A Highly Sterically Encumbered Boron Lewis Acid Enabled by an Organotellurium-Based Ligand.

Lewis acidic boron compounds are ubiquitous in chemistry due to their numerous applications, yet tuning and optimizing their properties towards different purposes is still a challenging field of research. In this work, the boron-based Lewis acid B[OTeF3(C6F5)2]3 was synthesized by reaction of the teflate derivative HOTeF3(C6F5)2 with BCl3 or BCl3 ⋅ SMe2. This new compound presents a remarkably high thermal stability up to 300 °C, as well as one of the most sterically encumbered boron centres known in the literature. Theoretical and experimental methods revealed that B[OTeF3(C6F5)2]3 exhibits a comparable Lewis acidity to that of the well-known B(C6F5)3. The affinity of B[OTeF3(C6F5)2]3 towards pyridine was accessed by Isothermal Titration Calorimetry (ITC) and compared to that of B(OTeF5)3 and B(C6F5)3. The ligand-transfer reactivity of this new boron compound towards different fluorides was demonstrated by the formation of an anionic Au(III) complex and a hypervalent iodine(III) species.

Fluorinated Dialkyl Chloronium Salts: Synthesis and Reactivity for Fluoroalkylation and Hydride Abstraction.

A new concept for the synthesis of dialkyl chloronium cations [R‒Cl‒R]+ is described (R = CH3, CH2CF3), that allows the formation of fluorinated derivatives. By utilizing the xenonium salt [XeOTeF5][M(OTeF5)n] (M = Sb, n = 6; M = Al, n = 4) chlorine atoms of chloroalkanes or the deactivated chlorofluoroalkane CH2ClCF3 are oxidized and removed as ClOTeF5 leading to the isolation of the corresponding chloronium salt. Since the resulting highly electrophilic cation [Cl(CH2CF3)2]+ is able to alkylate weak nucleophiles, this compound can be utilized for the introduction of a fluorinated alkyl group to those. In addition, the fluorinated alkyl chloronium cation displays a high hydride ion affinity, enabling the activation of linear hydrocarbons by hydride abstraction even at low temperatures ultimately leading to the formation of branched carbocations.

N, N-dimethyltryptamine forms oxygenated metabolites via CYP2D6 - an in vitro investigation.

N, N-dimethyltryptamine (DMT) is a psychedelic compound that has shown potential in the treatment of depression. Aside from the primary role of monoamine oxidase A (MAO-A) in DMT metabolism, the metabolic pathways are poorly understood. Increasing this understanding is an essential aspect of ensuring safe and efficacious use of DMT.This work aimed to investigate the cytochrome 450 (CYP) mediated metabolism of DMT by incubating DMT with recombinant human CYP enzymes and human liver microsomes (HLM) followed by analysis using high-resolution mass spectrometry for metabolite identification.DMT was rapidly metabolised by CYP2D6, while stable with all other investigated CYP enzymes. The metabolism of DMT in HLM was reduced after inclusion of harmine and SKF-525A whereas quinidine did not affect the metabolic rate, likely due to MAO-A residues present in HLM. Analysis of the CYP2D6 incubates showed formation of mono-, di- and tri-oxygenated metabolites, likely as a result of hydroxylation on the indole core.More research is needed to investigate the role of this metabolic pathway in vivo and any pharmacological activity of the proposed metabolites. Our findings may impact on safety issues following intake of ayahuasca in slow CYP2D6 metabolizers or with concomitant use of CYP2D6 inhibitors.

Insights on the Lewis Superacid Al(OTeF5 )3 : Solvent Adducts, Characterization and Properties.

Preparation and characterization of the dimeric Lewis superacid [Al(OTeF5 )3 ]2 and various solvent adducts is presented. The latter range from thermally stable adducts to highly reactive, weakly bound species. DFT calculations on the ligand affinity of these Lewis acids were performed in order to rank their remaining Lewis acidity. An experimental proof of the Lewis acidity is provided by the reaction of solvent-adducts of Al(OTeF5 )3 with [PPh4 ][SbF6 ] and OPEt3 , respectively. Furthermore, their reactivity towards chloride and pentafluoroorthotellurate salts as well as (CH3 )3 SiCl and (CH3 )3 SiF is shown. This includes the formation of the dianion [Al(OTeF5 )5 ]2- .

Unravelling the Role of the Pentafluoroorthotellurate Group as a Ligand in Nickel Chemistry.

The pentafluoroorthotellurate group (teflate, OTeF5 ) is able to form species, for which only the fluoride analogues are known. Despite nickel fluorides being widely investigated, nickel teflates have remained elusive for decades. By reaction of [NiCl4 ]2- and neat ClOTeF5 , we have synthesized the homoleptic [Ni(OTeF5 )4 ]2- anion, which presents a distorted tetrahedral structure, unlike the polymeric [NiF4 ]2- . This high-spin complex has allowed the study of the electronic properties of the teflate group, which can be classified as a weak/medium-field ligand, and therefore behaves as the fluoride analogue also in ligand-field terms. The teflate ligands in [NEt4 ]2 [Ni(OTeF5 )4 ] are easily substituted, as shown by the formation of [Ni(NCMe)6 ][OTeF5 ]2 by dissolving it in acetonitrile. Nevertheless, careful reactions with other conventional ligands have enabled the crystallization of nickel teflate complexes with different coordination geometries, i.e. [NEt4 ]2 [trans-Ni(OEt2 )2 (OTeF5 )4 ] or [NEt4 ][Ni(bpyMe2 )(OTeF5 )3 ].

The Tris(pentafluorophenyl)methylium Cation: Isolation and Reactivity.

Herein, we present two different routes for the synthesis of the perfluorinated trityl cation, which allowed the handling of the free, uncoordinated species in organic solvents for the first time. The usage of the weakly coordinating anion [Al(OTeF5 )4 ]- and its derivatives allows the characterization of this species by NMR spectroscopy and most importantly by single-crystal X-ray diffraction. The high hydride ion affinity of the cation is shown by hydrogen abstraction from isobutane. Furthermore, cyclic voltammetry reveals its oxidative potential which is supported by the reaction with tris(4-bromophenyl)amine, giving rise to the formation of the ammoniumyl radical cation, also known as "magic blue".

Factors Affecting the Pharmacokinetics of Pyrazinamide and Its Metabolites in Patients Coinfected with HIV and Implications for Individualized Dosing.

Pyrazinamide is a first-line drug used in the treatment of tuberculosis. High exposure to pyrazinamide and its metabolites may result in hepatotoxicity, whereas low exposure to pyrazinamide has been correlated with treatment failure of first-line antitubercular therapy. The aim of this study was to describe the pharmacokinetics and metabolism of pyrazinamide in patients coinfected with tuberculosis and HIV. We further aimed to identify demographic and clinical factors which affect the pharmacokinetics of pyrazinamide and its metabolites in order to suggest individualized dosing regimens. Plasma concentrations of pyrazinamide, pyrazinoic acid, and 5-hydroxypyrazinamide from 63 Rwandan patients coinfected with tuberculosis and HIV were determined by liquid chromatography-tandem mass spectrometry followed by nonlinear mixed-effects modeling. Females had a close to 50% higher relative pyrazinamide bioavailability compared to males. The distribution volumes of pyrazinamide and both metabolites were lower in patients on concomitant efavirenz-based HIV therapy. Furthermore, there was a linear relationship between serum creatinine and oral clearance of pyrazinoic acid. Simulations indicated that increasing doses from 25 mg/kg of body weight to 35 mg/kg and 50 mg/kg in females and males, respectively, would result in adequate exposure with regard to suggested thresholds and increase probability of target attainment to >0.9 for a MIC of 25 mg/liter. Further, lowering the dose by 40% in patients with high serum creatinine would prevent accumulation of toxic metabolites. Individualized dosing is proposed to decrease variability in exposure to pyrazinamide and its metabolites. Reducing the variability in exposure may lower the risk of treatment failure and resistance development.

Epigenetic program and transcription factor circuitry of dendritic cell development.

Dendritic cells (DC) are professional antigen presenting cells that develop from hematopoietic stem cells through successive steps of lineage commitment and differentiation. Multipotent progenitors (MPP) are committed to DC restricted common DC progenitors (CDP), which differentiate into specific DC subsets, classical DC (cDC) and plasmacytoid DC (pDC). To determine epigenetic states and regulatory circuitries during DC differentiation, we measured consecutive changes of genome-wide gene expression, histone modification and transcription factor occupancy during the sequel MPP-CDP-cDC/pDC. Specific histone marks in CDP reveal a DC-primed epigenetic signature, which is maintained and reinforced during DC differentiation. Epigenetic marks and transcription factor PU.1 occupancy increasingly coincide upon DC differentiation. By integrating PU.1 occupancy and gene expression we devised a transcription factor regulatory circuitry for DC commitment and subset specification. The circuitry provides the transcription factor hierarchy that drives the sequel MPP-CDP-cDC/pDC, including Irf4, Irf8, Tcf4, Spib and Stat factors. The circuitry also includes feedback loops inferred for individual or multiple factors, which stabilize distinct stages of DC development and DC subsets. In summary, here we describe the basic regulatory circuitry of transcription factors that drives DC development.

Enzyme-substrate complex structures of CYP154C5 shed light on its mode of highly selective steroid hydroxylation.

CYP154C5 from Nocardia farcinica is a bacterial cytochrome P450 monooxygenase active on steroid molecules. The enzyme has recently been shown to exhibit exclusive regioselectivity and stereoselectivity in the conversion of various pregnans and androstans, yielding 16α-hydroxylated steroid products. This makes the enzyme an attractive candidate for industrial application in steroid hormone synthesis. Here, crystal structures of CYP154C5 in complex with four different steroid molecules were solved at resolutions of up to 1.9 Å. These are the first reported P450 structures from the CYP154 family in complex with a substrate. The active site of CYP154C5 forms a flattened hydrophobic channel with two opposing polar regions, perfectly resembling the size and polarity distribution of the steroids and thus resulting in highly specific steroid binding with Kd values in the range 10-100 nM. Key enzyme-substrate interactions were identified that accounted for the exclusive regioselectivity and stereoselectivity of the enzyme. Additionally, comparison of the four CYP154C5-steroid structures revealed distinct structural differences, explaining the observed variations in kinetic data obtained for this P450 with the steroids pregnenolone, dehydroepiandrosterone, progesterone, androstenedione, testosterone and nandrolone. This will facilitate the generation of variants with improved activity or altered selectivity in the future by means of protein engineering.

Effects of artemisinin antimalarials on Cytochrome P450 enzymes in vitro using recombinant enzymes and human liver microsomes: potential implications for combination therapies.

1. Cytochrome P450 enzyme system is the most important contributor to oxidative metabolism of drugs. Modification, and more specifically inhibition, of this system is an important determinant of several drug-drug interactions (DDIs). 2. Effects of the antimalarial agent artemisinin and its structural analogues, artemether, artesunate and dihydroartemisinin, on seven of the major human liver CYP isoforms (CYP1A2, 2A6, 2B6, 2C9, 2C19, 2D6 and 3A4) were evaluated using recombinant enzymes (fluorometric assay) and human liver microsomes (LC-MS/MS analysis). Inhibitory potency (IC50) and mechanisms of inhibition were evaluated using nonlinear regression analysis. In vitro-in vivo extrapolation using the [I]/Ki ratio was applied to predict the risk of DDI in vivo. 3. All compounds tested inhibited the enzymatic activity of CYPs, mostly through a mixed type of inhibition, with CYP1A2, 2B6, 2C19 and 3A4 being affected. A high risk of interaction in vivo was predicted if artemisinin is coadministrated with CYP1A2 or 2C19 substrates. 4. With respect to CYP1A2 inhibition in vivo by artemisinin compounds, our findings are in line with previously published data. However, reported risks of interaction may be overpredicted and should be interpreted with caution.

2-Alkylphosphino-1-boraadamantanes.

Boraadamantanes are a privileged class of caged group 13 compounds having a trigonal pyramidal (non-VSEPR) ground-state. The Lewis acid-base chemistry of this type of compound is underdeveloped when compared to acyclic derivatives. This report provides the first examples of bifunctional boraadamantanes with an appended phosphine moiety (also the first boraadamantanes having a phosphorus-boron bond). Using this scaffold, boraadamantane coordination compounds are accessed for the first time.

[Xe(OTeF5)(pyF)]+: a strong oxidizing xenonium(II) teflate cation with N-donor bases.

Herein we report on the formation of the adduct salts [Xe(OTeF5)(pyF)][Al(OTeF5)4] (pyF = C5F5N, C5H3F2N) by abstraction of an -OTeF5 group from Xe(OTeF5)2 with the Lewis superacid Al(OTeF5)3 and subsequent adduct formation of the generated [XeOTeF5]+ cation with fluorinated pyridines. These salts represent the first xenonium cations with the weakly coordinating [Al(OTeF5)4]- anion. The strong oxidizing property of these compounds is further assessed.

Questing for homoleptic mononuclear manganese complexes with monodentate O-donor ligands.

Compounds containing Mn-O bonds are of utmost importance in biological systems and catalytic processes. Nevertheless, mononuclear manganese complexes containing all O-donor ligands are still rare. Taking advantage of the low tendency of the pentafluoroorthotellurate ligand (teflate, OTeF5) to bridge metal centers, we have synthesized two homoleptic manganese complexes with monomeric structures and an all O-donor coordination sphere. The tetrahedrally distorted MnII anion, [Mn(OTeF5)4]2-, can be described as a high spin d5 complex (S = 5/2), as found experimentally (magnetic susceptibility measurements and EPR spectroscopy) and using theoretical calculations (DFT and CASSCF/NEVPT2). The high spin d4 electronic configuration (S = 2) of the MnIII anion, [Mn(OTeF5)5]2-, was also determined experimentally and theoretically, and a square pyramidal geometry was found to be the most stable one for this complex. Finally, the bonding situation in both complexes was investigated by means of the Interacting Quantum Atoms (IQA) methodology and compared to that of hypothetical mononuclear fluoromanganates. Within each pair of [MnXn]2- (n = 4, 5) species (X = OTeF5, F), the Mn-X interaction is found to be comparable, therefore proving that the similar electronic properties of the teflate and the fluoride are also responsible for the stabilization of these unique species.

GES-18, a new carbapenem-hydrolyzing GES-Type ß-lactamase from Pseudomonas aeruginosa that contains Ile80 and Ser170 residues.

A clinical isolate of Pseudomonas aeruginosa recovered from the lower respiratory tract of an 81-year-old patient hospitalized in Belgium was sent to the national reference center to determine its resistance mechanism. PCR sequencing identified a new GES variant, GES-18, which differs from the carbapenem-hydrolyzing enzyme GES-5 by a single amino acid substitution (Val80Ile, in the numbering according to Ambler) and from GES-1 by two substitutions (Val80Ile and Gly170Ser). Detailed kinetic characterization showed that GES-18 and GES-5 hydrolyze imipenem and cefoxitin with similar kinetic parameters and that GES-18 was less susceptible than GES-1 to classical ß-lactamase inhibitors such as clavulanate and tazobactam. The overall structure of GES-18 is similar to the solved structures of GES-1 and GES-2, the Val80Ile and Gly170Ser substitutions causing only subtle local rearrangements. Notably, the hydrolytic water molecule and the Glu166 residue were slightly displaced compared to their counterparts in GES-1. Our kinetic and crystallographic data for GES-18 highlight the pivotal role of the Gly170Ser substitution which distinguishes GES-5 and GES-18 from GES-1.

A rapid and selective HPLC-UV method for the quantitation of efavirenz in plasma from patients on concurrent HIV/AIDS and tuberculosis treatments.

Owing to heterogeneity in therapeutic response, efavirenz is of research and clinical interest. There is a need to quantitate it using noncostly and selective methods. A method for efavirenz quantitation in plasma containing HIV and tuberculosis drugs was developed. Chromatographic separation was carried out using a C18 column. The mobile phase consisted of 0.1% formic acid and acetonitrile, and was pumped at a flow rate of 0.3 mL/min. Efavirenz and ritonavir (internal standard) were monitored at 247 nm. Plasma proteins were precipitated by centrifugation. The analysis time was 6 min. The response was linear (r = 0.9997). The accuracy ranged between 98 and 115% (intraday) and between 99 and 117% (interday). The precision ranged from 1.670 to 4.087% (intraday) and from 3.447 to 13.347% (interday). Recovery ranged from 98 to 132%. Stability ranged between 99 and 123%. The selectivity was proven by analysis of drugs used for the management of HIV/AIDS and tuberculosis. Plasma sample analysis showed an efavirenz retention time of 5.57 min and a peak plasma concentration of 2.4 µg/mL occurring at 2 h. This method is rapid and selective, and thus suitable for monitoring efavirenz in patients with HIV/AIDS alone or co-infected with tuberculosis in a less resourced setting.

Kinetic and crystallographic studies of extended-spectrum GES-11, GES-12, and GES-14 ß-lactamases.

GES-1 is a class A extended-spectrum ß-lactamase conferring resistance to penicillins, narrow- and expanded-spectrum cephalosporins, and ceftazidime. However, GES-1 poorly hydrolyzes aztreonam and cephamycins and exhibits very low k(cat) values for carbapenems. Twenty-two GES variants have been discovered thus far, differing from each other by 1 to 3 amino acid substitutions that affect substrate specificity. GES-11 possesses a Gly243Ala substitution which seems to confer to this variant an increased activity against aztreonam and ceftazidime. GES-12 differs from GES-11 by a single Thr237Ala substitution, while GES-14 differs from GES-11 by the Gly170Ser mutation, which is known to confer increased carbapenemase activity. GES-11 and GES-12 were kinetically characterized and compared to GES-1 and GES-14. Purified GES-11 and GES-12 showed strong activities against most tested ß-lactams, with the exception of temocillin, cefoxitin, and carbapenems. Both variants showed a significantly increased rate of hydrolysis of cefotaxime, ceftazidime, and aztreonam. On the other hand, GES-11 and GES-12 (and GES-14) variants all containing Ala243 exhibited increased susceptibility to classical inhibitors. The crystallographic structures of the GES-11 and GES-14 ß-lactamases were solved. The overall structures of GES-11 and GES-14 are similar to that of GES-1. The Gly243Ala substitution caused only subtle local rearrangements, notably in the typical carbapenemase disulfide bond. The active sites of GES-14 and GES-11 are very similar, with the Gly170Ser substitution leading only to the formation of additional hydrogen bonds of the Ser residue with hydrolytic water and the Glu166 residue.

Detection and characterization of VIM-31, a new variant of VIM-2 with Tyr224His and His252Arg mutations, in a clinical isolate of Enterobacter cloacae.

We report the first description of the metallo-ß-lactamase VIM-31, a new variant of VIM-2 with Tyr224His and His252Arg mutations, in Enterobacter cloacae 11236, which was isolated from blood specimens of a patient with colonic adenocarcinoma in Belgium. bla(VIM-31) was found on a class 1 integron located on a self-transferable but not typeable 42-kb plasmid. Compared to values published elsewhere for VIM-2, the purified VIM-31 enzyme showed weaker catalytic efficiency against all the tested beta-lactam agents (except for ertapenem), resulting from lower k(cat) (except for ertapenem) and higher K(m) values for VIM-31.

The CphAII protein from Aquifex aeolicus exhibits a metal-dependent phosphodiesterase activity.

The CphAII protein from the hyperthermophile Aquifex aeolicus shows the five conserved motifs of the metallo-ß-lactamase (MBL) superfamily and presents 28% identity with the Aeromonas hydrophila subclass B2 CphA MBL. The gene encoding CphAII was amplified by PCR from the A. aeolicus genomic DNA and overexpressed in Escherichia coli using a pLex-based expression system. The recombinant CphAII protein was purified by a combination of heating (to denature E. coli proteins) and two steps of immobilized metal affinity chromatography. The purified enzyme preparation did not exhibit a ß-lactamase activity but showed a metal-dependent phosphodiesterase activity versus bis-p-nitrophenyl phosphate and thymidine 5'-monophosphate p-nitrophenyl ester, with an optimum at 85°C. The circular dichroism spectrum was in agreement with the percentage of secondary structures characteristic of the MBL αßßα fold.

Development and application of a highly sensitive LC-MS/MS method for simultaneous quantification of N,N-dimethyltryptamine and two of its metabolites in human plasma.

A highly sensitive LC-MS/MS method for the quantification of N,N-dimethyltryptamine (DMT) and its metabolites indole-3-acetic acid and DMT N-oxide in human plasma has been developed and validated. Chromatography was performed using a diphenyl column with gradient elution (0.1% formic acid in methanol/water). The mass spectrometer was operated in multiple reaction monitoring mode. A methanolic solution containing internal standards 2-methylindole 3-acetic acid and deuterated DMT, was added to plasma samples, followed by protein precipitation with acetonitrile. The samples were centrifuged and supernatants transferred to new tubes and evaporated to dryness before reconstitution in aqueous mobile phase. The method was validated with regards to accuracy, precision, sensitivity, selectivity, recovery, matrix effects, stability, carry-over and dilution integrity. The validated linear range was 0.25-200 nM for DMT and 15-250 nM for DMT N-oxide. For the endogenous compound indole-3-acetic acid a different approach was taken due to its significant presence in blank samples. The change in signal response from a blank sample was used when constructing the calibration curve with linearity demonstrated between elevations of 500-5000 nM above the blank. Applicability of the described method was demonstrated through analysis of plasma samples from healthy volunteers having received intravenous injections of DMT. The presented method for rapid and sensitive quantification of DMT and its metabolites in human plasma can be applied to future studies aiming to characterize DMT disposition and its relationship to immediate psychedelic or long-term antidepressive effects.

Air-stable aryl derivatives of pentafluoroorthotellurate.

We report on two different sets of air-stable derivatives of pentafluoroorthotellurate containing fluorinated and non-fluorinated aryl groups. The acid cis-PhTeF4OH was obtained in gram scale and further transformed to Ag[cis-PhTeF4O], which was used as a cis-PhTeF4O transfer reagent to obtain [PPh4][cis-PhTeF4O]. Furthermore, the synthesis of trans-(C6F5)2TeF3OH was achieved by a selective hydrolysis of trans-(C6F5)2TeF4 in the presence of KF and subsequent protonation by aHF. Quantum-chemical calculations show a higher acidity and robustness against fluoride abstraction for trans-(C6F5)2TeF3OH compared to cis-PhTeF4OH.