Cytotoxicity of guanine-based degradation products contributes to the antiproliferative activity of guanine-rich oligonucleotides

Guanine-rich oligonucleotides (GROs) have attracted considerable attention as anticancer agents, because they exhibit cancer-selective antiproliferative activity and can form G-quadruplex structures with higher nuclease resistance and cellular uptake. Recently, a GRO, AS1411 has reached phase II clinical trials for acute myeloid leukemia and renal cell carcinoma. The antiproliferative activity of GROs has been associated with various protein targets; however the real mechanisms of action remain unclear. In this study, we showed evidence that antiproliferative activity of GROs (including AS1411) is mainly contributed by the cytotoxicity of their guanine-based degradation products, such as monophosphate deoxyguanosine (dGMP), deoxyguanosine (dG) and guanine. The GROs with lower nuclease resistance exhibited higher antiproliferative activity. Among nucleotides, nucleosides and nucleobases, only guanine-based compounds showed highly concentration-dependent cytotoxicity. Our results suggest that it is necessary to reconsider the cancer-selective antiproliferative activity of GROs. Since guanine-based compounds are endogenous substances in living organisms, systematic studies of the cytotoxicity of these compounds will provide new information for the understanding of certain diseases and offer useful information for drug design.     

Calcium ions tune the zinc-sequestering properties and antimicrobial activity of human S100A12

Human S100A12 is a host-defense protein expressed and released by neutrophils that contributes to innate immunity. Apo S100A12 is a 21 kDa antiparallel homodimer that harbors two Ca(ii)-binding EF-hand domains per subunit and exhibits two His₃Asp motifs for chelating transition metal ions at the homodimer interface. In this work, we present results from metal-binding studies and microbiology assays designed to ascertain whether Ca(ii) ions modulate the Zn(ii)-binding properties of S100A12 and further evaluate the antimicrobial properties of this protein. Our metal-depletion studies reveal that Ca(ii) ions enhance the ability of S100A12 to sequester Zn(ii) from microbial growth media. We report that human S100A12 has antifungal activity against Candida albicans, C. krusei, C. glabrata and C. tropicalis, all of which cause human disease. This antifungal activity is Ca(ii)-dependent and requires the His₃Asp metal-binding sites. We expand upon prior studies of the antibacterial activity of S100A12 and report Ca(ii)-dependent and strain-selective behavior. S100A12 exhibits in vitro growth inhibitory activity against Listeria monocytogenes. In contrast, S100A12 has negligible effect on the growth of Escherichia coli K-12 and Pseudomonas aeruginosa PAO1. Loss of functional ZnuABC, a high-affinity Zn(ii) import system, increases the susceptibility of E. coli and P. aeruginosa to S100A12, indicating that S100A12 deprives these mutant strains of Zn(ii). To evaluate the Zn(ii)-binding sites of S100A12 in solution, we present studies using Co(ii) as a spectroscopic probe and chromophoric small-molecule chelators in Zn(ii) competition titrations. We confirm that S100A12 binds Zn(ii) with a 2 : 1 stoichiometry, and our data indicate sub-nanomolar affinity binding. Taken together, these data support a model whereby S100A12 uses Ca(ii) ions to tune its Zn(ii)-chelating properties and antimicrobial activity.     

Unified total synthesis of the natural products endiandric acid A,kingianic acid E,and kingianins A,D, and F

A measure of the strength of a synthetic strategy is its versatility: specifically, whether it allows structurally distinct targets to be prepared. Herein we disclose a unified approach for the total synthesis of natural products of three distinct structural types, all of which occur naturally as racemic mixtures. The point of divergence involves the terminal alkylation of a conjugated tetrayne, and culminates in a significantly shortened synthesis of endiandric acid A (8 steps), the first total synthesis of kingianic acid E (8 steps), and a second-generation synthesis of kingianins A, D, and F (11 steps). Evidence for redox catalysis in the biosynthesis of kingianic acid E is presented.     

Controlling the activity of quorum sensing autoinducers with light

Bacteria use a communication system, called quorum sensing (QS), to organize into communities and synchronize gene expression to promote virulence and secure survival. Here we report on a proof-of-principle for externally interfering with this bacterial communication system, using light. By employing photoswitchable small molecules, we were able to photocontrol the QS-related bioluminescence in an Escherichia coli reporter strain, and the expression of target QS genes and pyocyanin production in Pseudomonas aeruginosa.     

Synthesis and structural analysis of nonstoichiometric ternary fulleride K 1.5 Ba 0.25 CsC 60

The existence of cation-vacancy sites in fullerides might lead to long-range ordering and generate a new vacancy-ordered superstructure. The purpose of this work is to search whether or not long-range ordering of vacant tetrahedral sites, namely superstructure emerges in nonstoichiometric K _1.5 Ba _0.25 CsC ₆₀ fulleride. Therefore, K _1.5 Ba _0.25 CsC ₆₀ with cation-vacancy sites is synthesized using a precursor method to avoid inadequate stoichiometry control and formation of impurity phases within the target composition. For this purpose, first, phase-pure K ₆ C ₆₀ , Ba ₆ C ₆₀ and Cs ₆ C ₆₀ precursors are synthesized. Stoichiometric quantities of these precursors are used for further reaction with C ₆₀ to afford K _1.5 Ba _0.25 CsC ₆₀ . Rietveld analysis of the high-resolution synchrotron X-ray powder diffraction data of the precursors and K _1.5 Ba _0.25 CsC ₆₀ confirms that K ₆ C ₆₀ , Ba ₆ C ₆₀ and Cs ₆ C ₆₀ are single-phase and they crystallize in a body-centered-cubic structure ( Im 3) as reported in the literature. The analysis also shows that K _1.5 Ba _0.25 CsC ₆₀ phase can be perfectly modeled using a face-centered cubic structure. No new peaks appear which could have implied the appearance of a superstructure. This suggests that there is no long-range ordered arrangement of vacant tetrahedral sites in K _1.5 Ba _0.25 CsC ₆₀ .     

A convergent total synthesis of ouabagenin

A convergent total synthesis of ouabagenin, an aglycon of cardenolide glycoside ouabain, was achieved by assembly of the AB-ring, D-ring and butenolide moieties. The multiply oxygenated cis-decalin structure of the AB-ring was constructed from (R)-perillaldehyde through the Diels–Alder reaction and sequential oxidations. The intermolecular acetal formation of the AB-ring and D-ring fragments, and combination of the intramolecular radical and aldol reactions, assembled the requisite steroidal skeleton in a stereoselective fashion. Finally, stereoselective installation of the C17-butenolide via the Stille coupling and hydrogenation led to ouabagenin.     

A soluble molecular variant of the semiconducting silicondiselenide

Silicondiselenide is a semiconductor and exists as an insoluble polymer (SiSe₂)_n which is prepared by reacting elemental silicon with selenium powder in the temperature range of 400–850 °C. Herein, we report on the synthesis, isolation, and characterization of carbene stabilized molecular silicondiselenide in the form of (cAAC)₂Si₂Se₄ (3) [cAAC = cyclic alkyl(amino)carbene]. 3 is synthesized via reaction of diatomic silicon(0) compound (cAAC)₂Si₂ (2) with black selenium powder at –78 °C to room temperature. The intensely orange colored compound 3 is soluble in polar organic solvents and stable at room temperature for a month under an inert atmosphere. 3 decomposes above 245 °C. The molecular structure of 3 has been confirmed by X-ray single crystal diffraction. It is also characterized by UV-vis, IR, Raman spectroscopy and mass spectrometry. The stability, bonding, and electron density distributions of 3 have been studied by theoretical calculations.     

In situ activation and monitoring of the evolution of the intracellular caspase family

The evolution of the intracellular caspase family is crucial in cell apoptosis. To evaluate this process, a universal platform of in situ activation and monitoring of the evolution of intracellular caspase is designed. Using well-known gold nanostructure as a model of both nanocarrier and matter inducing the cell apoptosis for photothermal therapy, a nanoprobe is prepared by assembly of two kinds of dye-labelled peptides specific to upstream caspase-9 and downstream caspase-3 as the signal switch, and folic acid as a targeting moiety. The energy transfer from dyes to the gold nanocarrier at two surface plasmon resonance absorption wavelengths leads to their fluorescence quenching. Upon endocytosis of the nanoprobe to perform the therapy against cancer cells, the peptides are successively cleaved by intracellular caspase activation with the evolution from upstream to downstream, which lights up the fluorescence of the dyes sequentially, and can be used to quantify both caspase-9 and caspase-3 activities in cancer cells and to monitor their evolution in living mice. The recovered fluorescence could also be used to assess therapeutic efficiency. This work provides a novel powerful tool for studying the evolution of the intracellular caspase family and elucidating the biological roles of caspases in cancer cell apoptosis.     

A supramolecular nanovehicle toward systematic,targeted cancer and tumor therapy

A supramolecular nanovehicle (denoted as SNV) was fabricated by encapsulating zinc phthalocyanine (ZnPc) and doxorubicin (DOX) into a copolymer (PVP-b-PAA-g-FA), so as to achieve systematic and synergistic chemotherapy-photodynamic therapy (PDT), targeted tumor imaging and therapy. The sophisticated copolymer designed in this work can load the PDT photosensitizer (ZnPc) and chemotherapy drug (DOX) simultaneously, which exhibits an excellent performance in chemotherapy-PDT targeted cancer and tumor therapy for both in vitro studies performed with HepG2 cells and in vivo tests with mice. This work provides a new drug formulation with a chemotherapy-PDT synergistic effect by virtue of the supramolecular material design, which possesses the advantages of an ultra-low drug dosage and highly-efficient in vivo targeted tumor imaging/therapy.     

Design,preparation, and selection of DNA-encoded dynamic libraries

We report a method for the preparation and selection of DNA-encoded dynamic libraries (DEDLs). The library is composed of two sets of DNA-linked small molecules that are under dynamic exchange through DNA hybridization. Addition of the protein target shifted the equilibrium, favouring the assembly of high affinity bivalent binders. Notably, we introduced a novel locking mechanism to stop the dynamic exchange and “freeze” the equilibrium, thereby enabling downstream hit isolation and decoding by PCR amplification and DNA sequencing. Our DEDL approach has circumvented the limitation of library size and realized the analysis and selection of large dynamic libraries. In addition, this method also eliminates the requirement for modified and immobilized target proteins.     

Addition of aluminium,zinc and magnesium hydrides to rhodium(iii)

We report the addition of M–H bonds (M = Al, Zn, Mg) to a Rh(iii) intermediate generated from the reductive elimination of triethylsilane from [Cp*Rh(H)₂(SiEt₃)₂]. A series of new heterobimetallic complexes possessing Rh–M bonds have been isolated and characterised by a number of spectroscopic (¹H, ²⁹Si, ¹³C, ¹⁰³Rh NMR, infrared, and X-ray diffraction) and computational techniques (NBO and QTAIM analysis). Experimental and computational data are consistent with cleavage of the M–H bond upon addition to rhodium with formation of new Rh–M and Rh–H bonds. Upon photolysis the Al analogue of this series undergoes a further elimination reaction producing triethylsilane and a highly unusual Rh₂Al₂H₄ containing cluster proposed to contain an Al(i) bridging ligand.     

Mechanism and selectivity of the dinuclear iron benzoyl-coenzyme A epoxidase BoxB

Benzoyl-CoA epoxidase is a dinuclear iron enzyme that catalyzes the epoxidation reaction of the aromatic ring of benzoyl-CoA with chemo-, regio- and stereo-selectivity. It has been suggested that this enzyme may also catalyze the deoxygenation reaction of epoxide, suggesting a unique bifunctionality among the diiron enzymes. We report a density functional theory study of this enzyme aimed at elucidating its mechanism and the various selectivities. The epoxidation is suggested to start with the binding of the O₂ molecule to the diferrous center to generate a diferric peroxide complex, followed by concerted O–O bond cleavage and epoxide formation. Two different pathways have been located, leading to (2S,3R)-epoxy and (2R,3S)-epoxy products, with barriers of 17.6 and 20.4 kcal mol^–1, respectively. The barrier difference is 2.8 kcal mol^–1, corresponding to a diastereomeric excess of about 99 : 1. Further isomerization from epoxide to phenol is found to have quite a high barrier, which cannot compete with the product release step. After product release into solution, fast epoxide–oxepin isomerization and racemization can take place easily, leading to a racemic mixture of (2S,3R) and (2R,3S) products. The deoxygenation of epoxide to regenerate benzoyl-CoA by a diferrous form of the enzyme proceeds via a stepwise mechanism. The C2–O bond cleavage happens first, coupled with one electron transfer from one iron center to the substrate, to form a radical intermediate, which is followed by the second C3–O bond cleavage. The first step is rate-limiting with a barrier of only 10.8 kcal mol^–1. Further experimental studies are encouraged to verify our results.     

Extending N-heterocyclic carbene ligands into the third dimension: a new type of hybrid phosphazane/NHC system

A simple, “click” synthetic approach to a new type of hybrid phosph(III)azane/NHC system is described. The presence of the phosphazane P₂N₂ ring unit, with P atoms flanking the NCN fragment and with this ring perpendicular to the binding site of the NHC, provides unique opportunities for modifying the electronic and steric character of these carbenes.     

A pH-activatable and aniline-substituted photosensitizer for near-infrared cancer theranostics

This work reports a newly designed pH-activatable and aniline-substituted aza-boron-dipyrromethene as a trifunctional photosensitizer to achieve highly selective tumor imaging, efficient photodynamic therapy (PDT) and therapeutic self-monitoring through encapsulation in a cRGD-functionalized nanomicelle. The diethylaminophenyl is introduced in to the structure for pH-activatable near-infrared fluorescence and singlet oxygen (¹O₂) generation, and bromophenyl is imported to increase the ¹O₂ generation efficiency upon pH activation by virtue of its heavy atom effect. After encapsulation, the nanoprobe can target α_vβ₃ integrin-rich tumor cells via cRGD and is activated by physiologically acidic pH for cancer discrimination and PDT. The fascinating advantage of the nanoprobe is near-infrared implementation beyond 800 nm, which significantly improves the imaging sensitivity and increases the penetration depth of the PDT. By monitoring the fluorescence decrease in the tumor region after PDT, the therapeutic efficacy is demonstrated in situ and in real time, which provides a valuable and convenient self-feedback function for PDT efficacy tracking. Therefore, this rationally designed and carefully engineered nanoprobe offers a new paradigm for precise tumor theranostics and may provide novel opportunities for future clinical cancer treatment.     

Synthesis,optical, and structural properties of bisphenol-bridged aromatic cyclic phosphazenes

Phenoxy- and naphthoxy-substituted bisphenol-bridged cyclic phosphazenes were synthesized in 2 steps and their thermal, photophysical, and electrochemical properties were investigated. The structures of the cyclic phosphazene compounds were determined by ESI-MS mass spectrometry and ¹ H, ¹³ C, and ³¹ P NMR spectroscopies. The photophysical studies of phenoxy- and naphthoxy-substituted bridged cyclophosphazenes were investigated by means of absorption and fluorescence spectroscopies in different solvents. Thermal and electrochemical properties of the target compounds were also studied. Furthermore, the excimer emissions through intramolecular interactions in solution and in solid state were investigated by fluorescence spectroscopy and the theoretical calculations were performed in detail using DFT.     

Heterologous expression of the avirulence gene ACE1 from the fungal rice pathogen Magnaporthe oryzae

The ACE1 and RAP1 genes from the avirulence signalling gene cluster of the rice blast fungus Magnaporthe oryzae were expressed in Aspergillus oryzae and M. oryzae itself. Expression of ACE1 alone produced a polyenyl pyrone (magnaporthepyrone), which is regioselectively epoxidised and hydrolysed to give different diols, 6 and 7, in the two host organisms. Analysis of the three introns present in ACE1 determined that A. oryzae does not process intron 2 correctly, while M. oryzae processes all introns correctly in both appressoria and mycelia. Co-expression of ACE1 and RAP1 in A. oryzae produced an amide 8 which is similar to the PKS-NRPS derived backbone of the cytochalasans. Biological testing on rice leaves showed that neither the diols 6 and 7, nor amide 8 was responsible for the observed ACE1 mediated avirulence, however, gene cluster analysis suggests that the true avirulence signalling compound may be a tyrosine-derived cytochalasan compound.     

Significant improvement of oxidase activity through the genetic incorporation of a redox-active unnatural amino acid

While nature employs various covalent and non-covalent strategies to modulate tyrosine (Y) redox potential and pK _a in order to optimize enzyme activities, such approaches have not been systematically applied for the design of functional metalloproteins. Through the genetic incorporation of 3-methoxytyrosine (OMeY) into myoglobin, we replicated important features of cytochrome c oxidase (CcO) in this small soluble protein, which exhibits selective O₂ reduction activity while generating a small amount of reactive oxygen species (ROS). These results demonstrate that the electron donating ability of a tyrosine residue in the active site is important for CcO function. Moreover, we elucidated the structural basis for the genetic incorporation of OMeY into proteins by solving the X-ray structure of OMeY specific aminoacyl-tRNA synthetase complexed with OMeY.     

Structure elucidation of nigricanoside A through enantioselective total synthesis

Nigricanoside A was isolated from green alga, and its dimethyl ester was found to display potent cytotoxicity. Its scarcity prevented a full structure elucidation, leaving total synthesis as the only means to determine its relative and absolute stereochemistry and to explore its biological activity. Here we assign the stereochemistry of the natural product through enantioselective total synthesis and provide initial studies of its cytotoxicity.     

A cell-penetrating protein designed for bimodal fluorescence and magnetic resonance imaging

Multimodal imaging is a highly desirable biomedical application since it can provide complementary information from each imaging modality. We propose a protein engineering-based strategy for the construction of a bimodal probe for fluorescence and magnetic resonance imaging. A recombinant protein was generated by the fusion of a supercharged green fluorescence protein (GFP³⁶⁺) with a lanthanide-binding tag (dLBT) that can stably bind two Gd³⁺ ions. The GFP³⁶⁺–dLBT fusion protein showed strong fluorescence and exhibited efficient contrast enhancement in magnetic resonance imaging. This protein probe improves the MR relaxation more efficiently than Gd-DTPA (gadopentetate dimeglumine). The superior cell-penetrating activity of GFP³⁶⁺ allows the efficient cellular uptake of this fusion protein and it can thus be used as a cellular imaging probe. Dual imaging was conducted in vitro and in mice. This result indicates that the fusion of different functional domains is a feasible approach for making multi-modal imaging agents.     

Synthesis and characterization of piperazine-substituted dihydrofuran derivatives viaMn(OAc) 3 mediated radical cyclizations

The aim of this study is to synthesize novel piperazine-containing dihydrofuran compounds (3a-n)from radical additions and cyclizations of diacyl and alkyl-acyl piperazine derivatives (1a-h) with 1,3-dicarbonyl compounds (2a-c) mediated by Mn(OAc) ₃ for the first time. From the reactions of 1a-c with dimedone (2a);1a, 1c, and 1d with acetylacetone (2b); and 1a with ethylacetoacetate(2c) ,the dihydrofuran-piperazine compounds 3a-c, 3d-f, and 3g were obtained in medium to high yields (31%–81%), respectively. In addition, dihydrofuran-piperazine compounds 3h-j and 3k-n were prepared at low to medium yields (20%–40%) from the reactions of 1e-g with 2a and 1e-h with 2c , respectively.