) and Ni(II). X-ray structures of [Cu(uracilato-N(1))(2)(NH(3))(2)].2(H(2)O), [Cu(5-chlorouracilato-N(1))(2)(NH(3))(2)](H(2)O)(2), [Ni(5-chlorouracilato-N(1))(2)(en)(2)].2H(2)O and [Zn(5-chlorouracilato-N(1))(NH(3))(3)].(5-chlorouracilato-N(1)).(H(2)O)

Four new complexes of uracilato and 5-halouracilato with the divalent metal ions Cu(II), Zn(II) and Ni(II) were obtained and structurally characterized. [Cu(uracilato- N(1))(2)(NH(3))(2)].2(H(2)O) (1) and [Cu(5-chlorouracilato-N(1))(2)(NH(3))(2)](H(2)O)(2) (2) complexes present distorted square planar co-ordination geometry around the metal ion. Although an additional axial water molecule is present [Cu(II)-OH(2)=2.89 A (for 1) and 2.52 A (for 2)] in both cases, only in the complex 2 would be considered in the limit of a bond distance. The Zn(II) in [Zn(5-chlorouracilato-N(1))(NH(3))(3)].(5-chlorouracilato-N(1)).(H(2)O) presents a tetrahedral co-ordination with three ammonia molecules and the N(1) of the corresponding uracilato moiety. A non-coordinated uracilato molecule is present as a counterion and a recognition between co-ordinated and free ligands, by means a tandem of H-bonds, should be mentioned. Finally, the complex [Ni(5-chlorouracilato-N(1))(2)(en)(2)] (H(2)O)(2) (where en is ethylenediamine) presents a typical octahedral trans co-ordination with additional hydrogen bonds between 5-chlorouracilato and the NH(2) groups of ethylenediamine units.     

Differential genotoxic effects of antitumor trans-[PtCl(2)(E-iminoether)(2)] and cisplatin in Escherichia coli

In the present work, we measured survival and the platinum on the genome after treatment of repair-proficient or repair-deficient Escherichia coli strains with trans-[PtCl₂(E-iminoether)₂] and compared these results with the effects of “classical” cisplatin. We found that toxicity of antitumor trans-[PtCl₂(E-iminoether)₂] in repair-deficient trains was much less than that of cisplatin. This markedly reduced toxicity was not a consequence of the reduced uptake or low levels of DNA binding in the bacteria cells but rather appeared to reflect DNA binding mode of this trans-platinum drug different from that of cisplatin.     

Heavy metal coprecipitation with hydrozincite [Zn(5)(CO(3))(2)(OH)(6)] from mine waters caused by photosynthetic microorganisms

An iron-poor stream of nearly neutral pH polluted by mine tailings has been investigated for a natural phenomenon responsible for the polishing of heavy metals in mine wastewaters. A white mineralized mat, which was determined to be hydrozincite [Zn(5)(CO(3))(2)(OH)(6)] by X-ray diffraction analysis, was observed in the stream sediments mainly in spring. The precipitate shows a total organic matter residue of 10% dry weight and contains high concentrations of Pb, Cd, Ni, Cu, and other metals. Scanning electron microscopy analysis suggests that hydrozincite is mainly of biological origin. Dormant photosynthetic microorganisms have been retrieved from 1-year-old dry hydrozincite. The autofluorescent microorganisms were imaged by a scanning confocal laser microscope. A photosynthetic filamentous bacterium, classified as Scytonema sp. strain ING-1, was found associated with microalga Chlorella sp. strain SA1. This microbial community is responsible for the natural polishing of heavy metals in the water stream by coprecipitation with hydrozincite.     

Platinum complexes with one radiosensitizing ligand [PtCl2(NH3) (sensitizer)]: radiosensitization and toxicity studies in vitro

Complexes of general formula [PtCl2(NH3)L] with one radiosensitizing ligand per platinum are compared with ligand L alone, complexes with two radiosensitizers per platinum [PtCl2L2], and their analogs with NH3 ligands, with respect to radiosensitizing properties and toxicity in CHO cells. Radiosensitizing ligands, L, were misonidazole, metronidazole, 4(5)-nitroimidazole, and 2-amino-5-nitrothiazole, and the ammine analogs were cis- and trans-DDP [diamminedichloroplatinum(II)] and the monoammine, K[PtCl3(NH3)]. Results are related to a previous study on plasmid DNA binding by these series. The toxicity of the mono series [PtCl2(NH3)L], attributable to DNA binding, is much higher than the corresponding bis complexes, [PtCl2L2]. For L = misonidazole, toxicity is similar to the monoammine, but higher in hypoxic than in aerobic cells. trans-[PtCl2(NH3)-(misonidazole)] is more toxic than the cis isomer. Except for L = 4(5)-nitroimidazole, the complexes [PtCl2(NH3)L] are more toxic than L in air and hypoxia. Hypoxic radiosensitization by the mono complexes is comparable to the monoammine and is not better than free sensitizers, again except for L = 4(5)-nitroimidazole. Significantly lower sensitization is observed in oxic cells. The bis complexes [PtCl2L2], which do not bind to DNA as well as the mono complexes, are less effective radiosensitizers and less toxic than the [PtCl2(NH3)L] series.     

Hypotensive properties and acute toxicity of trans-[Ru(NH(3))(4)P(OEt)(3)(NO)](PF(6))(3), a new nitric oxide donor.

The hypotensive effect and the acute toxicity of trans-[Ru(NH(3))(4)P(OEt)(3)(NO)](PF(6))(3) (RuNO) were investigated in conscious animals. The approximate lethal dose of RuNO is 257.5 micromol/kg in mice i.p. and the IC(50) values evaluated for V79 culture cell cytotoxicity were higher than 2.0 mM, suggesting that the ruthenium species are significantly less toxic than Na(2)[Fe(CN)(5)(NO)] (SNP) species. The RuNO hypotensive effect measured through in-bolus intravenous administration in chronically instrumented normotensive and hypotensive adult male Wistar rats is similar to that exhibited by equivalent doses of SNP. The hypotensive effect of the ruthenium complex is fully inhibited by methylene blue and PTIO, suggesting that the RuNO effect is likely to be primarily dependent on the NO-[cGMP] pathway in the smooth muscle cells.     

DNA binding by antitumor trans-[PtCl2(NH3)(thiazole)]. Protein recognition and nucleotide excision repair of monofunctional adducts

Antitumor effects of cis-diamminedichloroplatinum(II) (cisplatin) and the clinical inactivity of its trans isomer (transplatin) have been considered a paradigm for the classical structure-activity relationships of platinum drugs. However, several new analogues of transplatin which exhibit a different spectrum of cytostatic activity including activity in tumor cells resistant to cisplatin have been recently identified. Analogues containing the planar amine ligand of the general structure trans-[PtCl(2)(NH(3))(L)], where L = planar amine, represent an example of such compounds. DNA is believed to be the major pharmacological target of platinum compounds. To contribute to the understanding of mechanisms underlying the activation of trans geometry in transplatin analogues containing planar amine ligands, various biochemical and biophysical methods were employed in previous studies to analyze the global modifications of natural DNA by trans-[PtCl(2)(NH(3))(L)]. These initial studies have revealed some unique features of the DNA binding mode of this class of platinum drugs. As the monofunctional lesions represent a significant fraction of stable adducts formed in DNA by bifunctional antitumor trans-platinum compounds with planar ligands, we analyzed in the present work short DNA duplexes containing the single, site-specific monofunctional adduct of a representative of this class of platinum drugs, antitumor trans-[PtCl(2)(NH(3))(thiazole)]. It has been shown that, in contrast to the adducts of monodentate chlorodiethylenetriamineplatinum(II) chloride or [PtCl(NH(3))(3)]Cl, the monofunctional adduct of trans-[PtCl(2)(NH(3))(thiazole)] inhibits DNA synthesis and creates a local conformational distortion similar to that produced in DNA by the major 1,2-GG intrastrand CL of cisplatin, which is considered the lesion most responsible for its anticancer activity. In addition, the monofunctional adducts of trans-[PtCl(2)(NH(3))(thiazole)] are recognized by HMGB1 domain proteins and removed by the nucleotide excision repair system similarly as the 1,2-GG intrastrand CL of cisplatin. The results of the present work further support the view that the simple chemical modification of the structure of an inactive platinum compound alters its DNA binding mode into that of an active drug and that processing of the monofunctional DNA adducts of the trans-platinum analogues in tumor cells may be similar to that of the major bifunctional adducts of "classical" cisplatin.     

A new inorganic vasodilator, trans-[Ru(NO)(NH(3))(4)(POEt)(3)](PF(6))(3): hypotensive effect of endothelium-dependent and -independent vasodilators in different hypertensive animals models.

The hypotensive effect of RuNO was investigated in acute and chronic hypertensive rats, as well as in normotensive rats. Acute hypertension rats were used with 30% increase on basal BP (phenylephrine, angiotensin II (Ang II), N(G)-nitro-L-arginine methyl ester (L-NAME), and adult spontaneously hypertensive rats (SHR) (basal BP 168 +/- 3 mm Hg) were used as models for chronic hypertension. Rats were implanted with catheters (iv/ia) for BP measurements and for in bolus administration of RuNO, sodium nitroprusside (SNP), and acetylcholine (Ach) (10, 20, 40 nmol/kg, iv). The principal findings of this study were: (i) The hypotensive response to RuNO was 150% higher in acutely (phenylephrine and Ang II) and chronically (SHR) hypertensive rats than in normotensive rats, except in the case of L-NAME-induced hypertension (deltaMAP = 10 +/- 1.4 mm Hg). Chronic SHR showed 60% increase (deltaMAP = 19 +/- 0.8 mm Hg) in the effect compared to normotensive rats. (ii) The hypotensive response to SNP was lower (60%) in hypertensive rats than in normotensive rats, when compared to RuNO. However, the responses were similar in L-NAME-induced hypertension (deltaMAP = 30 +/- 2 mm Hg). (iii) The vasodilator response to Ach was increased in rats with Ang II-induced hypertension (deltaMAP = 53 +/- 1 mm Hg) and in SHR (deltaMAP = 67 +/- 3 mm Hg). RuNO response was more potent than SNP in hypertensive models and the increment in relation to normotensive was observed in the phenylephrine- and L-NAME-treated rats. This response could be correlated to the different endothelial dysfunction present in each model.     

Apoptosis induction and inhibition of H-ras overexpression by novel trans-[PtCl2(isopropylamine)(amine')] complexes

Hitherto, it has been generally accepted as a paradigm of the biochemical pharmacology of platinum antitumor drugs that a cis configuration of the leaving groups is necessary for antitumor activity of platinum compounds. However, it has been recently observed that certain trans-platinum complexes have both in vitro and in vivo antitumor activity. We previously reported the synthesis, characterization and cytotoxic activity against ras-transformed cells of several trans-[PtCl2LL'] complexes where L and L' are asymmetric aliphatic amines (L = dimethylamine and butylamine, L' = isopropylamine). The results reported in this paper show that the compounds trans-[PtCl2(isopropylamine)(dimethylamine)] and trans-[PtCl2(isopropylamine)(butylamine)] kill Pam 212-ras cisplatin resistant cells through apoptosis induction. Moreover, Western blot data show that both compounds inhibit overexpression of H-ras oncogene in Pam 212-ras cells. Altogether, these data indicate that, in contrast with cis-DDP, the apoptotic activity of these novel trans-Pt(II) compounds in ras-transformed cells is associated with their ability to abolish ras-overexpression.     

Conformation, protein recognition and repair of DNA interstrand and intrastrand cross-links of antitumor trans-[PtCl2(NH3)(thiazole)

Replacement of one ammine in clinically ineffective trans-[PtCl2(NH3)2] (transplatin) by a planar N-heterocycle, thiazole, results in significantly enhanced cytotoxicity. Unlike 'classical' cisplatin {cis-[PtCl2(NH3)2]} or transplatin, modification of DNA by this prototypical cytotoxic transplatinum complex trans-[PtCl2(NH3)(thiazole)] (trans-PtTz) leads to monofunctional and bifunctional intra or interstrand adducts in roughly equal proportions. DNA fragments containing site-specific bifunctional DNA adducts of trans-PtTz were prepared. The structural distortions induced in DNA by these adducts and their consequences for high-mobility group protein recognition, DNA polymerization and nucleotide excision repair were assessed in cell-free media by biochemical methods. Whereas monofunctional adducts of trans-PtTz behave similar to the major intrastrand adduct of cisplatin [J. Kasparkova, O. Novakova, N. Farrell and V. Brabec (2003) Biochemistry, 42, 792-800], bifunctional cross-links behave distinctly differently. The results suggest that the multiple DNA lesions available to trans-planaramine complexes may all contribute substantially to their cytotoxicity so that the overall drug cytotoxicity could be the sum of the contributions of each of these adducts. However, acquisition of drug resistance could be a relatively rare event, since it would have to entail resistance to or tolerance of multiple, structurally dissimilar DNA lesions.     

Complexes trans-[RuCl(2)(nic)(4)] and trans-[RuCl(2)(i-nic)(4)] as free radical scavengers

This study evaluates the action of the new ruthenium complexes trans-RuCl(2)(nic)(4)] (I) and trans-[RuCl(2)(i-nic)(4)] (II) as free radical scavengers. In our experiments, both compounds acted as scavengers of superoxide anion (O(2)*(-)), hydroxyl radicals (HO*) and nitrogen monoxide (formally known as 'nitric oxide'; NO*). In addition, complexes I and II potentiated the release of NO* from S-nitroso-N-acetyl-DL-penicilamine (SNAP), a NO* donor. Complex II, but not I, also decreased the nitrite levels in culture media of activated macrophages. A hypsochromic shift of lambda(max) and a significant change in half-wave potential (E(1/2)) was observed when NO* was added to the Complex II. Thiobarbituric reactive substance (TBARS) levels were significantly reduced in rats treated for 1 week with Complex II plus tert-butylhydroperoxide, when compared to rats treated only with tert-butylhydroperoxide. None of the complexes showed cytotoxicity. These findings support the suggestion that the new ruthenium complexes, especially trans-[RuCl(2)(i-nic)(4)] or its derivatives, might provide potential therapeutic benefits in disorders where reactive nitrogen (RNS) or oxygen (ROS) species are involved.     

Synthesis, crystal structures, and anti-convulsant activities of ternary [Zn(II)(3,5-diisopropylsalicylate)(2)], [Zn(II)(salicylate)(2)] and [Zn(II)(aspirinate)(2)] complexes

Following observations that bis(3,5-diisopropylsalicylato)diaquazinc(II), [Zn(II)(3,5-DIPS)(2)(H(2)O)(2)], had anti-convulsant activity, bis(acetylsalicylate)diaquazinc(II), [Zn(II)(aspirinate)(2)(H(2)O)(2)], and the Zn(II) ternary 1,10-phenanthroline (phen), 2,9-dimethyl-1,10-phenanthroline (neocuproine, NC) or dimethyl sulfoxide (DMSO) complexes of Zn(II)3,5-diisopropylsalicylate, salicylate, and acetylsalicylate were synthesized and spectroscopically characterized. Anti-convulsant and Rotorod toxicity activities of these complexes were determined to examine their anti-convulsant and undesirable central nervous stimulant or depressant activities of these Zn(II) non-steroidal anti-inflammatory agent complexes. Bis(3,5-diisopropylsalicylato)-1,10-phenanthorlinezinc(II), [Zn(II)(3,5-DIPS)(2)(phen)], (1) has one bidentate phen ligand and two mono-deprotonated 3,5-DIPS ligands. One of the carboxylates bonds in an asymmetric chelating mode. The Zn(II) atom exhibits a distorted bicapped rectangular pyramidal environment N(2)O(2)OO (4+1+1 *). In the neocuproine complex, bis(3,5-diisopropylsalicylato)-2,9-dimethyl-1,10-phenanthorlinezinc(II), [Zn(II)(3,5-DIPS)(2)(NC)] (2), the Zn(II) atom has a much more distorted bicapped rectangular pyramidal environment, N(2)O(2)O(2) (4+2 *), compared to 1. The two carboxylate ligands exhibit the same asymmetric coordinating mode with longer metalloelement-oxygen bond distances compared to 1. The space group of [Zn(II)(aspirinate)(2)(H(2)O)(2)] (3), which has been reported as Cc is corrected to C2/c. The zinc atom exhibits a (4+2 *) bicapped square pyramidal environment. While the two ternary phenanthroline-containing complexes, 1 and 2, evidenced weak protection against maximal electroshock (MES)- and subcutaneous Metrazol (scMET) induced seizures, [Zn(II)(3,5-DIPS)(2)(DMSO)(2)], [Zn(II)(aspirinate)(2)(H(2)O)(2)], and bis(salicylato)-1,10-phenanthorlinezinc(II), [Zn(II)(salicylate)(2)(phen)], were found to be particularly useful in protecting against MES and scMET seizures and [Zn(II)(aspirinate)(2)(H(2)O)(2)] and [Zn(II)(salicylate)(2)(phen)] were found to have activity in protecting against Psychomotor seizures, without causing Rotorod toxicity. Activities of these and other Zn(II) complexes of non-steroidal anti-inflammatory agents are consistent with the well-known anti-inflammatory responses of Zn(II)-dependent enzymes. There was also some evidence of Rotorod toxicity consistent with a mechanism of action involving sedative-hypnotic activity of recognized anti-epilepticdrugs.     

The interaction of aquated cis-[(NH3)2Pt(II)]with nucleic acid constituents. II. Modified nucleosides.

The binding interaction of aquated cis-(NH3)2Pt(II) with 1-methylguanosine (1-MeGuo), 7-methylguanosine (7-MeGuo), 1-methyladenosine (1-MeAdo+) and protonated adenosine has been studied using UV difference spectroscopy. The magnitude of the binding constants for the 1 : 1 interaction of cis-(NH3)2Pt(II) with 1-MeDuo and 1-MeAdo are log K = 3.5 and 3.7, respectively. These data are presented and compared to other cis-(NH3)2Pt(II)-nucleoside interactions.     

DNA and glutathione interactions in cell-free media of asymmetric platinum(II) complexes cis- and trans-[PtCl2(isopropylamine)(1-methylimidazole)]: relations to their different antitumor effects

The global modification of mammalian and plasmid DNAs by the novel platinum compounds cis-[PtCl₂(isopropylamine)(1-methylimidazole)] and trans-[PtCl₂(isopropylamine)(1-methylimidazole)] and the reactivity of these compounds with reduced glutathione (GSH) were investigated in cell-free media using various biochemical and biophysical methods. Earlier cytotoxicity studies had revealed that the replacement of the NH₃ groups in cisplatin by the azole and isopropylamine ligands lowers the activity of cisplatin in both sensitive and resistant cell lines. The results of the present work show that this replacement does not considerably affect the DNA modifications by this drug, recognition of these modifications by HMGB1 protein, their repair, and reactivity of the platinum complex with GSH. These results were interpreted to mean that the reduced activity of this analog of cisplatin in tumor cell lines is due to factors that do not operate at the level of the target DNA. In contrast, earlier studies had shown that the replacement of the NH₃ groups in the clinically ineffective trans isomer (transplatin) by the azole and isopropylamine ligands results in a radical enhancement of its activity in tumor cell lines. Importantly, this replacement also markedly alters the DNA binding mode of transplatin, which is distinctly different from that of cisplatin, but does not affect reactivity with GSH. Hence, the results of the present work are consistent with the view and support the hypothesis systematically tested by us and others that platinum drugs that bind to DNA in a fundamentally different manner from that of conventional cisplatin may have altered pharmacological properties.     

Studies on activities, cell up take and DNA binding of four multinuclear complexes of the form: [[trans-PtCl(NH(3))(2)](2)mu-[trans-Pd(NH(3))(2)-(H(2)N(CH(2))(n)NH(2))(2)]]Cl(4) where n=4-7

The activity against human cancer cell lines including ovarian: A2780, A2780(cisR), cell up take, DNA-binding and nature of interaction with pBR322 plasmid DNA have been studied for four multinuclear complexes code named DH4Cl, DH5Cl, DH6Cl and DH7Cl, having the general formula: [[trans-PtCl(NH(3))(2)](2)mu-[trans-Pd(NH(3))(2)-(H(2)N(CH(2))(n)NH(2))(2)]]Cl(4) where n=4, 5, 6 and 7 for DH4Cl, DH5Cl, DH6Cl and DH7Cl, respectively. The compounds are found to exhibit significant anticancer activity against ovarian cancer cell lines: A2780, A2780(cisR) and A2780(ZD0473R). DH6Cl in which the linking diamine has six carbon atoms is found to be the most active compound. As the number of carbon atoms in the linking diamine is decreased below six and increased above six, the activity is found to decrease, illustrating structure-activity relationship. All the multinuclear compounds are believed to form a plethora of long-range interstrand GG adducts with DNA dictated by the sequence of bases in the DNA strands. Increasing prevention of BamH1 digestion with the increase in concentration of the compounds is due to global changes in DNA conformation brought about by interstrand long-range binding of the compounds with DNA.     

The trans labilization of cis-[PtCl2(13CH3NH2)2] by glutathione can be monitored at physiological pH by [1H,13C] HSQC NMR

In order to monitor the trans labilization of cisplatin at physiological pH we have prepared the complex cis-[PtCl₂(¹³CH₃NH₂)₂] and studied its interactions with excess glutathione in aqueous solution at neutral pH by two-dimensional [1H,13C] heteronuclear single-quantum correlation (HSQC) NMR spectroscopy. [1H,13C] HSQC spectroscopy is a good method for following the release of ¹³CH₃NH₂ but is not so good for characterizing the Pt species in solution. In the reaction of cisplatin with glutathione, Pt–S bonds are formed and Pt–NH₃ bonds are broken. The best technique for following the formation of Pt–S bonds of cisplatin is by UV spectroscopy. [1H,13C] HSQC spectroscopy is the best method for following the breaking of the Pt–N bonds. [1H,15N] HSQC spectroscopy is the best method for characterizing the different species in solution. However, the intensity of the peaks in the ¹⁵NH₃–Pt–S region, in [1H,15N] HSQC, reflects a balance between the formation of Pt–S bonds, which increases the signal intensity, and the trans labilization, which decreases the signal intensity. [1H,15N] HSQC spectroscopy and [1H,13C] HSQC spectroscopy are complementary techniques that should be used in conjunction in order to obtain the most accurate information on the interaction of platinum complexes with sulfur-containing ligands.     

A high melting cis-[Pt(NH3)2[d(GpG)]]adduct of a decanucleotide duplex

The [cis-Pt(NH3)2(d(GCCGGATCGC)-N7(4), N7(5))]-d(GCGATCCGGC) duplex has been prepared with Tm = 49 degrees C (vs 58 degrees C for the unplatinated form). NMR of the ten observable imino protons supports a kinked structure with intact base pairing of the duplex on the 3'-side of the d(GpG).cis-Pt chelate (relative to the platinated strand) The modification of the B-DNA type CD spectrum, due to the platinum chelate, is comparable to that observed for the platination (at a 0.05 Pt:base ratio) of the Micrococcus Lysodeikticus DNA (72% GC).     

Influence of cis-[PtCl2(Me2SO)2] on the thermal denaturation of albumin

The interaction of cis-[PtCl2(Me2SO)2] with human serum albumin (HSA) and the sensitivity of the complex towards the thermal denaturation depending on the duration of incubation have been studied by absorption and fluorescence spectroscopy methods. Optimum conditions for cis-[PtCl2(Me2SO)2] binding to HSA have been determined. The results have been compared with the data obtained for HAS-cisplatin complex. It has been found that binding of HSA to cis-[PtCl2(Me2SO)2] does not result in significant structural changes of the protein.     

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.     

Interaction of cis-[Pt(NH3)2(H2O)2](NO3)2 with ribose deoxyribose diguanosine phosphates

The three diguanosine phosphates GpG (4 X 10(-4) M), d(GpG) (10(-5) M), and d(pGpG) (10(-5) M) have been reacted with cis-[Pt(NH3)2(H2O)2](NO3)2 (1 Pt/dinucleotide) in water at pH 5.5 and 37 degrees C. In each case a single product is formed. The three complexes have been characterized by proton nuclear magnetic resonance (1H NMR) and circular dichroism (CD) analyses. They are N(7)-N(7) chelates of the metal with an anti-anti configuration of the bases. They present a conformational change upon deprotonation of guanine N(1)H whose pKa is ca. 8.7 (D2O). Their CD spectra, compared to those of the free dinucleotides, exhibit an increase of ellipticity in the 275-nm region, which can be qualitatively related to the characteristic increase reported for platinated DNA and poly(dG) . poly(dC). These results are in favor of the hypothesis of intrastrand cross-linking of adjacent guanines, by the cis-PtII(NH3)2 moiety, after a local denaturation of DNA.