An organometallic compound which exhibits a DNA topology-dependent one-stranded intercalation mode

Understanding how small molecules interact with DNA is essential since it underlies a multitude of pathological conditions and therapeutic interventions. Many different intercalator compounds have been studied because of their activity as mutagens or drugs, but little is known regarding their interaction with nucleosomes, the protein-packaged form of DNA in cells. Here, using crystallographic methods and molecular dynamics simulations, we discovered that adducts formed by [(η⁶-THA)Ru(ethylenediamine)Cl][PF₆] (THA = 5,8,9,10-tetrahydroanthracene; RAED-THA-Cl[PF₆]) in the nucleosome comprise a novel one-stranded intercalation and DNA distortion mode. Conversely, the THA group in fact remains solvent exposed and does not disrupt base stacking in RAED-THA adducts on B-form DNA. This newly observed DNA binding mode and topology dependence may actually be prevalent and should be considered when studying covalently binding intercalating compounds

Nucleosome acidic patch-targeting binuclear ruthenium compounds induce aberrant chromatin condensation

© 2017 The Author(s). The 'acidic patch' is a highly electronegative cleft on the histone H2A-H2B dimer in the nucleosome. It is a fundamental motif for protein binding and chromatin dynamics, but the cellular impact of targeting this potentially therapeutic site with exogenous molecules remains unclear. Here, we characterize a family of binuclear ruthenium compounds that selectively target the nucleosome acidic patch, generating intra-nucleosomal H2A-H2B cross-links as well as inter-nucleosomal cross-links. In contrast to cisplatin or the progenitor RAPTA-C anticancer drugs, the binuclear agents neither arrest specific cell cycle phases nor elicit DNA damage response, but rather induce an irreversible, anomalous state of condensed chromatin that ultimately results in apoptosis. In vitro, the compounds induce misfolding of chromatin fibre and block the binding of the regulator of chromatin condensation 1 (RCC1) acidic patch-binding protein. This family of chromatin-modifying molecules has potential for applications in drug development and as tools for chromatin research

Fighting cancer with transition metal complexes: from naked DNA to protein and chromatin targeting strategies

Many transition metal complexes have unique physicochemical properties that can be efficiently exploited in medicinal chemistry for cancer treatment. Traditionally, double-stranded DNA has been assumed to be the main binding target; however, recent studies have shown that nucleosomal DNA as well as proteins can act as dominant molecular binding partners. This has raised new questions about the molecular determinants that govern DNA versus protein binding selectivity, and has offered new ways to rationalize their biological activity and possible side effects. To address these questions, molecular simulations at an atomistic level of detail have been used to complement, support, and rationalize experimental data. Herein we review some relevant studies\u2014focused on platinum and ruthenium compounds\u2014to illustrate the power of state-of-the-art molecular simulation techniques and to demonstrate how the interplay between molecular simulations and experiments can make important contributions to elucidating the target preferences of some promising transition metal anticancer agents. This contribution aims at providing relevant information that may help in the rational design of novel drug-discovery strategies

Pseudo electron-deficient organometallics: limited reactivity towards electron-donating ligands

YesHalf-sandwich metal complexes are of considerable interest in medicine, material, and nanomaterial chemistry. The design of libraries of such complexes with particular reactivity and properties is therefore a major quest. Here, we report the unique and peculiar reactivity of eight apparently 16-electron half-sandwich metal (ruthenium, osmium, rhodium, and iridium) complexes based on benzene-1,2-dithiolato and 3,6-dichlorobenzene-1,2-dithiolato chelating ligands. These electron-deficient complexes do not react with electron-donor pyridine derivatives, even with the strong σ-donor 4-dimethylaminopyridine (DMAP) ligand. The Ru, Rh, and Ir complexes accept electrons from the triphenylphosphine ligand (σ-donor, π-acceptor), whilst the Os complexes were found to be the first examples of non-electron-acceptor electron-deficient metal complexes. We rationalized these unique properties by a combination of experimental techniques and DFT/TDFT calculations. The synthetic versatility offered by this family of complexes, the low reactivity at the metal center, and the facile functionalization of the non-innocent benzene ligands is expected to allow the synthesis of libraries of pseudo electron-deficient half-sandwich complexes with unusual properties for a large range of applications

Fighting Cancer with Transition Metal Complexes: From Naked DNA to Protein and Chromatin Targeting Strategies

Many transition metal complexes have unique physicochemical properties that can be efficiently exploited in medicinal chemistry for cancer treatment. Traditionally, double-stranded DNA has been assumed to be the main binding target; however, recent studies have shown that nucleosomal DNA as well as proteins can act as dominant molecular binding partners. This has raised new questions about the molecular determinants that govern DNA versus protein binding selectivity, and has offered new ways to rationalize their biological activity and possible side effects. To address these questions, molecular simulations at an atomistic level of detail have been used to complement, support, and rationalize experimental data. Herein we review some relevant studies—focused on platinum and ruthenium compounds—to illustrate the power of state-of-the-art molecular simulation techniques and to demonstrate how the interplay between molecular simulations and experiments can make important contributions to elucidating the target preferences of some promising transition metal anticancer agents. This contribution aims at providing relevant information that may help in the rational design of novel drug-discovery strategies

Characterizing the nucleosome site selectivity, adduct structures and impact on chromatin for a variety of metal-based anticancer compounds and structural studies of nucleosome-linker histone interactions

The enormous length of DNA molecules is managed in the nucleus of eukaryotic cells by wrapping the double helix through histone protein association to form chromatin. The basic repeating unit of chromatin is the nucleosome. Due to this packaging, nucleosomal DNA and histones have different conformations and accessibility compared to that of naked DNA or chaperone-bound histones. The structure and dynamics of chromatin are important for many cellular functions. Different nuclear factors bind to nucleosomes to regulate chromatin dynamics and signal downstream cellular processes, like DNA replication and transcription. As a consequence, nucleosomes and their binding partners can be specifically targeted to modulate these functions. This is especially relevant to cancer cell function, whereby numerous epigenetic differences, such as alterations in nucleosome positioning, between transformed and healthy cells result from extensive changes in gene expression and the frequency of DNA replication (cell division). Distinguishing epigenetic features of cancer cells could be potentially exploited through the design of site-specific chromatin-targeting anticancer agents. Because of the severe side effects and resistance problems associated with platinum anticancer drugs, many different compounds based on alternative heavy metals have been explored as potential therapeutic agents and many more are presently under investigation. However, the cellular targets and mechanisms of action for these newer agents are largely unknown. Based on studies with cisplatin and other platinum drugs, DNA is generally assumed to be the cellular target for reactive metal-based compounds. However, this thesis work and other recent evidence show that proteins are the main targets for at least some of these compounds. Since the nucleosome contains both DNA and protein components and is an important therapeutic target, it is useful for investigating the DNA- versus protein-binding preferences of different metal-based agents. We investigated the site selectivity, adduct structures and impact on chromatin for many different metal-based compounds, including ruthenium, osmium, gold and rhodium agents. This involved a broad spectrum of biochemical, X-ray crystallographic and electron microscopic (EM) techniques in conjunction with collaborations on cellular, analytical chemistry and computational approaches. The striking discovery is that, with the exception of one class of ruthenium agent, the compounds investigated show a preference for binding to histone protein sites as opposed to the DNA of the nucleosome. Moreover, by comparing two structurally similar ruthenium compounds, one a cytotoxic anti-primary tumor agent and the other an effectively non-cytotoxic antimetastasis agent, we could show the basis for their differential DNA versus histone protein targeting and consequently linkages to their distinct cellular impact. For the compounds that preferentially form adducts at histone protein sites, we also discovered that there are two distinct classes of binding site, which are distinguishable by virtue of electrostatic components, affinity for histidine, steric access and hydrophobic interactions. Beyond this, the results shed light on the basis for cleavage of ‘carrier’ ligands, the influence of different metal centers and the basis for a novel nucleosomal drug-drug synergy. Finally, we show that adducts formed by certain metalloagents can have a direct and dramatic influence on the structure and dynamics of the chromatin fiber. Together with the other findings, this suggests strong therapeutic potential surrounding many different features of chromatin, yielding new leads for the rational design of improved metal-based anticancer agents. By compacting chromatin in a site specific manner, linker histone association with the nucleosome changes the structure and accessibility of defined genomic sites. However, the details of linker histone binding location on the nucleosome are still not clear, and in particular a high-resolution atomic structure for a nucleosome-linker histone assembly has not been solved. In addition to the biological significance, structural characterization of linker histone-nucleosome interactions could hold importance for drug development initiatives, for instance new targets for metal-based compounds. In this thesis, we investigated binding and crystallization of complexes of nucleosome with linker histones, focusing largely on the minimal sized assembly, the ‘chromatosome’. We found different affinities for nucleosome binding to the full length versus globular domain-only linker histone and also some degree of DNA length and sequence dependence in the interaction. Different crystals of the chromatosome were obtained, however the best so far diffract X-rays to only ~5.5 Å resolution. Nonetheless, the initial results of the study provide a platform for further optimization of chromatosome crystallizations and possible structural characterization.DOCTOR OF PHILOSOPHY (SBS

Near-atomic resolution structures of interdigitated nucleosome fibres

Chromosome structure at the multi-nucleosomal level has remained ambiguous in spite of its central role in epigenetic regulation and genome dynamics. Recent investigations of chromatin architecture portray diverse modes of interaction within and between nucleosome chains, but how this is realized at the atomic level is unclear. Here we present near-atomic resolution crystal structures of nucleosome fibres that assemble from cohesive-ended dinucleosomes with and without linker histone. As opposed to adopting folded helical ‘30 nm’ structures, the fibres instead assume open zigzag conformations that are interdigitated with one another. Zigzag conformations obviate extreme bending of the linker DNA, while linker DNA size (nucleosome repeat length) dictates fibre configuration and thus fibre–fibre packing, which is supported by variable linker histone binding. This suggests that nucleosome chains have a predisposition to interdigitate with specific characteristics under condensing conditions, which rationalizes observations of local chromosome architecture and the general heterogeneity of chromatin structure.Ministry of Education (MOE)Published versionThis project was funded by the Singapore Ministry of Education Academic Research Fund Tier 1 (grants 2014-T1-001-049 and 2017-T1-002-020), Tier 2 (grant MOE2015-T2-2- 089) and Tier 3 (grant MOE2012-T3-1-001) Programmes. The research leading to these results has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement #730872, project CALIPSOplus. C.A.D. dedicates this work to the memory of Agnieszka Marie Mordas Martin