ReV Complexes Research Articles

Rhenium(V) Complexes as Cysteine-Targeting Coordinate Covalent Warheads.

Interest in covalent enzyme inhibitors as therapeutic agents has seen a recent resurgence. Covalent enzyme inhibitors typically possess an organic functional group that reacts with a key feature of the target enzyme, often a nucleophilic cysteine residue. Herein, the application of small, modular ReV complexes as inorganic cysteine-targeting warheads is described. These metal complexes were found to react with cysteine residues rapidly and selectively. To demonstrate the utility of these ReV complexes, their reactivity with SARS-CoV-2-associated cysteine proteases is presented, including the SARS-CoV-2 main protease and papain-like protease and human enzymes cathepsin B and L. As all of these proteins are cysteine proteases, these enzymes were found to be inhibited by the ReV complexes through the formation of adducts. These findings suggest that these ReV complexes could be used as a new class of warheads for targeting surface accessible cysteine residues in disease-relevant target proteins.

Oxidorhenium(V) and Rhenium(III) Complexes with m‐Terphenyl Isocyanides

Reactions of the sterically encumbered m‐terphenyl isocyanides CNArDipp2 (Dipp = 2,6‐diisopropylphenyl) and CNArMes2 (Mes = 2,4,6‐trimethylphenyl) with (NBu4)[ReOCl4] in CH2Cl2 form stable complexes of the composition (NBu4)[ReOCl3(CNArR)] or [ReOCl3(CNArR)2] depending on the amount of isocyanide added. In the [ReOCl3(CNArR)2] complexes, cis coordination of the two isocyanides is observed for CNArMes2, while the sterically more demanding CNArDIPP2 ligands are found in trans positions. The rhenium(III) species [ReCl3(PPh3)(CNArMes2)2] was obtained from the reaction of [ReOCl3(PPh3)2] and CNArMes2. The ν(CN) IR frequencies measured for the ReV complexes appear at higher wavenumbers than for the uncoordinated isocyanides, which suggests a low degree of backdonation into anti‐bonding orbitals of these ligands.

Bioavailable inhibitors of HIV-1 RNA biogenesis identified through a Rev-based screen

New antiretroviral agents with alternative mechanisms are needed to complement the combination therapies used to treat HIV-1 infections. Here we report the identification of bioavailable molecules that interfere with the gene expression processes of HIV-1. The compounds were detected by screening a small library of FDA-approved drugs with an assay based on measuring the displacement of Rev, and essential virus-encoded protein, from its high-affinity RNA binding site. The antiretroviral activity of two hits was based on interference with post-integration steps of the HIV-1 cycle. Both hits inhibited RRE–Rev complex formation in vitro, and blocked LTR-dependent gene expression and viral transcription in cellular assays. The best compound altered the splicing pattern of HIV-1 transcripts in a manner consistent with Rev inhibition. This mechanism of action is different from those used by current antiretroviral agents. The screening hits recognized the Rev binding site in the viral RNA, and the best compound did so with substantial selectivity, allowing the identification of a new RNA-binding scaffold. These results may be used for developing novel antiretroviral drugs.

Host Cell Interactome of HIV-1 Rev Includes RNA Helicases Involved in Multiple Facets of Virus Production

The HIV-1 Rev protein plays a key role in the late phase of virus replication. It binds to the Rev Response Element found in underspliced HIV mRNAs, and drives their nuclear export by the CRM1 receptor pathway. Moreover, mounting evidence suggests that Rev has additional functions in viral replication. Here we employed proteomics and statistical analysis to identify candidate host cell factors that interact with Rev. For this we studied Rev complexes assembled in vitro with nuclear or cytosolic extracts under conditions emulating various intracellular environments of Rev. We ranked the protein-protein interactions by combining several statistical features derived from pairwise comparison of conditions in which the abundance of the binding partners changed. As a validation set, we selected the eight DEAD/H box proteins of the RNA helicase family from the top-ranking 5% of the proteins. These proteins all associate with ectopically expressed Rev in immunoprecipitates of cultured cells. From gene knockdown approaches, our work in combination with previous studies indicates that six of the eight DEAD/H proteins are linked to HIV production in our cell model. In a more detailed analysis of infected cells where either DDX3X, DDX5, DDX17, or DDX21 was silenced, we observed distinctive phenotypes for multiple replication features, variously involving virus particle release, the levels of unspliced and spliced HIV mRNAs, and the nuclear and cytoplasmic concentrations of these transcripts. Altogether the work indicates that our top-scoring data set is enriched in Rev-interacting proteins relevant to HIV replication. Our more detailed analysis of several Rev-interacting DEAD proteins suggests a complex set of functions for the helicases in regulation of HIV mRNAs. The strategy used here for identifying Rev interaction partners should prove effective for analyzing other viral and cellular proteins.

Rhenium(IV) and rhenium(V) complexes with 3,5-dimethylpyrazole

Mono-and dinuclear ReIV and ReV complexes with 3,5-dimethylpyrazole (Me2pzH) were synthesized. The cis-[Re2O3Cl4(3,5-Me2pzH)4] complex (cis-1) was prepared by the reaction of NH4ReO4 with K[HB(Me2pz)3] in concentrated HCl or by refluxing of [ReCl3(MeCN)(PPh3)2] with Me2pzH in air. The bromide complex trans-[Re2O3Br4(3,5-Me2pzH)4] (trans-2) was synthesized by passing dry HBr through a solution of [Re2O3Br2(μ-3,5-Me2pz)2(3,5-Me2pzH)2] (4) in chloroform. The pyrazolate-bridged complex [Re2O3Cl2(μ-3,5-Me2pz)2(3,5-Me2pzH)2] (3) was prepared from (Et4N)2[ReOCl5] or Cs2[ReOCl5] and Me2pzH. The corresponding bromide and iodide complexes [Re2O3X2(3,5-Me2pz)2(3,5-Me2pzH)2] · C6H6 (X = Br (4) or I (5)) were synthesized by the reactions of (NH4)2[ReBr6] or K2[ReI6], respectively, with Me2pzH. The [ReO(OMe)(3,5-Me2pzH)4]Br2 · · 3,5-Me2pzH · 4H2O complex (6) was obtained as a by-product in the synthesis of complex 4. The reaction of [ReNCl2(PPh3)2] with Me2pzH was accompanied by hydrolytic denitration giving rise to the mixed-ligand complex [Re2O3Cl2(μ-3,5-Me2pz)2(3,5-Me2pzH)(PPh3)] (7). The reaction of (NH4)2[ReBr6] with a Me2pzH melt gave the trans-[ReBr4(3,5-Me2pzH)2] · · Me2CO complex (8). The structures of complexes 2 and 4–8 were established by X-ray diffraction. All compounds were characterized by elemental analysis, electronic absorption spectroscopy, 1H NMR and IR spectroscopy, mass spectrometry, and cyclic voltammetry.

Synthesis and Structural Studies of Rhenium(V) Complexes Stabilized by a Monoanionic Cyclen Ligand

AbstractThe novel cationic (oxo)ReV complexes [ReO(cyclen‐H)Cl]X [X = Cl (3a), BPh4 (3b)] and [ReO(cyclen‐H)I]I (4) have been prepared by treating cyclen (1,4,7,10‐tetrazacyclododecane) with (Bu4N)[ReOCl4] (1) and cis‐[ReO2I(PPh3)2] (2), respectively. The X‐ray structural analyses of 3a and 4, as well as 1H NMR spectroscopy, showed that the macrocyclic ligand is coordinated in the unprecedented monoanionic form. 1H and 13C NMR spectroscopy also demonstrated that 3a,b and 4 exist in solution as single isomers. To the best of our knowledge, 3a and 4 are the first examples of structurally characterized oxo complexes of d transition elements with cyclen bound to the metal atom through one amido and three amine groups. In both complexes, the geometry around the ReV center is best described as distorted octahedral. The terminal oxo and halo ligands are coordinated in relative cis positions, and the amido and three amino nitrogen atoms from cyclen occupy the four remaining coordination positions. The Re−N(amido) bonds [1.894(9) Å in 3a and 1.888(12) Å in 4] are trans to the halo ligands and, as expected, are considerably shorter than the Re−N(amine) bonds, which span from 2.110(6) to 2.246(8) Å and from 2.124(8) to 2.268(10) Å for cations the cations of 3a and 4, respectively. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)

New Mixed-Ligand ReV Complexes with Bis(2-mercaptoethyl) Sulfide and Functionalized Thioimidazolyl Ligands

The preparation of the mixed-ligand ReV complexes [Re(O)(κ3-SSS)(κ1-Simz)] (1), [Re(O)(κ3-SSS)(κ1-Simz)]·HCl (2) and [Re(O)(κ3-SSS)(κ1-SimzCOGlyGly)] (3) are described. These [3+1] type compounds are stabilized by tridentate bis(2-mercaptoethyl) sulfide (HSSSH) and by monodentate functionalized thioimidazolyl ligands. Complexes 1, 2 and 3 were fully characterized by IR and 1H NMR spectroscopy, and by X-ray diffraction analysis in the case of 1 and 2. In complexes 1 and 2 the Re atom is five-coordinate, presenting a distorted square pyramidal coordination geometry. Based on the angular structural parameter, τ, this distortion lies towards a trigonal bipyramidal geometry (τ = 0.40, 1; τ = 0.46, 2). (© Wiley-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002)

Rhenium complexes with pyrazoles. Synthesis and crystal structure of trans-[Re(O)(OMe)L4]Br2·L·4H2O (L is 3,5-dimethylpyrazole)

The first mononuclear ReV complex with four pyrazole ligands, viz., [Re(O)(OMe)L4]Br2 (L is 3,5-dimethylpyrazole), and its molecular adduct with L, viz., [Re(O)(OMe)L4]Br2·L·4H2O, were synthesized. The complex is resistant to hydrolysis in a neutral aqueous solution. The structure of the adduct was established by X-ray diffraction analysis.

Inhibition of rev·RRE complexation by triplex tethered oligonucleotide probes

We have described a class of molecules, called tethered oligonucleotide probes (TOPs), that bind RNA on the basis of both sequence and structure. TOPs consist of two short oligonucleotides joined by a tether whose length and composition may be varied using chemical synthesis. In a triplex TOP, one oligonucleotide recognizes a short single-stranded region in a target RNA through the formation of Watson-Crick base pairs; the other oligonucleotide recognizes a short double-stranded region through the formation of Hoogsteen base pairs. Binding of triplex TOPs to an HIV-1 Rev Response Element RNA variant (RRE AU) was measured by competition electrophoretic mobility shift analysis. Triplex TOP·RRE AU stabilities ranged between −9.6 and −6.1 kcal mol 1 under physiological conditions of pH, salt, and temperature. Although the most stable triplex TOP·RRE AU complex contained 12 contiguous U·AU triple helical base pairs, complexes containing only six or nine triple helical base pairs also formed. Triplex TOPs inhibited formation of the RRE·Rev complex with IC 50 values that paralleled the dissociation constants of the analogous triplex TOP·RRE AU complexes. In contrast to results obtained with TOPs that target two single-stranded RRE regions, inhibition of Rev·RRE AU complexation by triplex TOPs did not require pre-incubation of RRE AU and a TOP: triplex TOPs competed efficiently with Rev for RRE AU and inhibited RRE AU·Rev complexation at equilibrium.

Inhibition of HIV-1 replication in lymphocytes by mutants of the Rev cofactor eIF-5A.

Eukaryotic initiation factor 5A(eIF-5A) is a cellular cofactor require d for the function of the human immunodeficiency virus type-1 (HIV-1) Rev trans-activator protein. The majority of a set of eIF-5A mutants did not support growth of yeast cells having an inactivated genomic copy of eIF-5A, indicating that the introduced mutation eliminated eIF-5A activity. Two nonfunctional mutants, eIF-5AM13 and eIF-5AM14, retained their binding capacity for the HIV-1 Rev response element:Rev complex. Both mutants were constitutively expressed in human T cells. When these T cells were infected with replication-competent HIV-1, virus replication was inhibited. The eIF-5AM13 and eIF5AM14 proteins blocked Rev trans-activation and Rev-mediated nuclear export.

Inhibition of HIV-1 Rev-RRE interaction by diphenylfuran derivatives.

The interactions between RNA structures, such as RRE in the HIV-1 genome, and proteins, such as Rev of HIV-1, are essential for efficient viral replication. Compounds that bind specifically to such RNAs and disrupt their protein complexes offer a novel mechanism for inhibition of replication of the virus. As a step in this approach, we have designed and characterized a series of synthetic diphenylfuran cations that selectively inhibit Rev binding to RRE. Fluorescence titrations and gel band-shift results indicate that the diphenylfurans bind to RRE and inhibit Rev complex formation in a structure-dependent manner. The derivative with the greatest affinity for RRE has an association constant of greater than 10(7) M-1 and inhibits formation of the Rev--RRE complex at concentrations below 1 microM. It binds to RRE considerably more strongly than it binds to simple RNA duplexes. Spectral changes and energy transfer results on complex formation suggest that the compound has a nonclassical intercalation binding mode. CD studies with modified RRE hairpins indicate that the active diphenylfurans bind at the structured internal loop of RRE and cause a conformational change. The most active diphenylfurans are tetracations that appear to bind to RRE by a threading intercalation mode and cause a conformational change in the RNA that is essential for inhibition of Rev complex formation with RRE.

Multinuclear NMR Studies of Ligand‐Exchange Reactions on Analogous Technetium(V) and Rhenium(V) Complexes. Relevance to nuclear medicine

AbstractThe kinetics of pyridine exchange on trans‐[MO2(py)4]+ have been followed by 1H‐NMR in CD3NO2 for M = Re, Tc: k298S−1 = (5.5 ± 0.1) × 10−6, 0.04 ± 0.02; ΔH≠/kJmol−1 = 111 ± 3, 101 ± 9; ΔS≠/JK−1mol−1 = +28 ± 10, +68 ± 35. For the Rev complex, pyridine and oxygen exchanges have been measured simultaneously by 1H‐ and 17O‐NMR in deuterated water: k298/s−1 = (8.6 ± 0.2) × 10−6 (py), (14.5 ± 0.3) × 10−6 (oxygen); ΔH≠/kJmol−1 = 111 ± 1, 91 ± 1; ΔS /JK−1mol−1 = +32 ± 3, −32 ± 4. For both complexes, the rate law for pyridine exchange is first‐order in complex and zero‐order in pyridine; together with the activation parameter values, and the fact that the rate does not depend significantly on the nature of the solvent, this strongly implies the operation of a dissociative mechanism. The ratio of pyridine exchange rates for the Tc and Re complexes at room temperature is ca. 8000. The consequences of these observations for radiopharmaceutical synthesis are discussed.

Kinetic and equilibrium study of substitution reactions oftrans-tetracyanodioxorhenate(V) ions with monodentate nucleophiles

The kinetics of the substitution reactions of the protonated froms oftrans-tetracyanodioxorthenate(V) with thiourea (TU),N-methylthiourea (NMTU),N, N′-dimethylthiourea (NNDMTU) and hydrazoic acid (HN3) were studied. The results were compared with those obtained for similar reactions of [WO2(CN)4]4−. This study showed that the diprotonated form [ReO(H2O)(CN)4]− is the only species reactive towards substitution reactions (and not the [ReO(OH)(CN)4]2− ion) and that only the aqua ligand in [ReO(H2O)(CN)4]− is substituted by the incoming group. A dissociative mechanism is proposed for the substitution reactions between [ReO(H2O)(CN)4]− and the monodentate nucleophiles. The i.r. data for these Rev complexes are reported and discussed in terms of the relativetrans influence of the various monodentate ligands.

Alkyl and Alkylidyne Complexes of Rhenium

The multiplicity of its oxidation states enables the element rhenium to fix numerous ligand types in stable complexes. The synthesis of “Cp*ReClMe3” now completes the series of Rev complexes “Cp*ReClnMe4–n”. The neopentyl complex 1 reacts with C5H5TiCl3 to give the paramagnetic carbyne complex 2, which can be smoothly oxidized to the diamagnetic complex 3. Complexes 2 and 3 are the first π-cyclopentadienylcarbyne complexes with rhenium in high oxidation states (ReVI and ReVII, resp.). Cp* = η5-C5(CH3)5.

Concerning the Role of Organotin Compounds in Olefin Metathesis: Synthesis, Structure, and Lewis Acidity of [(η5‐C5Me5)CH3ReCl3

A severely flattened pyramid characterizes the structure and determines the reactivity of the Rev complex 1, which is formed upon reaction of tetramethyltin with [(η5-C5Me4Et)ReCl4]. Lewis bases can attack at the trans position to the π-bound ring system of 1 and its η5-C5Me5 analogue. This finding could contribute to an understanding of the role of Sn-cocatalytis for olefin metathesis.

Hexahydrido(pentamethylcyclopentadienyl)rhenium, [(C5Me5)ReH6

The novel organometallic polyhydride 2, formally a ReVII derivative, is surprisingly formed upon reaction of the ReV complex 1 with LiAlH4 followed by careful methanolysis. Complex 2 has a fluxional structure at room temperature; at lower temperatures, the complex has a rigid, pentagonal-bipyramidal geometry.