Nuclear factor-kappa B (NF-кB) comprises a family of protein transcription factors that have a regulatory function in numerous cellular processes and are implicated in the cancer cell response to antineoplastic drugs, including cisplatin. We characterized the effects of DNA adducts of cisplatin and ineffective transplatin on the affinity of NF-кB proteins to their consensus DNA sequence (кB site). Although the кB site-NF-κB protein interaction was significantly perturbed by DNA adducts of cisplatin, transplatin adducts were markedly less effective both in cell-free media and in cellulo using a decoy strategy derivatized-approach. Moreover, NF-κB inhibitor JSH-23 [4-methyl-N¹-(3-phenylpropyl)benzene-1,2-diamine] augmented cisplatin cytotoxicity in ovarian cancer cells and the data showed strong synergy with JSH-23 for cisplatin. The distinctive structural features of DNA adducts of the two platinum complexes suggest a unique role for conformational distortions induced in DNA by the adducts of cisplatin with respect to inhibition of the binding of NF-кB to the platinated кB sites. Because thousands of κB sites are present in the DNA, the mechanisms underlying the antitumor efficiency of cisplatin in some tumor cells may involve downstream processes after inhibition of the binding of NF-κB to κB site(s) by DNA adducts of cisplatin, including enhanced programmed cell death in response to drug treatment.
- MeSH
- adukty DNA chemie genetika metabolismus MeSH
- cisplatina aplikace a dávkování chemie farmakologie MeSH
- DNA nádorová chemie genetika metabolismus MeSH
- fenylendiaminy aplikace a dávkování farmakologie MeSH
- HEK293 buňky MeSH
- konformace nukleové kyseliny MeSH
- konsenzuální sekvence MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nádory vaječníků farmakoterapie genetika metabolismus MeSH
- NF-kappa B antagonisté a inhibitory chemie metabolismus MeSH
- protinádorové látky aplikace a dávkování chemie farmakologie MeSH
- sekvence nukleotidů MeSH
- stereoizomerie MeSH
- synergismus léků MeSH
- vazebná místa genetika MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The effect of replacement of the N,N-chelating ligand 1,10-phenanthroline (phen) in the Ir(III) pentamethylcyclopentadienyl (Cp*) complex [(η(5)-Cp*)(Ir)(phen)Cl](+) (2) with the C,N-chelating ligand 7,8-benzoquinoline (bq) to give [(η(5)-Cp*)(Ir)(bq)Cl] (1) on the cytotoxicity of these Cp*Ir(III) complexes toward cancer cell lines was investigated. Complex 2 is inactive, similar to other Cp*Ir(III) complexes containing the N,N-chelating ligands. In contrast, a single atom change (C(-) for N) in the chelating N,N ligand resulted in potency in human ovarian carcinoma cisplatin-sensitive A2780 cells, and, strikingly, 1 is active in the cisplatin-resistant human breast cancer MCF-7 and A2780/cisR cells. Replacement of the N,N-chelating ligand with the C,N-chelating ligand gives rise to increased hydrophobicity, leading to higher cellular accumulation, higher DNA-bound iridium in cells and higher cytotoxicity. The pathways involved in cellular accumulation of 1 have been further explored and compared with conventional cisplatin. The results show that both energy-independent passive diffusion and energy-dependent transport play a role in accumulation of 1. Further results were consistent with involvement of p-glycoprotein, multidrug resistance-associated protein 1 and glutathione metabolism in the efflux of 1. In contrast, the internalization of 1 mediated by the endocytotic uptake pathway(s) seems less likely. Understanding the factors which contribute to the mechanism of cellular accumulation of this Ir(III) complex can now lead to the design of structurally similar metal complexes for antitumor chemotherapy.
- MeSH
- chelátory chemie farmakokinetika farmakologie MeSH
- chemorezistence MeSH
- cisplatina farmakologie MeSH
- iridium chemie farmakokinetika farmakologie MeSH
- komplexní sloučeniny chemie farmakokinetika farmakologie MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nádory prsu farmakoterapie metabolismus MeSH
- proteiny spojené s mnohočetnou rezistencí k lékům metabolismus MeSH
- protinádorové látky chemie farmakokinetika farmakologie MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The new trinuclear tridentate Pt(II) complex [Pt(3)Cl(3)(hptab)](3+) (1; hptab = N,N,N',N',N'',N''-hexakis(2-pyridylmethyl)-1,3,5-tris(aminomethyl)benzene) exhibits promising cytotoxic effects in human and mouse tumor cells including those resistant to conventional cisplatin (Dalton Trans. 2006, 2617; Chem. Eur. J. 2009, 15, 5245). The present study is focused on the molecular pharmacology of 1, in particular on its interactions with DNA (which is the major pharmacological target of platinum antitumor drugs), to elucidate more deeply the mechanism underlying its antitumor effects. Results obtained with the aid of methods of molecular biophysics and pharmacology reveal new details of DNA modifications by 1. Complex 1 binds to DNA forming in the absence of proteins and molecular crowding agents mainly trifunctional intrastrand cross-links. In these DNA adducts all three Pt(II) centers of 1 are coordinated to DNA base residues, which leads to extensive conformational alterations in DNA. An intriguing aspect of the DNA-binding mode of this trinuclear Pt(II) complex 1 is that it can cross-link proteins to DNA. Even more interestingly, 1 can cross-link in the presence of molecular crowding agent, which mimics environmental conditions in cell nucleus, two DNA duplexes in a high yield--a feature observed for the first time for antitumor trinuclear platinum complexes. Thus, the concept for the design of agents capable of forming intramolecular tridentate DNA adducts, DNA-protein and interduplex DNA-DNA cross-links based on trinuclear tridentate Pt(II) complexes with semirigid aromatic linkers may result in new compounds which exhibit a variety of biological effects and can be also useful in nucleic acids research.
- MeSH
- DNA chemie MeSH
- lidé MeSH
- molekulární struktura MeSH
- myši MeSH
- organoplatinové sloučeniny chemie metabolismus MeSH
- protinádorové látky chemie metabolismus MeSH
- reagencia zkříženě vázaná chemie metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- nádory radioterapie terapie MeSH
- radioterapie metody trendy využití MeSH
- Publikační typ
- abstrakt z konference MeSH
A combination of biophysical, biochemical, and computational techniques was used to delineate mechanistic differences between the platinum-acridine hybrid agent [PtCl(en)(L)](NO(3))(2) (complex 1, en = ethane-1,2-diamine, L = 1-[2-(acridin-9-ylamino)ethyl]-1,3-dimethylthiourea) and a considerably more potent second-generation analogue containing L' = N-[2-(acridin-9-ylamino)ethyl]-N-methylpropionamidine (complex 2). Calculations at the density functional theory level provide a rationale for the binding preference of both complexes for guanine-N7 and the relatively high level of adenine adducts observed for compound 1. A significant rate enhancement is observed for binding of the amidine-based complex 2 with DNA compared with the thiourea-based prototype 1. Studies conducted with chemical probes and on the bending and unwinding of model duplex DNA suggest that adducts of complex 2 perturb B-form DNA more severely than complex 1, however, without denaturing the double strand and significantly less than cisplatin. Circular and linear dichroism spectroscopies and viscosity measurements suggest that subtle differences exist between the intercalation modes and adduct geometries of the two complexes. The adducts formed by complex 2 most efficiently inhibit transcription of the damaged DNA by RNA polymerase II. Not only do complexes 1 and 2 cause less distortion to DNA than cisplatin, they also do not compromise the thermodynamic stability of the modified duplex. This leads to a decreased or negligible affinity of HMG domain proteins for the adducts formed by either Pt-acridine complex. In a DNA repair synthesis assay the lesions formed by complex 2 were repaired less efficiently than those formed by complex 1. These significant differences in DNA adduct formation, structure, and recognition between the two acridine complexes and cisplatin help to elucidate why compound 2 is highly active in cisplatin-resistant, repair proficient cancer cell lines.
- MeSH
- adukty DNA chemie MeSH
- akridiny chemie metabolismus farmakologie MeSH
- amidiny chemie metabolismus farmakologie MeSH
- B-DNA chemie metabolismus MeSH
- cisplatina analogy a deriváty chemie metabolismus farmakologie MeSH
- DNA chemie metabolismus MeSH
- genetická transkripce účinky léků MeSH
- HeLa buňky MeSH
- interkalátory chemie metabolismus farmakologie MeSH
- kinetika MeSH
- konformace nukleové kyseliny účinky léků MeSH
- lidé MeSH
- oprava DNA účinky léků MeSH
- organoplatinové sloučeniny chemie metabolismus farmakologie MeSH
- protein - isoformy metabolismus MeSH
- protein HMGB1 metabolismus MeSH
- protinádorové látky chemie metabolismus farmakologie MeSH
- racionální návrh léčiv MeSH
- thiomočovina chemie metabolismus farmakologie MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
- srovnávací studie MeSH
The primary objective was to understand more deeply the molecular mechanism underlying different antitumor effects of dinuclear Pt(II) complexes containing aromatic linkers of different length, {[cis-Pt(NH(3))(2)Cl](2)(4,4'-methylenedianiline)}(2+) (1) and {[cis-Pt(NH(3))(2)Cl](2)(alpha,alpha'-diamino-p-xylene)}(2+) (2). These complexes belong to a new generation of promising polynuclear platinum drugs resistant to decomposition by sulfur nucleophiles which hampers clinical use of bifunctional polynuclear trans Pt(II) complexes hitherto tested. Results obtained with the aid of methods of molecular biophysics and pharmacology reveal differences and new details of DNA modifications by 1 and 2 and recognition of these modifications by cellular components. The results indicate that the unique properties of DNA interstrand cross-links of this class of polynuclear platinum complexes and recognition of these cross-links may play a prevalent role in antitumor effects of these metallodrugs. Moreover, the results show for the first time a strong specific recognition and binding of high-mobility-group-domain proteins, which are known to modulate antitumor effects of clinically used platinum drugs, to DNA modified by a polynuclear platinum complex.
- MeSH
- adukty DNA chemie metabolismus MeSH
- cisplatina chemie metabolismus farmakologie MeSH
- HeLa buňky MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- poškození DNA účinky léků fyziologie MeSH
- protinádorové látky chemie metabolismus farmakologie MeSH
- reagencia zkříženě vázaná chemie metabolismus farmakologie MeSH
- viabilita buněk účinky léků fyziologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
The global modification of mammalian and plasmid DNAs by the novel platinum compounds cis-[PtCl(2)(isopropylamine)(1-methylimidazole)] and trans-[PtCl(2)(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 NH(3) 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 NH(3) 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.
- MeSH
- bezbuněčný systém MeSH
- cirkulární dichroismus MeSH
- DNA chemie účinky léků MeSH
- financování organizované MeSH
- glutathion chemie účinky léků MeSH
- kultivační média chemie MeSH
- lidé MeSH
- organoplatinové sloučeniny farmakologie chemie MeSH
- protinádorové látky farmakologie chemie MeSH
- spektrofotometrie ultrafialová MeSH
- stereoizomerie MeSH
- vazebná místa MeSH
- Check Tag
- lidé MeSH
The different antitumor and other biological effects of the third-generation antitumor platinum drug oxaliplatin [(1R,2R-diamminocyclohexane)oxalatoplatinum(II)] in comparison with those of conventional cisplatin [cis-diamminedichloridoplatinum(II)] are often explained by the ability of oxaliplatin to form DNA adducts of different conformation and consequently to exhibit different cytotoxic effects. This work describes, for the first time, the structural and biochemical characteristics of the interstrand cross-links of oxaliplatin. We find that: 1) DNA bending, unwinding, thermal destabilization, and delocalization of the conformational alteration induced by the cross-link of oxaliplatin are greater than those observed with the cross-link of cisplatin; 2) the affinity of high-mobility-group proteins (which are known to mediate the antitumor activity of platinum complexes) for the interstrand cross-links of oxaliplatin is markedly lower than for those of cisplatin; and 3) the chirality at the carrier 1,2-diaminocyclohexane ligand can affect some important structural properties of the interstrand cross-links of cisplatin analogues. Thus, the information contained in the present work is also useful for a better understanding of how the stereochemistry of the carrier amine ligands of cisplatin analogues can modulate their anticancer and mutagenic properties. The significance of this study is also reinforced by the fact that, in general, interstrand cross-links formed by various compounds of biological significance result in greater cytotoxicity than is expected for monofunctional adducts or other intrastrand DNA lesions. Therefore, we suggest that the unique properties of the interstrand cross-links of oxaliplatin are at least partly responsible for this drug's unique antitumor effects.
- MeSH
- cirkulární dichroismus MeSH
- cisplatina farmakologie metabolismus MeSH
- cyklohexylaminy farmakologie chemie MeSH
- denaturace nukleových kyselin účinky léků MeSH
- DNA genetika chemie MeSH
- financování organizované MeSH
- konformace nukleové kyseliny účinky léků MeSH
- ligandy MeSH
- organoplatinové sloučeniny farmakologie chemie MeSH
- protein HMGB1 metabolismus MeSH
- protinádorové látky farmakologie chemie MeSH
- reagencia zkříženě vázaná chemie MeSH
- sekvence nukleotidů MeSH
- stereoizomerie MeSH
- substrátová specifita MeSH
- tranzitní teplota účinky léků MeSH
Downstream processes that discriminate between DNA adducts of a third generation platinum antitumor drug oxaliplatin and conventional cisplatin are believed to be responsible for the differences in their biological effects. These different biological effects are explained by the ability of oxaliplatin to form DNA adducts more efficient in their biological effects. In this work conformation, recognition by HMG domain protein and DNA polymerization across the major 1,2-GG intrastrand cross-link formed by cisplatin and oxaliplatin in three sequence contexts were compared with the aid of biophysical and biochemical methods. The following major differences in the properties of the cross-links of oxaliplatin and cisplatin were found: i), the formation of the cross-link by oxaliplatin is more deleterious energetically in all three sequence contexts; ii), the cross-link of oxaliplatin bends DNA slightly but systematically less in all sequence contexts tested; iii), the affinity of HMG domain protein to the cross-link of oxaliplatin is considerably lower independent of the sequence context; and iv), the Klenow fragment of DNA polymerase I pauses considerably more at the cross-link of oxaliplatin in all sequence contexts tested. We have also demonstrated that the chirality at the carrier ligand of oxaliplatin can affect its biological effects.
