DNA–protein crosslinks
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- MeSH
- aminokyseliny MeSH
- desmosin MeSH
- glykosylace MeSH
- kolagen enzymologie patofyziologie MeSH
- oxidační stres MeSH
- stárnutí MeSH
- Publikační typ
- přehledy MeSH
- MeSH
- adukty DNA chemie MeSH
- aminokyseliny chemie MeSH
- cisplatina chemie MeSH
- finanční podpora výzkumu jako téma MeSH
- krysa rodu rattus MeSH
- molekulární modely MeSH
- protein HMGB1 chemie izolace a purifikace MeSH
- protinádorové látky chemie MeSH
- reagencia zkříženě vázaná MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
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
- MeSH
- adukty DNA chemie metabolismus MeSH
- cisplatina chemie toxicita MeSH
- DNA biosyntéza MeSH
- financování vládou MeSH
- konformace nukleové kyseliny MeSH
- oprava DNA MeSH
- organoplatinové sloučeniny chemie toxicita MeSH
- proteiny s vysokou pohyblivostí metabolismus MeSH
- protinádorové látky chemie toxicita MeSH
- reagencia zkříženě vázaná chemie toxicita MeSH
- thiazoly chemie toxicita MeSH
Recognition and processing by cellular proteins of DNA modified by platinum complexes have been suggested to be relevant to the mechanism of their antitumor activity. Platinum complexes form on DNA various mono- and bifunctional adducts. It has already been described by other authors that intrastrand cross-links formed on DNA by antitumor cis-diamminedichloroplatinum(II) (cisplatin) between neighboring purine residues are recognized by several DNA-binding proteins. In contrast, these proteins do not recognize the intrastrand cross-links formed on DNA by cisplatin or its clinically ineffective trans isomer (transplatin) between nonadjacent base residues. An eventuality heretofore not addressed is that DNA interstrand cross-links (ICLs) of platinum compounds may be recognized by and bound to DNA-binding proteins. DNA probes of 110 base pairs (bp) were constructed containing five equally spaced ICLs of cisplatin or transplatin. These ICLs were formed at specific sites at which these adducts are preferentially formed in natural DNA. Gel electrophoresis mobility shift and competition assays with these probes were used to investigate the specific recognition and binding of the calf thymus HMG1 protein to the DNA ICLs of both platinum isomers. The ICL of antitumor cisplatin was recognized by and bound to the HMG1 protein with a similar affinity as the 1,2-intrastrand d(GpG) cross-link of this drug. The protein binding to the ICL is selective for the DNA modification by cisplatin, but not by chemotherapeutically inactive transplatin.(ABSTRACT TRUNCATED AT 250 WORDS)
- MeSH
- adukty DNA * metabolismus MeSH
- cisplatina * metabolismus MeSH
- DNA footprinting MeSH
- DNA vazebné proteiny * metabolismus MeSH
- endodeoxyribonukleasy metabolismus MeSH
- kompetitivní vazba MeSH
- konformace nukleové kyseliny MeSH
- molekulární sekvence - údaje MeSH
- poškození DNA MeSH
- proteiny s vysokou pohyblivostí metabolismus MeSH
- protinádorové látky * metabolismus MeSH
- reagencia zkříženě vázaná * metabolismus MeSH
- sekvence nukleotidů MeSH
- substrátová specifita MeSH
- vazba proteinů MeSH
- Publikační typ
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Závěrečná zpráva o řešení grantu Interní grantové agentury MZ ČR
Přeruš. str. : tab., grafy ; 32 cm
Cytotoxicita platinových komplexů závisí v nádorových buňkách na typu,počtu a distribuci neopravitelných poškození DNA.Oprava poškozené DNA je tedy důležitým faktorem,který určuje klinickou účinnost.Testovány budou následující pracovní hypotézy. Rozpoznání aduktů vytvořených na DNA komplexy platiny složkami nukleotidové excizní opravy(NER),vyskytujícími se v lidských buňkách,závisí na schopnosti těchto aduktů destabilizovat dvoušroubovici DNA. Aby bylo dosaženo klinicky využitelné protinádorové účinnosti,je nezbytný vznik takového aduktu,který narušuje pravidelné uspořádání dvojité šroubovice DNA,aniž by došlo ve výrazné míře k vyvolání dějů vedoucích k NER. S cílem prověřit tyto hypotézy experimentálně,budou připraveny próby DNA obsahující jediný adukt komplexu platiny v předem zvolených sekvencích nukleotidů.Testována bude účinnost složek lidského NER tyto adukty opravovat.; Cytotoxicity of platinum antitumor drugs depends on the type, number and distribution of unrepaired DNA lesions in cancer cells.Dna repair is likely to be an important determinant of clinical efficacy.The following hypothesis will be tested: Recognitionof platinum-DNA adducts by human nucleotide excision repair (NER) depends on their ability to destabilize DNA double-helix. To achieve biological and clinical efficacy,it is necessary to obtain platinum-DNA adducts that distort the double helix without eliciting substantial exision repair responses. To test these hypotheses experimentally,a series of DNA substrates containing a single site-specific platinum adduct at a specific position will be designed and purified.The capacity of human NER to processthese site-directed adducts will be tested.
Linear or branched 1,3-diketone-linked thymidine 5'-O-mono- and triphosphate were synthesized through CuAAC click reaction of diketone-alkynes with 5-azidomethyl-dUMP or -dUTP. The triphosphates were good substrates for KOD XL DNA polymerase in primer extension synthesis of modified DNA. The nucleotide bearing linear 3,5-dioxohexyl group (HDO) efficiently reacted with arginine-containing peptides to form stable pyrimidine-linked conjugates, whereas the branched 2-acetyl-3-oxo-butyl (PDO) group was not reactive. Reaction with Lys or a terminal amino group formed enamine adducts that were prone to hydrolysis. This reactive HDO modification in DNA was used for bioconjugations and cross-linking with Arg-containing peptides or proteins (e.g. histones).
- MeSH
- arginin chemie MeSH
- DNA chemická syntéza chemie MeSH
- histony chemie MeSH
- ketony chemická syntéza chemie MeSH
- nádorový supresorový protein p53 chemie MeSH
- peptidy chemie MeSH
- proteiny chemie MeSH
- reagencia zkříženě vázaná chemická syntéza chemie MeSH
- sérový albumin hovězí chemie MeSH
- skot MeSH
- thiminnukleotidy chemická syntéza chemie MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
We show by x-ray crystallography that the complex trans, trans, trans-[Pt(N(3))(2)(OH)(2)(NH(3))(py)] (1) contains an octahedral Pt(IV) center with almost linear azido ligands. Complex 1 is remarkably stable in the dark, even in the presence of cellular reducing agents such as glutathione, but readily undergoes photoinduced ligand substitution and photoreduction reactions. When 1 is photoactivated in cells, it is highly toxic: 13-80 x more cytotoxic than the Pt(II) anticancer drug cisplatin, and ca. 15 x more cytotoxic toward cisplatin-resistant human ovarian cancer cells. Cisplatin targets DNA, and DNA platination levels induced in HaCaT skin cells by 1 were similar to those of cisplatin. However, cisplatin forms mainly intrastrand cis diguanine cross-links on DNA between neighboring nucleotides, whereas photoactivated complex 1 rapidly forms unusual trans azido/guanine, and then trans diguanine Pt(II) adducts, which are probably mainly intrastrand cross-links between two guanines separated by a third base. DNA interstrand and DNA-protein cross-links were also detected. Importantly, DNA repair synthesis on plasmid DNA platinated by photoactivated 1 was markedly lower than for cisplatin or its isomer transplatin (an inactive complex). Single-cell electrophoresis experiments also demonstrated that the DNA damage is different from that induced by cisplatin or transplatin. Cell death is not solely dependent on activation of the caspase 3 pathway, and, in contrast to cisplatin, p53 protein did not accumulate in cells after photosensitization of 1. The trans diazido Pt(IV) complex 1 therefore has remarkable properties and is a candidate for use in photoactivated cancer chemotherapy.
- MeSH
- adukty DNA chemie MeSH
- DNA chemie MeSH
- elektroforéza MeSH
- farmaceutická chemie * metody MeSH
- fotochemie metody MeSH
- geny p53 MeSH
- guanin chemie MeSH
- konformace nukleové kyseliny MeSH
- krystalografie rentgenová MeSH
- lidé MeSH
- molekulární konformace MeSH
- nádorové buněčné linie MeSH
- nádorový supresorový protein p53 metabolismus MeSH
- organoplatinové sloučeniny * farmakologie chemie MeSH
- platina * chemie MeSH
- protinádorové látky * farmakologie chemie MeSH
- reagencia zkříženě vázaná chemie MeSH
- světlo MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Chemoresistance poses one of the most significant challenges of cancer therapy. Carboplatin (CbPt) is one of the most used chemotherapeutics in ovarian cancer (OVC) treatment. MRE11 constitutes a part of homologous recombination (HR), which is responsible for the repair of CbPt-induced DNA damage, particularly DNA crosslinks. The study's main aim was to address the role of HR in CbPt chemoresistance in OVC and to evaluate the possibility of overcoming CbPt chemoresistance by Mirin-mediated MRE11 inhibition in an OVC cell line. Lower expression of MRE11 was associated with better overall survival in a cohort of OVC patients treated with platinum drugs (TCGA dataset, P < 0.05). Using in vitro analyses, we showed that the high expression of HR genes drives the CbPt chemoresistance in our CbPt-resistant cell line model. Moreover, the HR inhibition by Mirin not only increased sensitivity to carboplatin (P < 0.05) but also rescued the sensitivity in the CbPt-resistant model (P < 0.05). Our results suggest that MRE11 inhibition with Mirin may represent a promising way to overcome OVC resistance. More therapy options will ultimately lead to better personalized cancer therapy and improvement of patients' survival.
- MeSH
- chemorezistence * genetika účinky léků MeSH
- homologní protein MRE11 * genetika MeSH
- karboplatina * farmakologie terapeutické užití MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nádory vaječníků * farmakoterapie genetika MeSH
- poškození DNA účinky léků MeSH
- protinádorové látky farmakologie terapeutické užití MeSH
- regulace genové exprese u nádorů účinky léků MeSH
- rekombinační oprava DNA * účinky léků MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
