Reaction of cis and trans isomers of platinum(II) diamminedichloride with purine,adenine and its derivatives in dilute solutions

Reactions of the cytostatic and antitumour agent, cis-platinum(II) diamminedichloride, and its trans isomer with purine, adenine and its N-9 derivatives were followed spectrophotometrically in dilute aqueous solutions. While the cis isomer formed with all compounds studied complexes with the metal-to-ligand ratios ML, ML₂ and M₂L, it was found that the trans isomer did not form the complexes ML₂ with adenosine and AMP. The values of dissociation constants, reaction rate constants and activation energies of the reactions of the cis and trans isomers with purine, adenine and its derivatives were in most cases comparable. Lower stability was exhibited only by the complexes of trans-platinum(II) diamminedichloride (trans-Pt(II)) with AMP as well as by its complexes M₂L with adenine and adenosine. The ability of cis-platinum(II) diamminedichloride (cis-Pt(II)) to react with two adenine residues can explain the greater tendency of the cis isomer to form intrastrand cross-links in nucleic acids as compared with the trans isomer.     

Modulation of oxidative DNA damage and DNA-crosslink formation induced by cis-diammine-tetrachloro-platinum(IV) in the presence of endogenous reductants

Platinum(IV) [Pt(IV)] complex, satraplatin, is currently in clinical trials for the treatment of various cancers. As a key step of the anti-cancer effect exertion, satraplatin is supposed to be reduced by endogenous reductants to platinum(II) [Pt(II)] complex. In this study, we investigated the interaction of DNA, Pt(IV), and the endogenous reductants such as ascorbic acid (AsA) and glutathione (GSH). As a model Pt(IV) compound, cis-diammine-tetrachloro-Pt(IV) [cis-Pt(IV)], which is a prodrug of cisplatin [cis-diammine-dichloro-Pt(II), cis-Pt(II)], was incubated with calf thymus DNA in the presence of AsA or GSH. In the presence of AsA, cis-Pt(IV) induced oxidative DNA damage. Hydroxyl radical scavengers suppressed the AsA-associated oxidative damage, thereby suggesting that hydroxyl radicals are involved in the DNA oxidation. cis-Pt(II)-like CD spectral change and crosslink formation in calf thymus DNA were also observed during this DNA oxidation, suggesting cis-Pt(IV) reduction by AsA and DNA conformational change induced by the newly formed cis-Pt(II) binding to DNA. GSH did not induce oxidative DNA damage likely due to its own hydroxyl radical scavenging ability. Further, GSH suppressed the Pt(II)-mediated DNA conformational change and crosslink formation, suggesting that GSH sequesters the cis-Pt(II) away from DNA by GSH-cis-Pt(II) complex formation.     

The induction of chromosomal structural changes in male chickens by the alkylating agents: triethylene melamine and ethyl methanesulfonate

E. coli chromosomal DNA wastreated with various Pt co-ordiantion compounds and then used as donor DNA in E. coli transformation. Genetic analysis of transformants obtained with Pt-treated DNA showed effects of cis-diamminedichloroplatinum(II) (cis-Pt(II)) and cis-dimethyl-1,3-diaminopropane CL₄ (cis-Pt(IV) (DMDAP) on the processing of DNA. With trans-diamminedichloroplatinum(II) (trans-Pt(II)) appllied in similar concentrations no effects were found.The effects of cis-Pt(II) and cis-Pt(IV) (DMDAP) on the genetic processing were different. The effects of cis-Pt(II) could be explained by assuming intra-strand crosslinks as an important lesion.     

In vitro antitumor activity and interaction with DNA model bases of cis-[PtCl2(iPram)(azole)] complexes and comparison with their trans analogues

Asymmetric cis-platinum(II) complexes with isopropylamine and two different azole ligands were synthesized and characterized with different techniques. In addition, for one of the complexes the X-ray structure was determined. Cytotoxicity tests using several human tumor cell lines, including the cisplatin-sensitive cell line A2780 and its cisplatin-resistant analogue. These results were compared with the results obtained for the trans isomers of the presented complexes and a relation between the structure and the activity was established. It was found that complexes with cis geometry are less active than their trans analogues, in particular in the resistant cell line A2780R. However, complex 1 can overcome cisplatin resistance to a certain extent. In the interaction with GMP, the asymmetric cis-Pt(II) complexes react with similar rates as their trans analogues and they behave as bifunctional species.     

Electronic structure of platinum(II) antitumor complexes and their interactions with nucleic acid bases. I

Using the semi-empirical all-valence method (GRINDOL) (recently modified and extended to transition series elements), electronic structure and intermolecular interactions of the model antitumor Pt(II) compounds with guanine and thioguanine have been calculated. Several possible models of antitumor action of platinum compounds are discussed. It is concluded that cis-Pt(II) complexes with guanine form stable intrastrand N7N7 cross-links (but chelation to the O6 atom is also possible). The trans-isomers of platinum(II) exclusively form interstrand cross-links, but the cis-Pt(II) complexes with thioguanine form almost entirely the N7S five-membered chelates.     

An invitro characterization of interstrand cross-links in DNA exposed to the antitumor drug cis-dichlorodiammineplatinum(II)

The $\text{cis}$-isomer of the antitumor drug dichlorodiammineplatinum(II) [$\text{cis}$-Pt(II)] was tested for its abilty to introduce nicks (single-strand breaks) into supercoiled PM2 DNA. Whereas incubations up to 24 h show no indication of $\text{cis}$-Pt(II)-treated DNA having single-strand breaks, DNA interstrand cross-links were detected in the first 15 min of incubation. Furthermore, the formation of DNA interstrand cross-links was $\text{both}$ inhibited and fully reversed after incubation with 2 mM thiourea.     

Interferon induction by platinum(II)-poly(I)·poly(C) complexes

The effect of the interaction between poly(I)·poly(C) and cis-dichloro-diammineplatinum(II) (cis-Pt), its trans analogue and chloro-diethylene-triamminoplatinum(II) (dien-Pt) on interferon induction activity was investigated. The covalent monodentate fixation of the three compounds on N⁷ of inosine has different effects on the structure and thermostability of poly(I)·poly(C) which is well reflected by the interferon induction activity of the samples. Thus, the sandwich stabilization by dien-Pt at low binding ratios is manifested by an increased interferon induction and a high resistance towards RNAase degradation. The destabilization of the duplex by cis-Pt decreases interferon induction, accompanied by an increase in RNAase sensitivity of the complexes. In the case of trans-Pt the duplex structure is little perturbed and interferon induction is essentially maintained.     

Steric control of DNA interstrand cross-link sites of <Emphasis Type="Italic">trans</Emphasis> platinum complexes: specificity can be dictated by planar nonleaving groups

trans -dichloroplatinum(II) complexes exhibit antitumor activity violate the classical structure-activity relationships of platinum(II) complexes. These novel “nonclassical”trans platinum complexes also comprise those containing planar aromatic amines. Initial studies have shown that these compounds form a considerable amount of DNA interstrand cross-links (up to ∼30%) with a rate markedly higher than clinically ineffective transplatin. The present work has shown, using Maxam-Gilbert footprinting, that trans-[PtCl₂(NH₃)(quinoline)] and trans-[PtCl₂(NH₃)(thiazole)], representatives of the group of new antitumor trans-dichloroplatinum complexes containing planar amines, preferentially form DNA interstrand cross-links between guanine residues at the 5′-GC-3′ sites. Thus, DNA interstrand cross-linking by trans-[PtCl₂(NH₃)(quinoline)] and trans-[PtCl₂(NH₃)(thiazole)] is formally equivalent to that by antitumor cisplatin, but different from clinically ineffective transplatin which preferentially forms these adducts between complementary guanine and cytosine residues. This result shows for the first time that simple chemical modification of the structure of an inactive compound alters its DNA binding site into a DNA adduct of an active drug. Received: 6 January 2000 / Accepted: 8 March 2000     

Differential sensitivity of Fanconi anaemia lymphocytes to the clastogenic action of cis-diamminedichloroplatinum (II) and trans-diamminedichloroplatinum (II)

Summary Fanconi anaemia (FA) lymphocytes were tested for their susceptibility to chromosomal breakage by cis-diamminedichloroplatinum (II) [cis-Pt(II)] and its stereoisomer trans-diamminedichloroplatinum (II) [trans-Pt(II)]. Unlike trans-Pt(II), which is a rather inefficient clastogen, cis-Pt(II) is very efficient in inducing chromosomal breakage in FA cells at concentrations that hardly affect control cells. As both cis-Pt(II) and trans-Pt(II) are capable of inducing DNA interstrand crosslinks but only cis-Pt(II) can induce DNA intra-strand crosslinks, this result suggests that FA cells may be specifically sensitive to the intrastrand type of DNA crosslink.     

Unusual response to bifunctional alkylating agents in a case of Fanconi anaemia

Summary Chromosomal breakage frequencies were determined in Fanconi anaemia (FA) blood cultures treated with various concentrations of the polyfunctional alkylating agents mitomycin C, diepoxybutane, and cis-platinum(II)-diammine-dichloride, for which FA cells have a characteristic hypersensitivity. At concentrations that hardly affected control cultures, three out of four patients tested exhibited a concentration-dependent increase of cells with aberrant chromosomes, with a concomitant increase in the number of chromosomal aberrations per aberrant cell. The fourth patient, a 22-year-old male, was exceptional because with all three clastogens only 40% of his cultured cells exhibited a typical concentration-dependent response, while 60% of his cells responded like those from normal healthy controls. The possible nature and significance of this unusual response is discussed.     

Cytotoxicity, mutagenicity, cellular uptake, DNA and glutathione interactions of lipophilic trans-platinum complexes tethered to 1-adamantylamine

Cytotoxicity and mutagenicity of trans,trans,trans-[PtCl₂(CH₃COO)₂(NH₃)(1-adamantylamine)] [trans-adamplatin(IV)] and its reduced analog trans-[PtCl₂(NH₃)(1-adamantylamine)] [trans-adamplatin(II)] were examined. In addition, the several factors underlying biological effects of these trans-platinum compounds using various biochemical methods were investigated. A notable feature of the growth inhibition studies was the remarkable circumvention of both acquired and intrinsic cisplatin resistance by the two lipophilic trans-compounds. Interestingly, trans-adamplatin(IV) was considerably less mutagenic than cisplatin. Consistent with the lipophilic character of trans-adamplatin complexes, their total accumulation in A2780 cells was considerably greater than that of cisplatin. The results also demonstrate that trans-adamplatin(II) exhibits DNA binding mode markedly different from that of ineffective transplatin. In addition, the reduced deactivation of trans-adamplatin(II) by glutathione seems to be an important determinant of the cytotoxic effects of the complexes tested in the present work. The factors associated with cytotoxic and mutagenic effects of trans-adamplatin complexes in tumor cell lines examined in the present work are likely to play a significant role in the overall antitumor activity of these complexes.     

Platinum(II) and palladium(II) complexes with 2-Acetyl pyridine 4N-ethyl thiosemicarbazone able to overcome the cis-Platin resistance. Structure, antibacterial activity and DNA strand breakage

The reactions of Pd(II) and Pt(II) with 2-Acetyl Pyridine N(4)-Ethyl-Thiosemicarbazones, HAc4Et and 2-Acetyl Pyridine N(4)-1-(2-pyridyl)-piperazinyl Thiosemicarbazone, HAc4PiPiz and 2-Formyl Pyridine N(4)-1-(2-pyridyl)-piperazinyl Thiosemicarbazone, HFo4PiPiz afforded the complexes, [Pd(Ac4Et)], 1, [Pd(HAc4Et)₂]Cl₂, 2 and [Pd(Ac4Et)₂], 3[Pt(Ac4Et)], 4, [Pt(HAc4Et)₂]Cl₂, 5, [Pt(Ac4Et)₂], 6 and [Pd(Fo4PipePiz)Cl], 7, [Pd(Fo4PipePiz)₂], 8, [Pd(Ac4PipePiz)Cl], 9 and [Pd(Ac4PipePiz)₂], 10. The crystal structure of the complex [Pt(Ac4Et)₂], 6 has been solved. The platinum(II) atom is in a square planar environment surrounded by two cis nitrogen atoms and two cis sulfur atoms. The ligands are not equivalent, one being tridentate with (N,N,S) donation, the other being monodentate using only the sulfur atom to coordinate to the metal. The tridentate ligand shows a Z, E, Z configuration while the monodentate ligand shows an E, E, Z. Inter-molecular hydrogen bonds stabilize the structure, while the crystal packing is determined by –, and Pt – C interactions. The antibacterial effect of Pd(II) and Pt(II) complexes were studied in vitro. The complexes were found to have effect on Gram(+) bacteria, while the same complexes showed no bactericidal effect on Gram(–) bacteria. The effect of the Pd(II) and Pt(II) complexes on the in vitro DNA strand breakage was studied by agarose gel electrophoresis. The complexes 1-6 were found to exhibit a cytotoxic potency in a very low micromolar range and to be able to overcome the cisplatin resistance of A2780/Cp8 cells (Kovala-Demertzi et al. 2000).     

Renaturation effects of cis and trans platinum II and IV compounds on calf thymus deoxyribonucleic acid.

The effects of cis dichlorodiammine platinum [cis Pt(II)], trans dichlorodiammine platinum (trans Pt(II)], cis tetrachlorodiammine platinum [cis Pt(IV)], trans tetrachlorodiammine platinum [trans Pt(IV)], and ethylenediaminedichloride platinum [Pt(II)en] on the absorption spectra, and thermal hyper- and hypochromicity of calf thymus DNA were investigated. Platinum-induced renaturation was studied as one parameter of interstrand cross-linking. Based on a DNA cross-linking hypothesis, the tumor-inhibitory platinum compounds cis Pt(II), cis Pt(IV) and Pt(II)en would be expected to induce renaturation following thermal denaturation, whereas the ineffective drugs, trans Pt(II) and trans Pt(IV) would not. All five bind to DNA in such a way as to induce renaturation. However, cis Pt(IV) requires at least a 3- to 4-fold longer incubation time than is required by the other compounds to form the coordination bonds necessary for renaturation. Maximum renaturation with all compounds was observed at a molar Pt/base ratio of 0.05 except cis Pt(IV), with which it was 0.25. The rate of the formation of the platinum-coordinated cross-links by fresh cis Pt(II) suggests two reactions or types of reactions occur. The first is rapid and destabilizes the DNA helix, whereas the second is slow and responsible for renaturation following thermal denaturation. These results suggest that cis Pt(IV) may be activated cellularly and that cross-linking is not the primary mechanism of action of the tumor-inhibitory platinum compounds.     

Specificity of the interaction of DNA-platinum, amount of platinum, and pH measurement]

Interaction between the sodium salt of a DNA extracted from salmon sperm (41% GC) with [Pt(NH₃)₄]Cl₂, [Pt(NH₂? (CH₂)₂? NH? (CH₂)₂? NH₂Cl]Cl, cis-Pt(NH₂? (CH₂)₂? NH₂)Cl₂, cis-Pt(NH₃)₂Cl₂, trans-Pt(NH₃)₂Cl₂, K[Pt(C₂H₄)Cl₃], and K₂[PtCl₄) indicates at least three types of complexation. A correlation is found between the change of pH and the number of platinum atoms fixed per (AT + GC) unit. The first binding site is located on the G-C pairs (guanine–cytosine), most likely the N-7(G) site, as it was shown in a previous study of the guanosine-platinum salts. The fixation of the second platinum atom by the pair (AT + GC) takes place with liberation of protons. In the case of the complexes cis-Pt(NH₂? (CH₂)₂? NH₂)Cl₂, cis-Pt(NH₃)₂Cl₂, and trans-Pt(NH₃)₂Cl₂ the second interaction seems to involve simultaneously the N-7(A) and the N-1(G) and N-3(C) sites. This latter intercrosslink between guanine and cytosine obviously liberates protons and the decrease of pH is related in this case to the trans effect of the platinum compounds. The first two platinum atoms in the reaction of K₂PtCl₄] or the Zeise salt, K[Pt(C₂H₄)Cl₃] with DNA are fixed on the G-C pairs. A maximum of six platinum atoms per (AT + GC) unit were fixed in this case. Preliminary experiments with a DNA extracted from bacteria Micrococcus lysodeikticus (72% GC) give similar results.     

The meiotic drive system on maize abnormal chromosome 10 contains few essential genes

In maize, a distal portion of abnormal chromosome 10 (Ab10) causes the meiotic drive of itself as well as many unlinked heterochromatic regions known as knobs. The Ab10 drive system, which encodes trans- as well as cis-acting components, occupies a large region of chromosome 10L equivalent to 3% of the genome. Here we describe five new structural mutations of Ab10 (five deletions and a duplication) that arose from a screen for meiotic drive mutants. The high frequency of breakage events, detected both genetically and cytologically, suggest that the chromosome may be especially unstable. Very large deletions within the drive system are female-transmissible and plants homozygous for deficiencies lacking much of this interval can be grown to maturity. The data suggest that few genes required for normal growth and development lie within the portion of Ab10 responsible for meiotic drive. These and other published data suggest that meiotic drive systems tend to evolve in gene-sparse or otherwise information-poor regions of the genome where they are less likely to negatively affect individual fitness.     

Mechlorethamine-induced DNA-protein cross-linking in human fibrosarcoma (HT1080) cells

Antitumor nitrogen mustards, such as bis(2-chloroethyl)methylamine (mechlorethamine), are useful chemotherapeutic agents with a long history of clinical application. The antitumor effects of nitrogen mustards are attributed to their ability to induce DNA-DNA and DNA-protein cross-links (DPCs) that block DNA replication. In the present work, a mass spectrometry-based methodology was employed to characterize in vivo DNA-protein cross-linking following treatment of human fibrosarcoma (HT1080) cells with cytotoxic concentrations of mechlorethamine. A combination of mass spectrometry-based proteomics and immunological detection was used to identify 38 nuclear proteins that were covalently cross-linked to chromosomal DNA following treatment with mechlorethamine. Isotope dilution HPLC-ESI(+)-MS/MS analysis of total proteolytic digests revealed a concentration-dependent formation of N-[2-(S-cysteinyl)ethyl]-N-[2-(guan-7-yl)ethyl]methylamine (Cys-N7G-EMA) conjugates, indicating that mechlorethamine cross-links cysteine thiols within proteins to N-7 positions of guanine in DNA.     

Multinuclear magnetic resonance studies of the reactions of the antitumor complexes cis-Pt(L)2X2 and cis-Pt(NH3)(L)X2 (L = cyclobutylamine and cyclopentylamine) with guanosine and other bases and crystal structures of Pt(cyclopentylamine)2I2

The crystal structures of two Pt(cyclopentylamine)₂I₂ compounds were determined by X-ray diffraction methods. Both crystals contain disordered cyclopentylamine ligands. Crystal I contains two independent trans-Pt(cyclopentylamine)₂I₂ molecules and all the C atoms are disordered on two positions. The second crystal (II) is most interesting since it contains both cis- and trans-Pt(cyclopentylamine)₂I₂ isomers in the same unit cell. It was prepared from the recrystallization of the cis isomer in acetone. The C atoms of the trans molecule in crystal II are disordered on two positions, while only one position was determined in the cis molecule, although some of the C thermal factors are quite high. The reactions of cis-Pt(amine)₂X₂ and cis-Pt(NH₃)(amine)X₂ (amine = cyclobutylamine and cyclopentylamine) with guanosine in water were studied in different Pt:guanosine proportions by multinuclear (¹H, ¹⁹⁵Pt and ¹⁵N) magnetic resonance spectroscopy. The presence of several species in solution was observed. For the mixed-cyclobutylamine compound, ¹⁵N NMR has shown that some of the NH₃ ligands have been displaced from the coordination sphere in the presence of an excess of guanosine. The reactions of the two mixed-ligand complexes cis-Pt(NH₃)(amine)Cl₂ with 9-methylguanine, inosine and 9-methylhypoxanthine were also studied in water and the results are discussed.     

Susceptibility of heterochromatin to aphidicolin-induced chromosomal breakage

The distribution of aphidicolin-induced chromosomal lesions was analyzed to determine the relative breakage susceptibility of euchromatin and heterochromatin in the cactus mouse, Peromyscus eremicus. The observed breakage was tested against expected distributions corresponding to the karyotypic proportions of autosomal euchromatin, autosomal heterochromatin, X-chromosomal euchromatin, and X-chromosomal heterochromatin. The distribution of induced breakage was independent of sex but dependent on the individual. In all individuals, there was a highly significant (P0.0001) deficiency in the number of breaks observed as compared to expected in autosomal heterochromatin. Sparse observations in the X chromosome and the absence of breaks in the Y chromosome precluded valid statistical tests of the sex-chromosomal distribution of induced breakage. These data indicate that the autosomal heterochromatin of Peromyscus is resistant to aphidicolin-induced chromosomal breakage and argue against a simple relationship between late replication and a general mechanism for chromosomal fragility.     

Binding study of the drug cis-dichlorodiammineplatinum(II)to G p5′ and dGp5′ by high resolution proton and carbon-13 NMR spectroscopy

The molecular mode of action leading to the anticancer activity of the drug cis-diamminedichloroplatinum(II), cis-DDP or cis-platinum is still the subject of speculation. In the present high field (400 MHz) ¹H NMR study the results on coupling constants for cis- and trans-diammine bis(guanosine- 5′-monophosphate) and (d-guanosine-5′-monophosphate)platinum(II) complexes are presented and discussed. The ¹H and ¹³C NMR chemical shifts obtained are consistent with the drug binding to N7 of each guanine. It has been found that the drug induces different conformational changes in the nucleotide from the trans-DDP isomer.