Obtusaquinone: A Cysteine-Modifying Compound That Targets Keap1 for Degradation.

We have previously identified the natural product obtusaquinone (OBT) as a potent antineoplastic agent with promising in vivo activity in glioblastoma and breast cancer through the activation of oxidative stress; however, the molecular properties of this compound remained elusive. We used a multidisciplinary approach comprising medicinal chemistry, quantitative mass spectrometry-based proteomics, functional studies in cancer cells, and pharmacokinetic analysis, as well as mouse xenograft models to develop and validate novel OBT analogs and characterize the molecular mechanism of action of OBT. We show here that OBT binds to cysteine residues with a particular affinity to cysteine-rich Keap1, a member of the CUL3 ubiquitin ligase complex. This binding promotes an overall stress response and results in ubiquitination and proteasomal degradation of Keap1 and downstream activation of the Nrf2 pathway. Using positron emission tomography (PET) imaging with the PET-tracer 2-[18F]fluoro-2-deoxy-d-glucose (FDG), we confirm that OBT is able to penetrate the brain and functionally target brain tumors. Finally, we show that an OBT analog with improved pharmacological properties, including enhanced potency, stability, and solubility, retains the antineoplastic properties in a xenograft mouse model.

Measurement of drug-target engagement in live cells by two-photon fluorescence anisotropy imaging.

The ability to directly image and quantify drug-target engagement and drug distribution with subcellular resolution in live cells and whole organisms is a prerequisite to establishing accurate models of the kinetics and dynamics of drug action. Such methods would thus have far-reaching applications in drug development and molecular pharmacology. We recently presented one such technique based on fluorescence anisotropy, a spectroscopic method based on polarization light analysis and capable of measuring the binding interaction between molecules. Our technique allows the direct characterization of target engagement of fluorescently labeled drugs, using fluorophores with a fluorescence lifetime larger than the rotational correlation of the bound complex. Here we describe an optimized protocol for simultaneous dual-channel two-photon fluorescence anisotropy microscopy acquisition to perform drug-target measurements. We also provide the necessary software to implement stream processing to visualize images and to calculate quantitative parameters. The assembly and characterization part of the protocol can be implemented in 1 d. Sample preparation, characterization and imaging of drug binding can be completed in 2 d. Although currently adapted to an Olympus FV1000MPE microscope, the protocol can be extended to other commercial or custom-built microscopes.

One-Carbon Homologation of Primary Alcohols to Carboxylic Acids, Esters, and Amides via Mitsunobu Reactions with MAC Reagents.

A method is reported for the one-carbon homologation of an alcohol to the extended carboxylic acid, ester, or amide. The process involves the Mitsunobu reaction with an alkoxymalononitrile, followed by unmasking in the presence of a suitable nucleophile. The homologation and unmasking can even be performed in a one-pot process in high yield.

Access to Spirocyclized Oxindoles and Indolenines via Palladium-Catalyzed Cascade Reactions of Propargyl Carbonates with 2-Oxotryptamines and Tryptamines.

Reported here are methods for the direct construction of a range of spirocyclized oxindoles and indolenines in good to excellent yields. Specifically, we report the palladium-catalyzed reactions of oxindoles and indoles, both functioning as bis-nucleophiles, with propargyl carbonates to afford spirocyclic products having an exocyclic double bond on the newly formed ring. The reaction proceeds through a process wherein the first nucleophilic unit on the oxindole or indole reacts with an allenyl-palladium species, formed from oxidative addition of Pd(0) to propargyl carbonates, to generate a π-allyl palladium intermediate that then reacts further with the second nucleophilic component of the oxindole or indole. The cascade process forges two bonds en route to spirocyclized oxindole and indolenine products. The use of chiral phosphines renders the cyclization sequence enantioselective, providing spirocyclic products with modest to good enantioselectivities.

A biomimetic strategy to access the silybins: total synthesis of (-)-isosilybin A.

We report the first asymmetric, total synthesis of (-)-isosilybin A. A late-stage catalytic biomimetic cyclization of a highly functionalized chalcone is employed to form the characteristic benzopyranone ring. A robust and flexible approach to this chalcone provides an entry to the preparation of the entire isomeric family of silybin natural products.

Rapid access to spirocyclized indolenines via palladium-catalyzed cascade reactions of tryptamine derivatives and propargyl carbonate.

We report the intermolecular palladium-catalyzed reaction of tert-butyl propargyl carbonate with tryptamine derivatives or other indole-containing bis-nucleophiles. The reaction proceeds under mild conditions and with low catalyst loadings to afford novel spiroindolenine products in good to high yields.

Squaramide-catalyzed enantioselective Michael addition of masked acyl cyanides to substituted enones.

Masked acyl cyanide (MAC) reagents are shown to be effective umpolung synthons for enantioselective Michael addition to substituted enones. The reactions are catalyzed by chiral squaramides and afford adducts in high yields (90-99%) and with excellent enantioselectivities (85-98%). The addition products are unmasked to produce dicyanohydrins that, upon treatment with a variety of nucleophiles, provide γ-keto acids, esters, and amides. The use of this umpolung synthon has enabled, in enantiomerically enriched form, the first total synthesis of the prenylated phenol (+)-fornicin C.

Asymmetric Methods for the Synthesis of Flavanones, Chromanones, and Azaflavanones.

Flavanones, chromanones, and related structures are privileged natural products that display a wide variety of biological activities. Although flavanoids are abundant in nature, there are a limited number of available general and efficient synthetic methods for accessing molecules of this class in a stereoselective manner. Their structurally simple architectures belie the difficulties involved in installation and maintenance of the stereogenic configuration at the C2 position, which can be sensitive and can undergo epimerization under mildly acidic, basic, and thermal reaction conditions. This review presents the methods currently used to access these related structures. The synthetic methods include manipulation of the flavone/flavanone core, carbon-carbon bond formation, and carbon-heteroatom bond formation.

Concise syntheses of the abyssinones and discovery of new inhibitors of prostate cancer and MMP-2 expression.

Hydrogen-bonding catalysis is an emerging field that facilitates rapid access to medicinally relevant enantioenriched small molecules. Here, we report the first asymmetric total syntheses of four members of the abyssinone class of natural products (I, II, III, and IV 4´-OMe) via quinine- or quinidine-derived thiourea-catalyzed intramolecular conjugate additions of ß-keto ester alkylidenes. This concise strategy includes a tandem deprotection/decarboxylation final step that delivers all four natural products and their corresponding antipodes. A preliminary evaluation of all of these small molecules against a metastatic human prostate cancer cell line has identified that these compounds selectively and differentially inhibit cell growth and downregulate the expression of matrix metalloproteinase-2 (MMP-2) at non-toxic concentrations.

Catalytic asymmetric alkylation of substituted isoflavanones.

The asymmetric alkylation of isoflavanones (3-aryl-chroman-4-ones) and protected 3-phenyl-2,3-dihydroquinolin-4(1H)-ones catalyzed by a novel cinchonidine-derived phase transfer catalyst E is reported. This functionalization occurs at the unactivated C3 methine to afford novel products that can easily be functionalized to generate more complex fused ring systems. The process accommodates a variety of isoflavanones and activated electrophiles and installs a stereogenic quaternary center in high yield and with good-to-excellent selectivity. Isoflavanones are a privileged class of natural products with a broad spectrum of biological activities including insecticidal, antimicrobial, antibacterial, estrogenic, antitumor, and anti-HIV activity. (1) Isoflavanones are also precursors for more complex natural products such as pterocarpans and rotenones. (1) Given their therapeutic promise, selective strategies to access new classes of isoflavanones and related structures has high value. (2) The functionalization of the C3 position could promote beneficial interactions with biological targets of interest. Specifically, an alkylation at C3 can rapidly access new members of the general class of biologically active homoisoflavanones. (3).