Stereochemical Inversion of a Cyano-Stabilized Grignard Reagent: Remarkable Effects of the Ethereal Solvent Structure and Concentration

Chiral organometallic reagents are useful in asymmetric synthesis, and configurational stability of these species is critical to success. In this study we followed the epimerization of a chiral Grignard reagent, prepared by Mg/Br exchange of bromonitrile <i>trans</i>-<b>2b</b>. This compound underwent highly retentive Mg/Br exchange in Et₂O; less retention was observed in 2-MeTHF and THF. Epimerization rate constants <i>k</i>_tc were determined at 195 K by measuring the diastereomer ratio of deuteration product <i>d</i>₁-<b>3b</b> as a function of the delay time before quench. Studies were also performed at varying concentrations of Et₂O in toluene. Remarkable dynamic range in <i>k</i>_tc was seen: relative to reaction at 0.12 M Et₂O in toluene, epimerization was 26-, 800-, and 1300-fold faster in Et₂O, 2-MeTHF, and THF, respectively. Thus, the identity and concentration of an ethereal solvent can dramatically affect configurational stability. Reaction stoichiometry experiments suggested that, in Et₂O, the Grignard reagent derived from <i>trans</i>-<b>2b</b> exists as an <i>i</i>-PrMgCl heterodimer; the invariance of <i>k</i>_tc over a 20-fold range in [Mg]_total ruled out mandatory deaggregation (or aggregation) on the epimerization path. Analysis of the dependency of <i>k</i>_tc on [Et₂O] and temperature in Et₂O/toluene solution at 195, 212, and 231 K indicated fast incremental solvation before rate-limiting ion-pair separation and provided an estimate of the entropic cost of capturing a solvent ligand (−13 ± 3 eu). Calculations at the MP2/6-31G*­(PCM)//B3LYP/6-31G* level provide support for these conclusions and map out a possible “ionogenic conducted tour” pathway for epimerization

Access to “Friedel–Crafts-Restricted” <i>tert</i>-Alkyl Aromatics by Activation/Methylation of Tertiary Benzylic Alcohols

Herein we describe a two-step protocol to prepare <i>m</i>-<i>tert</i>-alkylbenzenes. The appropriate tertiary benzylic alcohols are activated with SOCl₂ or concentrated HCl and then treated with trimethylaluminum, affording the desired products in 68–97% yields (22 examples). This reaction sequence is successful in the presence of a variety of functional groups, including acid-sensitive and Lewis-basic groups. In addition to <i>t</i>-Bu groups, 1,1-dimethylpropyl and 1-ethyl-1-methylpropyl groups can also be installed using this method

Enantioselective Deprotonative Ring Contraction of <i>N</i>1‑Methyl‑<i>N</i>4‑Boc-benzo[<i>e</i>][1,4]diazepine-2,5-diones

<i>N</i>1-Methyl-<i>N</i>4-Boc-benzo­[<i>e</i>]­[1,4]­diazepine-2,5-diones were prepared in good yield and high stereochemical purity from five amino acids. Upon deprotonation, these compounds undergo ring contraction to the corresponding quinolone-2,4-diones with high enantioselectivity, providing efficient entry to a potentially useful drug scaffold. Mechanistic commentary and comparisons to related reactions are provided

Select Small Core Structure Carbamates Exhibit High Contact Toxicity to “Carbamate-Resistant” Strain Malaria Mosquitoes, <em>Anopheles gambiae</em> (Akron)

<div><p>Acetylcholinesterase (AChE) is a proven target for control of the malaria mosquito (<em>Anopheles gambiae</em>). Unfortunately, a single amino acid mutation (G119S) in <em>An. gambiae</em> AChE-1 (<em>Ag</em>AChE) confers resistance to the AChE inhibitors currently approved by the World Health Organization for indoor residual spraying. In this report, we describe several carbamate inhibitors that potently inhibit G119S <em>Ag</em>AChE and that are contact-toxic to carbamate-resistant <em>An. gambiae</em>. PCR-RFLP analysis was used to confirm that carbamate-susceptible G3 and carbamate-resistant Akron strains of <em>An. gambiae</em> carry wild-type (WT) and G119S AChE, respectively. G119S <em>Ag</em>AChE was expressed and purified for the first time, and was shown to have only 3% of the turnover number (<em>k</em>_cat) of the WT enzyme. Twelve carbamates were then assayed for inhibition of these enzymes. High resistance ratios (>2,500-fold) were observed for carbamates bearing a benzene ring core, consistent with the carbamate-resistant phenotype of the G119S enzyme. Interestingly, resistance ratios for two oxime methylcarbamates, and for five pyrazol-4-yl methylcarbamates were found to be much lower (4- to 65-fold). The toxicities of these carbamates to live G3 and Akron strain <em>An. gambiae</em> were determined. As expected from the enzyme resistance ratios, carbamates bearing a benzene ring core showed low toxicity to Akron strain <em>An. gambiae</em> (LC₅₀>5,000 μg/mL). However, one oxime methylcarbamate (aldicarb) and five pyrazol-4-yl methylcarbamates (<b>4a</b>–<b>e</b>) showed good to excellent toxicity to the Akron strain (LC₅₀ = 32–650 μg/mL). These results suggest that appropriately functionalized “small-core” carbamates could function as a resistance-breaking anticholinesterase insecticides against the malaria mosquito.</p> </div

Biological Studies and Target Engagement of the 2‑<i>C</i>‑Methyl‑d‑Erythritol 4‑Phosphate Cytidylyltransferase (IspD)-Targeting Antimalarial Agent (1<i>R</i>,3<i>S</i>)‑MMV008138 and Analogs

Malaria continues to be one of the deadliest diseases worldwide, and the emergence of drug resistance parasites is a constant threat. <i>Plasmodium</i> parasites utilize the methylerythritol phosphate (MEP) pathway to synthesize isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), which are essential for parasite growth. Previously, we and others identified that the Malaria Box compound MMV008138 targets the apicoplast and that parasite growth inhibition by this compound can be reversed by supplementation of IPP. Further work has revealed that MMV008138 targets the enzyme 2-<i>C</i>-methyl-d-erythritol 4-phosphate cytidylyltransferase (IspD) in the MEP pathway, which converts MEP and cytidine triphosphate (CTP) to cytidinediphosphate methylerythritol (CDP-ME) and pyrophosphate. In this work, we sought to gain insight into the structure–activity relationships by probing the ability of MMV008138 analogs to inhibit <i>Pf</i>IspD recombinant enzyme. Here, we report <i>Pf</i>IspD inhibition data for fosmidomycin (FOS) and 19 previously disclosed analogs and report parasite growth and <i>Pf</i>IspD inhibition data for 27 new analogs of MMV008138. In addition, we show that MMV008138 does not target the recently characterized human IspD, reinforcing MMV008138 as a prototype of a new class of species-selective IspD-targeting antimalarial agents

Electrophoretic analysis (SDS-PAGE) of the purified r<i>Ag</i>AChE (WT and G119S).

<p>A: Protein standard (Da); B: r<i>Ag</i>AChE-WT (“n”); C: r<i>Ag</i>AChE-G119S mutant (“m”).</p

Enzyme resistance ratios and <i>Ag</i>AChE vs <i>h</i>AChE selectivity of selected carbamates.

a<p>Recombinant sources of <i>Ag</i>AChE are r<i>Ag</i>AChE-WT and r<i>Ag</i>AChE-G119S; homogenates are G3 (WT) and Akron (G119S). ^bResistance ratio is calculated as <i>k</i>_i(WT)/<i>k</i>_i(G119S); values are taken from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0046712#pone-0046712-t002" target="_blank">Table 2</a>. Standard error in the ratio is calculated according to a standard propagation of error formula <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0046712#pone.0046712-Andraos1" target="_blank">[61]</a>. ^cSelectivity for inhibiting <i>Ag</i>AChE (WT) vs <i>h</i>AChE, calculated as <i>k</i>_i(<i>Ag</i>AChE)/<i>k</i>_i(<i>h</i>AChE), with standard error in the ratio calculated according to a standard propagation of error formula <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0046712#pone.0046712-Andraos1" target="_blank">[61]</a>.</p

Tarsal contact toxicity (24 h) to G3 and Akron strain <i>An. gambiae</i>, and toxicity resistance ratios.

a<p>No mortality at this concentration. ^bLess than 10% mortality at this concentration. ^cDefined by LC₅₀ (Akron)/LC₅₀ (G3).</p

Carbamate inactivation rate constants <i>k</i>_i for r<i>Ag</i>AChE (WT & G119S), <i>An. gambiae</i> homogenates (G3 & Akron), and r<i>h</i>AChE.^a

a<p>Measured at 23±1°C, pH 7.7, 0.1% (v/v) DMSO. ^bG3 strain <i>An. gambiae</i> carry WT <i>Ag</i>AChE and possess a carbamate-susceptible phenotype. ^cAkron strain <i>An. gambiae</i> carry G119S mutant <i>Ag</i>AChE and possess a carbamate-resistant phenotype.</p

Computational modeling of tetrahedral intermediates formed by addition of the <i>Ag</i>AChE catalytic serine (S199) Oγ to carbamates.

<p>A) terbam with WT enzyme; location of catalytic serine oxygen Oγ is highlighted with the green arrow. B) terbam with G119S mutant enzyme; steric clash of the hydroxyl group of S119 with aromatic ring of terbam is noted with a blue arrow. C) Aldicarb with G119S mutant enzyme; note the absence of a steric clash with the hydroxyl group of S119. D) Pyrazol-4-yl methylcarbamate <b>4c</b> ((<i>S</i>)-enantiomer) with the G119S mutant enzyme; note the absence of a steric clash with the hydroxyl group of S119. Nonbonded contact distances in B and D are given in Å.</p