We present the synthesis and characterization of six new heteroleptic osmium(II) complexes of the type [Os(C^N)(N^N)2]OTf (N^N = 2,2'-bipyridine and dipyrido[3,2-d:2',3'-f]quinoxaline; C^N = deprotonated methyl 1-butyl-2aryl-benzimidazolecarboxylate) with varying substituents in the R3 position of the phenyl ring of the cyclometalating C^N ligand. The new compounds are highly kinetically inert and absorb a full-wavelength range of visible light. An investigation of the antiproliferative activity of the new compounds has been performed using a panel of human cancer and noncancerous 2D cell monolayer cultures under dark conditions and green light irradiation. The results demonstrate that the new Os(II) complexes are markedly more potent than conventional cisplatin. The promising antiproliferative activity of selected Os(II) complexes was also confirmed using 3D multicellular tumor spheroids, which have the characteristics of solid tumors and can mimic the tumor tissue microenvironment. The mechanism of antiproliferative action of complexes has also been investigated and revealed that the investigated Os(II) complexes activate the endoplasmic reticulum stress pathway in cancer cells and disrupt calcium homeostasis.
- MeSH
- benzimidazoly farmakologie MeSH
- homeostáza MeSH
- komplexní sloučeniny * farmakologie MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nádory * MeSH
- osmium farmakologie MeSH
- protinádorové látky * farmakologie MeSH
- vápník MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Compounds containing vicinal diol (glycol) groups, including saccharides, could be modified with sixvalent osmium complexes with nitrogenous ligands, particularly with N,N,N',N'-tetramethylethylenediamine (Os(VI)tem). The modification products are electrochemically active. Here we show that aminosaccharides can also be modified by Os(VI)tem. We studied chitosan oligosaccharides in their acetylated and deacetylated form in 0.2 M Na-phosphate, pH 6.9. Deacetylated chitosan oligosaccharides with free amino groups modified by Os(VI)tem yielded two peaks (peak I' at -0.15 V and peak II' at about -0.38 V) despite the fact that these oligomers contain only one glycol group on the non-reducing end of the molecule. The electrochemical behavior of Os(VI)tem modified deacetylated chitosan oligomers differs from Os(VI)tem modified simple saccharides, containing only glycol groups, predominantly in peak I'. Our results suggest that free amino groups are involved in Os(VI)tem modification of chitosan oligomers.
Conventional chemotherapy is mostly effective in the treatment of rapidly-dividing differentiated tumor cells but has limited application toward eliminating cancer stem cell (CSC) population. The presence of a very small number of CSCs may contribute to the development of therapeutic resistance, metastases, and relapse. Thus, treatment failure by developing novel anticancer drugs capable of effective targeting of CSCs is at present a major challenge for research focused on chemotherapy of cancer. Here, we show that Os(II) complex 2 [Os(η6-pcym)(bphen)(dca)]PF6 (pcym = p-cymene, bphen = bathophenanthroline, and dca = dichloroacetate), is capable of efficient and selective killing CSCs in heterogeneous populations of human breast cancer cells MCF-7 and SKBR-3. Notably, its remarkable submicromolar potency to kill CSCs is considerably higher than that of its Ru analog, [Ru(η6-pcym)(bphen)(dca)]PF6 (complex 1) and salinomycin, one of the most selective CSC-targeting compounds hitherto identified. Furthermore, Os(II) complex 2 reduces the formation, size, and viability of three-dimensional mammospheres which more closely reflect the tumor microenvironment than cells in traditional two-dimensional cultures. The antiproliferation studies and propidium iodide staining using flow cytometry suggest that Os(II) complex 2 induces human breast cancer stem cell death predominantly by necroptosis, a programmed form of necrosis. The results of this study demonstrate the promise of Os(II) complex 2 in treating human breast tumors. They also represent the foundation for further preclinical and clinical studies and applications of Os(II) complex 2 to comply with the emergent need for human breast CSCs-specific chemotherapeutics capable to treat chemotherapy-resistant and relapsed human breast tumors.
- MeSH
- apoptóza účinky léků MeSH
- chloracetáty farmakologie MeSH
- cymeny farmakologie MeSH
- fenantroliny farmakologie MeSH
- komplexní sloučeniny farmakologie MeSH
- lidé MeSH
- lokální recidiva nádoru patologie MeSH
- nádorové buněčné linie MeSH
- nádorové kmenové buňky účinky léků metabolismus MeSH
- nádorové mikroprostředí účinky léků MeSH
- nádory prsu farmakoterapie patologie MeSH
- nekroptóza účinky léků MeSH
- nekróza metabolismus MeSH
- organoplatinové sloučeniny farmakologie MeSH
- osmium farmakologie MeSH
- proliferace buněk účinky léků MeSH
- protinádorové látky farmakologie MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
This review is focused on the reaction of 1,2-diols with ligand complexes of six-valent osmium [Os(VI)L] (where L is a nitrogenous ligand) and possibilities of electrochemical analysis of yielded products. A number of biologically important molecules, such as mono-, oligo- and polysaccharides, RNA and glycoproteins, belong to compounds containing 1,2-diol in their structure. These compounds react with Os(VI)L yielding relatively stable ligand osmate esters which are electrochemically active and suitable to the electrochemical analysis. The ligand osmate esters give redox peaks at the mercury and carbon electrodes. The redox peaks are due to the electrochemical reduction or oxidation of osmium atoms. The osmate esters with some ligands give catalytic peaks at the mercury electrodes. The catalytic peak is due to the catalytic hydrogen evolution and is very sensitive. With the catalytic peak it is possible to measure picomolar concentrations in some cases. We have used reactions of Os(VI)L for the analysis of glycans and glycoproteins in relation to their great importance in biomedicine.
We report on the preparation and thorough characterization of cytotoxic half-sandwich complexes [Ru(η⁶-pcym)(bphen)(dca)]PF₆ (Ru-dca) and [Os(η⁶-pcym)(bphen)(dca)]PF₆ (Os-dca) containing dichloroacetate(1-) (dca) as the releasable O-donor ligand bearing its own cytotoxicity; pcym = 1-methyl-4-(propan-2-yl)benzene (p-cymene), bphen = 4,7-diphenyl-1,10-phenanthroline (bathophenanthroline). Complexes Ru-dca and Os-dca hydrolyzed in the water-containing media, which led to the dca ligand release (supported by ¹H NMR and electrospray ionization mass spectra). Mass spectrometry studies revealed that complexes Ru-dca and Os-dca do not interact covalently with the model proteins cytochrome c and lysozyme. Both complexes exhibited slightly higher in vitro cytotoxicity (IC50 = 3.5 μM for Ru-dca, and 2.6 μM for Os-dca) against the A2780 human ovarian carcinoma cells than cisplatin (IC50 = 5.9 μM), while their toxicity on the healthy human hepatocytes was found to be IC50 = 19.1 μM for Ru-dca and IC50 = 19.7 μM for Os-dca. Despite comparable cytotoxicity of complexes Ru-dca and Os-dca, both the complexes modified the cell cycle, mitochondrial membrane potential, and mitochondrial cytochrome c release by a different way, as revealed by flow cytometry experiments. The obtained results point out the different mechanisms of action between the complexes.
- MeSH
- fenantroliny chemie farmakologie MeSH
- komplexní sloučeniny chemie farmakologie MeSH
- kyselina dichloroctová chemie farmakologie MeSH
- lidé MeSH
- ligandy MeSH
- nádory vaječníků farmakoterapie MeSH
- osmium chemie MeSH
- proliferace buněk účinky léků MeSH
- ruthenium chemie MeSH
- uvolňování léčiv MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
In this paper, we present an electrochemical DNA-protein interaction assay based on a combination of protein-specific immunoprecipitation at magnetic beads (MBIP) with application of oligonucleotide (ON) probes labeled with an electroactive oxoosmium complex (Os,bipy). We show that double-stranded ONs bearing a dT20 tail labeled with Os,bipy are specifically recognized by the tumor suppressor p53 protein according to the presence or absence of a specific binding site (p53CON) in the double-stranded segment. We demonstrate the applicability of the Os,bipy-labeled probes in titration as well as competition MBIP assays to evaluate p53 relative affinity to various sequence-specific or structurally distinct unlabeled DNA substrates upon modulation of the p53-DNA binding by monoclonal antibodies used for the immunoprecipitation. To detect the p53-bound osmium-labeled probes, we took advantage of a catalytic peak yielded by Os,bipy-modified DNA at the mercury-based electrodes, allowing facile determination of subnanogram quantities of the labeled oligonucleotides. Versatility of the electrochemical MBIP technique and its general applicability in studies of any DNA-binding protein is discussed.
- MeSH
- DNA chemie MeSH
- elektrochemické techniky přístrojové vybavení metody MeSH
- elektrody MeSH
- katalýza MeSH
- lidé MeSH
- nádorový supresorový protein p53 chemie MeSH
- oligonukleotidové sondy chemie MeSH
- osmium chemie MeSH
- rtuť chemie MeSH
- vazba proteinů MeSH
- vodík chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
Glycoproteins participate in various biological events, including disease progression. Currently, there is a pressing need for development of new simple and inexpensive methods for glycoprotein carbohydrate component (mostly oligosaccharides, OLSs) analysis and electrochemical methods were little applied in their analysis. Polysaccharides and OLS were long time considered as electroinactive compounds. We show that OLS adducts with six-valent osmium complexes are electroactive and can be determined at mercury and carbon electrodes. Adducts of OLSs with complex of Os(VI) with N,N,N',N'-tetramethylethylenediamine (tmen) can be prepared by mixing of OLS with [Os(VI)tmen] either at 37°C overnight or at 75°C in 10-15min. We modified 3α,6α-mannopentaose (MPO), stachyose and γ-cyclodextrin with [Os(VI)tmen]. The OLS adducts produced CV redox couples at hanging mercury drop electrode (HMDE) and at pyrolytic graphite electrode (PGE). 6nM MPO was determined by conventional adsorptive stripping at HMDE with RSD 5.3% directly in the reaction mixture. Similar determination at PGE was much less sensitive. Using adsorptive transfer (ex situ) stripping at PGE, μL volumes of OLS were sufficient for the analysis. Protein glycosylation stands at present in focus of medicinal chemistry because of its importance in various diseases and their diagnostics. Our paper represents first steps toward application of electrochemical methods in biomedical analysis of OLS.
- MeSH
- adsorpce MeSH
- alkeny chemie MeSH
- časové faktory MeSH
- elektrochemie přístrojové vybavení metody MeSH
- elektrody MeSH
- limita detekce MeSH
- oligosacharidy analýza chemie MeSH
- organokovové sloučeniny chemie MeSH
- osmium chemie MeSH
- rtuť chemie MeSH
- teplota MeSH
- uhlík chemie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
A complex OsO(4), 2,2'-bipyridine (Os,bipy), has been used for electroactive labeling of biopolymers as well as for probing of nucleic acids and protein structure and interactions. In DNA, Os,bipy forms electrochemically active adducts with pyrimidine nucleobases, exhibiting highly selective modification of thymine residues in single-stranded DNA. Here, we show that modification of rare thymine residues (one thymine among several tens of unreactive purine bases) can easily be detected by means of a simple ex situ voltammetric analysis using carbon electrodes. Based on this remarkable sensitivity of detection, Os,bipy has been used as an electroactive probe for unpaired and/or mismatched thymine residues within DNA heteroduplexes. Site-specific chemical modification of the DNA with the Os,bipy has allowed a clear distinction between perfectly base-paired DNA homoduplexes and mismatched heteroduplexes, as well as discrimination among heteroduplexes containing one or two mispaired thymines, a single thymine insertion, or combination of a mispair and an insertion.
Modified 2'-deoxynucleoside triphosphates (dNTPs) bearing [Ru(bpy)(3)](2+) and [Os(bpy)(3)](2+) complexes attached via an acetylene linker to the 5-position of pyrimidines (C and U) or to the 7-position of 7-deazapurines (7-deaza-A and 7-deaza-G) have been prepared in one step by aqueous cross-couplings of halogenated dNTPs with the corresponding terminal acetylenes. Polymerase incorporation by primer extension using Vent (exo-) or Pwo polymerases gave DNA labeled in specific positions with Ru(2+) or Os(2+) complexes. Square-wave voltammetry could be efficiently used to detect these labeled nucleic acids by reversible oxidations of Ru(2+/3+) or Os(2+/3+). The redox potentials of the Ru(2+) complexes (1.1-1.25 V) are very close to that of G oxidation (1.1 V), while the potentials of Os(2+) complexes (0.75 V) are sufficiently different to enable their independent detection. On the other hand, Ru(2+)-labeled DNA can be independently analyzed by luminescence. In combination with previously reported dNTPs bearing ferrocene, aminophenyl, and nitrophenyl tags, the Os-labeled dATP has been successfully used for "multicolor" redox labeling of DNA and for DNA minisequencing.
- MeSH
- barva MeSH
- barvení a značení metody MeSH
- DNA-dependentní DNA-polymerasy chemie MeSH
- DNA chemie MeSH
- elektrochemie MeSH
- luminiscence MeSH
- oligonukleotidy chemie MeSH
- osmium chemie MeSH
- oxidace-redukce MeSH
- reagencia zkříženě vázaná chemie MeSH
- ruthenium chemie MeSH
- Publikační typ
- práce podpořená grantem MeSH
