Synthesis, biological evaluation, and modeling of dimeric PPI analogues as novel DNA minor groove binders

A series of symmetrical dimeric proton pump inhibitor (PPI) analogues, designed as novel type DNA minor groove binders, was synthesized and evaluated for anti-tumor activity. Some of these new compounds showed IC(50) values below 10 microM in an in vitro anti-tumor test. A molecular modeling study was performed to confirm the sequence selectivity of these compounds towards AT base pairs in DNA. Two effective compounds were selected and docked into the minor groove of DNA. The snug binding may be responsible for their cytotoxic and anti-tumor effects.     

Guanidiniocarbonylpyrrole–Aryl Derivatives: Structure Tuning for Spectrophotometric Recognition of Specific DNA and RNA Sequences and for Antiproliferative Activity

We present a systematic study of different guanidiniocarbonylpyrrole‐aryl derivatives designed to interact with DNA or RNA both through intercalation of an aromatic moiety into the base stack of the nucleotide and through groove binding of a guanidiniocarbonylpyrrole cation. We varied 1) the size of the aromatic ring (benzene, naphthalene, pyrene and acridine), 2) the length and flexibility of the linker connecting the two binding groups, and 3) the total number of positive charges present at different pH values. The compounds and their interactions with DNA and RNA were studied by UV/Vis, fluorescence and CD spectroscopy. Antiproliferative activities against human tumour cell lines were also determined. Our studies show that efficient interaction with, for example, DNA requires a significantly large aromatic ring (pyrene) connected through a flexible linker to the pyrrole moiety. However, a positive charge, as in 12 , is also needed. Compound 12 allows for base‐pair‐selective recognition of ds‐DNA at physiological pH values. The antiproliferative activities of these compounds correlate with their binding affinities towards DNA, suggesting that their biological effects are most probably due to DNA binding.     

Antiviral Activity of Synthetic Aminopyrrolic Carbohydrate Binding Agents: Targeting the Glycans of Viral gp120 to Inhibit HIV Entry

The binding abilities of a set of structurally related aminopyrrolic synthetic receptors for mannosides, endowed with antimycotic activity against yeast and yeast‐like pathogens bearing mannoproteins on their cell surface, have been investigated towards the highly mannosylated gp120 and gp41 glycoproteins of the HIV envelope. A pronounced binding interaction with both glycoproteins was observed by SPR for most of the investigated compounds. Comparison of their binding properties towards the glycoproteins with their binding affinities toward mannosides revealed a direct correlation, supporting their role as carbohydrate binding agents (CBAs). Cytostatic activity studies revealed antiproliferative activity dependent on the nature and the structure of compounds. Antiviral activity studies against a broad panel of DNA and RNA viruses showed inhibitory effect against HIV infection of the T‐lymphocyte CEM cell line for two compounds, suggesting antiviral activity similar to other CBAs, such as the nonpeptidic pradimicin antibiotics.     

Novel Bleomycin Analogues: Synthesis,Antitumor Activity,and Interaction with DNA

The novel analogues 11 – 16 of bleomycin A₆ ( 3 ) were obtained by selective protection of the primary‐amine function of the β‐aminoalaninamide moiety of 3 by means of coordination with Cu^II ions, condensation with an aliphatic or aromatic acid R′COOH in the presence of dicyclohexylcarbodiimide, and demetalization (Scheme). The antitumor activity against HeLa and BGC‐823 in vitro, binding property with CT‐DNA, and cleavage potency towards pBR322 DNA were also studied (Tables 1–3). All the compounds 11 – 16 displayed significant antitumor activity, which was enhanced as the hydrophobicity of the C‐terminus substituent R′ increased, but decreased as the DNA‐binding affinity increased. There was a negative relationship between DNA‐cleavage potency and binding affinity to DNA in this series of compounds.     

Interactions of Phenanthroline Compounds with i-Motif DNA

The interactions of each of three phenanthroline derivatives 1, 2 and 3 with the human telomeric/-motif DNA were investigated. The results suggest these compounds are potent binders. The compounds could stabilize the structure of i-motif DNA by π-π stacking. Moreover, the binding constants of the compounds with/-motif DNA were （2.71-8.12）×10^4 L·mol^-1, and the binding stoichiometry ratio was 1：1. CD studies reveal that the binding by phenanthroline comoounds perturbs the conformation of i-motif DNA.     

A mass spectrometric investigation of non-covalent interactions between ruthenium complexes and DNA

Electrospray ionisation mass spectrometry was used to investigate reactions between six ruthenium compounds and three different non self-complementary duplex oligonucleotides containing 16 base pairs. Each of the compounds studied formed non-covalent complexes containing between one and five ruthenium molecules bound to DNA. Competition experiments involving duplex 16mers and pairs of ruthenium compounds were used to determine the order of relative binding affinities of the metal compounds. Other competition experiments involving ruthenium compounds, and the organic DNA binding agents daunomycin and distamycin, provided information about the sites and modes of DNA binding of the ruthenium compounds.     

Studies on the Interaction of Novel Organogermanium Sesquioxides with DNA

The affinity and mode of interaction of four novel organogermanium sesquioxides with calf thymus DNA(CT-DNA) and two synthetic oligonucleotides, d(AT)₂₂d(AT)₂₂ and d(GC)₂₂d(GC)₂₂, were investigated by a combination of absorption spectroscopy, DNA thermal denaturalization method, viscosity method, fluorometric technique, and competitive binding study with ethidium bromide(EB). The results show that the organogermanium compounds can interact with DNA by intercalation, the binding ability of the compounds to CT-DNA and the synthetic oligonucleotides was found to be modest(in comparison to the proven intercalators), with binding constants on the order of 10³―10⁵ L/mol, respectively. Generally, the binding of the organogermanium sesquioxides with naphthalene moiety to DNA is stronger than that of the compounds with anthraquinone moiety. And the compounds with anthraquinone moiety have preference for binding to AT-rich duplexes, whereas the compounds with naphthalene moiety have a little preference for binding to GC-rich duplexes. DNA may be the primary effect target.     

Biological fingerprinting analysis by liquid chromatography/mass spectrometry for evaluation of DNA structural selectivity of multiple compounds in natural products

A method by combination of centrifugal ultrafiltration (CUF) sampling with liquid chromatography-mass spectrometry (LC-MS) analysis was established to evaluate the DNA structure and sequence selectivity of the multiple compounds in a small molecule library. The developed method was applied to analyze the extracts of natural products Coptis chinensis Franch and Rheum palmatum (L.). From the obtained biological fingerprinting chromatograms, 7 compounds in C. chinensis Franch and 11 in R. palmatum (L.) were screened out as DNA binding agents. Most of these compounds were identified by standards and LC-MS analysis after the sample pretreatment with the DNA immobilized cartridge. DNA structural binding preference of the multiple active compounds in these two extracts was then evaluated simultaneously without purification.     

Novel cytotoxic oxovanadium(IV) complexes: Influence of pyrazole‐incorporated heterocyclic scaffolds on their biological response

Homoleptic VO^IV complexes with bidentate heterocycles, namely bipyrazole/pyrimidin‐2‐amine/triazolopyrimidine derivatives, are novel square pyramidal oxovanadium(IV) complexes. Their identities have been established using physicochemical techniques, namely elemental and spectral (ESI‐MS, IR, UV–visible) analyses. The in vitro antibacterial activities against Gram‐negative and Gram‐positive microorganisms have been studied for all compounds, exhibiting good inhibition as compared to the ligands. In addition, all compounds exhibit significant cytotoxicity towards brine shrimp with LD₅₀ values obtained in the range 8–24 μg ml⁻¹. Cellular level cytotoxicity has been investigated using bioassay of Schizosaccharomyces pombe , the complexes showing inevitable viability as concentration increases. Notably, UV absorption spectral titrations of the synthesized complexes with DNA reveal that the complexes bind to calf thymus DNA (CT‐DNA) through intercalation mode (K _b = 10⁴–10⁵ M⁻¹). Molecular docking reveals that the compounds are stacked between the base pairs of DNA. The results suggest that the complexes show promising binding affinity compared to the ligands towards CT‐DNA.     

Probing of effective pyrazolone based metallonucleases: Molecular docking and in vitro biological critiques

Four novel tryptophan based metal (II) complexes of the type [ML (Try)₂] were prepared by using pyrazolone derived Schiff base ligand. The proposed structure was confirmed by physicochemical methods which reveal octahedral coordination environment around the metal center. Intercalative binding mode of the complexes with CT DNA was confirmed by electronic absorption titration, viscosity measurements and fluorescence spectroscopy. Efficiency of DNA cleavage ability of the metal complexes was explored by the gel electrophoresis technique. The antimicrobial activities of the metal complexes showed potent biocidal activity. The percentage of free radical scavenging activity shows that the complexes are very reactive towards DPPH. Moreover, their cytotoxicity was tested against the two cancer cell lines (MCF‐7 and HepG2) and one normal cell line (NHDF) respectively. The MTT assay shows that the complexes have the anticancer efficacy. Moreover, the complexes exhibit a limited cytotoxicity effect on normal cell line NHDF. The morphological changes of apoptosis cell death were investigated by using Hoechst 33258 staining method. In addition, the Molecular docking studies was executed to consider the nature of binding and binding affinity of the synthesized compounds with DNA (PDB: 1BNA) and protein (PDB: 3hb5).     

Water‐soluble Schiff base Cu(II) and Zn(II) complexes: Synthesis,DNA targeting ability and chemotherapeutic potential of Cu(II) complex for hepatocellular carcinoma – in vitro and in vivo approach

Reliable compounds with low toxicity are tempting potential chemotherapeutics. With an aim of achieving less toxic but more potent metallodrugs, four new‐generation hydrophilic Cu(II) and Zn(II) complexes with DNA‐targeting properties were synthesized and characterized using various physicochemical data. The excellent DNA binding and cleavage results confirmed the mode of binding of DNA with the complexes and their ability to denature it. The profound in vitro cytotoxicity exhibited by complex 3 against a panel of cell lines (HeLa, MCF‐7 and HepG‐2) along with NHDF (normal human dermal fibroblasts) with distinct activity towards HepG‐2 and low toxicity to NHDF prompted in vivo studies of induced hepatocellular carcinoma‐affected Swiss albino rats. On evaluating various serum hepatic, biological and histopathological parameters, complex 3 showed excellent activity in restoring the damaged liver to normal. As a means of identifying the pathway of DNA damage, flow cytometric evaluation of cell cycle analysis was performed, which revealed S phase arrest‐induced apoptosis in HepG‐2 cells by complex 3 , making it a cell cycle‐specific drug.     

C-2 Arylamino substituted purine ara-carbocyclic nucleosides as potential anti-cytomegalovirus agents

There are reports in the literature that placement of an arylamino side chain at the C-2 position of purine nucleosides produces compounds capable of inhibiting DNA polymerase. To evaluate the potential of this class of compounds as antiviral agents that act by inhibiting viral DNA polymerase, ara-carbocyclic purine nucleosides possessing a 4-(l-butyl)phenylamino and a 3,5-dichlorophenylamino substituent at C-2 were chosen as the prototype structures and have been prepared from 2,4,6-trichloropyrimidine in 6 steps. For the antiviral analysis, human cytomegalovirus served as the principal virus since it expresses a virally specific DNA polymerase. None of the compounds showed activity towards this virus, but they were found to display some toxicity towards one or more cell lines.     

2'-Spiro ribo- and arabinonucleosides: synthesis, molecular modelling and incorporation into oligodeoxynucleotides

We have synthesized four conformationally restricted bicyclic 2'-spiro nucleosides via 2'-C-allyl nucleosides as key intermediates. The ribo-configured 2'-spironucleosides 9b and 14b were obtained by a convergent strategy starting from 2-ketofuranose 1 whereas the arabino-configured 2'-spironucleosides 21 and 27 were obtained by a linear strategy with a 2'-ketouridine derivative as starting material. The furanose ring of 9b/14b adopts N-type conformations whereas the furanose ring of 21/27 exists as an N<==>S equilibrium. These compounds showed no anti-HIV-1 activity or cytotoxicity. Incorporation of the four 2'-spironucleosides (as monomers X4 and X5) into oligodeoxynucleotides was accomplished using the phosphoramidite approach on an automated DNA synthesizer. Irrespective of monomeric configuration, hybridization studies revealed that these 2'-spironucleotide monomers (X4 and X5) induce decreased duplex thermostabilities compared with the corresponding DNA:DNA and DNA:RNA duplexes. Molecular modelling indicated that steric constraints are a possible reason for the lowered binding affinities of the modified oligodeoxynucleotides towards complementary single-stranded DNA and single-stranded RNA complements.     

Effects of the nature of counterions in solution on interactions of phenoxazones, xanthones, and carbazoles containing benzo-crown groups with DNA

The interactions of DNA with phenoxazones, xanthones, and carbazoles containing the (benzo-18-crown-6)-4′-yl and (benzo-15-crown-5)-4′-yl radicals bonded to the chromophore via spacers of different lengths in the presence of Na⁺ and K⁺ ions were studied by spectrophotometry, circular dichroism, and dynamic birefringence. The thermodynamic parameters of the binding of the compounds with DNA and changes in the macromolecular parameters of the DNA molecule during complexation were determined. Based on the results of these studies, we suggested the models of bonding of these compounds to the double helix of DNA. It is shown that the mode of DNA binding with a phenoxazone derivative containing two (benzo-15-crown-5)-4′-yl radicals bonded via a fragment of glycine to chromophore depends on the type of the counterion in solution. In the presence of Na⁺, the chromophore is intercalated into the double helix of DNA; in the presence of K⁺, it is bound to DNA in the form of a dimer outside the double helix. The type of the counterion does not affect the mode of binding of other crown-containing compounds of actinocin with DNA. For compounds containing the (benzo-18-crown-6)-4′-yl radical, the mode of binding to DNA adepends only on the spacer length.     

New transition metal complexes of 9,10‐phenanthrenequinone p‐toluyl hydrazone Schiff base: Synthesis,spectroscopy, DNA and HSA interactions,antimicrobial, DFT and docking studies

The new complexes of Cu (II) and Ni (II) of a tridentate Schiff base ligand derived from 9,10‐phenanthrenequinone and p‐toluic hydrazide have been synthesized and characterized by elemental analysis, electrical conductometry, FT‐IR, Mass, NMR and UV–Vis. The DFT calculations were carried out at B3LYP/6‐31G*(d) level for the determination of the optimized structure of the ligand and its complexes. The as‐synthesized compounds were screened for their antimicrobial activity. Also, their binding behavior with fish salmon‐DNA (FS‐DNA) and human serum albumin (HSA) were studied by different kinds of spectroscopic and molecular modeling techniques. The fluorescence data at different temperatures were applied in order to estimate the thermodynamics parameters of interactions of ligand and its complexes with DNA and HSA. The results showed that the as‐made compounds could bind to FS‐DNA and HSA via the groove binding as the major binding mode. According to molecular docking calculation and competitive binding experiments, these compounds bind to the minor groove of DNA and hydrophobic residues located in the subdomain IB of HSA. In addition, the molecular docking results kept in good consistence with experimental data.     

Nucleotide oxidation mediated by naphthalimide excited states with covalently attached viologen cosensitizers

The ground- and excited-state interactions of polymethylene-linked 1,8-naphthalimide-viologen dyads with calf-thymus DNA have been investigated. By virtue of the covalently attached viologen, the compounds represent the first example of linked chromophore/cosensitizer systems in the photooxidation of duplex DNA. The compounds associate strongly with DNA. Analysis of ground-state spectral changes yield binding constants of 0.7-2.5 x 10(6) M-1. Upon 355 nm pulsed irradiation of the compounds in the presence of calf-thymus DNA, reduced viologen is observed within the laser pulse. Photoproducts are not observed on this time scale in the absence of DNA. Since ground-state bleaching of the naphthalimide was not observed, the results suggest that DNA nucleobases are the species being oxidized. The quantum efficiency of radical production increases with the extent of binding to DNA. Under conditions where the compounds are bound predominantly to DNA, the quantum efficiencies were found to range from 0.02 to 0.03. Although small, the values represent a substantial increase in charge-separation yield compared to 1,8-naphthalimide compounds that lack the covalently attached viologen. The mechanism of radical production and effect of number of intervening methylenes are discussed.     

Screening and investigation of triplex DNA binders from Stephania tetrandra S. Moore by a combination of peak area‐fading ultra high‐performance liquid chromatography with orbitrap mass spectrometry and optical spectroscopies

The identification and screening of triplex DNA binders are important because these compounds, in many cases, are potential anticancer agents as well as promising drug candidates. Therefore, the ability to screen for these compounds in a high‐throughput mode could dramatically improve the drug screening process. A method involving a combination of 96‐well plate format and peak area‐fading ultra high‐performance liquid chromatography coupled with Orbitrap mass spectrometry was employed for screening bioactive compounds binding to the triplex DNA from the extracts of Stephania tetrandra S. Moore. Two compounds were screened out and identified as fangchinoline and tetrandrine based on the comparison of retention time and tandem mass spectrometry data with those of standards. The binding mechanisms of fangchinoline and tetrandrine at the molecular level were explored using tandem mass spectrometry, fluorescence spectroscopy, ultraviolet‐visible spectroscopy, and circular dichroism. Collision‐induced dissociation experiments showed that the complexes with fangchinoline and tetrandrine were dissociated by ligand elimination. According to these measurements, an intercalating binding is the most appropriate binding mode of these two alkaloids to the triplex DNA. The current work provides not only deep insight into alkaloid‐triplex DNA complexes but also useful guidelines for the design of efficient anticancer agents.     

Ligational behavior of new mononucleating NOO ethyl pyruvate Schiff base towards 3d metal(II) ions: an emphasis on antiproliferative and photocleavage property

A series of Co(II), Ni(II), Cu(II), and Zn(II) complexes of a tridentate hydrazone were prepared and characterized by various spectro‐analytical techniques and magnetic moment studies. The complexes were found to be monomeric and non‐electrolytes. The copper complex is electrochemically active in the applied potential range. The compounds synthesized in the present study have shown promising antiproliferative activity when screened using the in vitro method against two human cancer cell lines: HeLa and HepG2. The Escherichia coli DNA‐binding properties of all the compounds were investigated with UV–visible absorption spectrophotometric titrations, viscosity measurements, DNA melting experiments and gel electrophoreses measurements. The compounds were demonstrated to act as DNA intercalators with appreciable DNA‐binding constant values. Copyright © 2012 John Wiley & Sons, Ltd.     

Drug screening of pharmaceutical discovery compounds by micro-size exclusion chromatography/mass spectrometry.

Micro-size exclusion chromatography coupled with capillary liquid chromatography (capLC) and mass spectrometry (MS) provides a rapid and simple approach to the preliminary screening of active ligands toward a specific target macromolecule. In this study, the effectiveness of this technique is demonstrated by a number of small molecule ligands with known binding affinities towards the protein target. All ligands were incubated together with a target protein under native conditions. Separation was then achieved by microcentrifugation where the high molecular weight (MW) compounds were selectively passed through the size-exclusion material. The retained low MW compounds were then recovered and analyzed by capLC/MS. The absence of the ligand indicated strong affinity towards the target, while ligand detection indicated inactivity. This assay demonstrated the drugs that were acting as strong inhibitors of Co-PDF from those showing to be comparatively inactive. The relative binding rank order of the drugs towards Co-PDF was also determined. The results were validated by a corresponding set of control experiments in which the target molecules were excluded from the process. In principle, high-throughput micro-size exclusion chromatography, coupled with capLC/MS, offers a powerful technique as a preliminary screen in determining both the strong binding affinity and the relative affinity rank ordering of ligands towards a specific target macromolecule, and is complementary with other analytical drug screening techniques.