Site-Selective Functionalization of Unactivated Allylic C-H Bonds via Direct Deprotonation with KTMP: Application to the Formal Total Synthesis of (+)-Artemisinin from Amorphadiene.

The site-selective functionalization of unactivated allylic C-H bonds via direct deprotonation using KTMP is described. The conversion of amorphadiene to artemisinic alcohol via a simple, highly regioselective deprotonation over 4 other possible allylic sites is shown with further extrapolation to the first large-scale telescoped chemical synthesis of artemisinic acid from amorphadiene. Finally, application of the method for the successful site-selective functionalization of unactivated allylic C-H bonds in other terpene-based natural products is also highlighted.

A Metal-Free Reductive N-Alkylation of Indoles with Aldehydes.

A simple metal-free method has been developed for the reductive N-alkylation of indoles employing aldehydes as the alkylating agent and inexpensive Et3SiH as the reductant. A wide range of aromatic and aliphatic aldehydes are viable substrates along with a variety of substituted indoles. In addition, the method was applied to a one-pot sequential 1,3-alkylation of a substituted indole and successfully demonstrated on a 100 mmol scale.

Synthesis and Biological Evaluations of Electrophilic Steroids Inspired by the Taccalonolides.

Natural products have served as inspirational scaffolds for the design and synthesis of novel antineoplastic agents. Here we present our preliminary efforts on the synthesis and biological evaluation of a new class of electrophilic steroids inspired by the naturally occurring taccalonolides. We demonstrate that these simplified analogs exhibit highly persistent antiproliferative properties similar to the taccalonolides and retain activity against resistant cancer cell lines that warrants further preclinical development.

Dysregulated ribonucleoprotein granules promote cardiomyopathy in RBM20 gene-edited pigs.

Ribonucleoprotein (RNP) granules are biomolecular condensates-liquid-liquid phase-separated droplets that organize and manage messenger RNA metabolism, cell signaling, biopolymer assembly, biochemical reactions and stress granule responses to cellular adversity. Dysregulated RNP granules drive neuromuscular degenerative disease but have not previously been linked to heart failure. By exploring the molecular basis of congenital dilated cardiomyopathy (DCM) in genome-edited pigs homozygous for an RBM20 allele encoding the pathogenic R636S variant of human RNA-binding motif protein-20 (RBM20), we discovered that RNP granules accumulated abnormally in the sarcoplasm, and we confirmed this finding in myocardium and reprogrammed cardiomyocytes from patients with DCM carrying the R636S allele. Dysregulated sarcoplasmic RBM20 RNP granules displayed liquid-like material properties, docked at precisely spaced intervals along cytoskeletal elements, promoted phase partitioning of cardiac biomolecules and fused with stress granules. Our results link dysregulated RNP granules to myocardial cellular pathobiology and heart failure in gene-edited pigs and patients with DCM caused by RBM20 mutation.

Pd- and Ni-Based Systems for the Catalytic Borylation of Aryl (Pseudo)halides with B2(OH)4.

Despite recent advancements in metal-catalyzed borylations of aryl (pseudo)halides, there is a continuing need to develop robust methods to access both early-stage and late-stage organoboron intermediates amendable for further functionalization. In particular, the development of general catalytic systems that operate under mild reaction conditions across a broad range of electrophilic partners remains elusive. Herein, we report the development and application of three catalytic systems (two Pd-based and one Ni-based) for the direct borylation of aryl (pseudo)halides using tetrahydroxydiboron (B2(OH)4). For the Pd-based catalyst systems, we have identified general reaction conditions that allow for the sequestration of halide ions through simple precipitation that results in catalyst loadings as low as 0.01 mol % (100 ppm) and reaction temperatures as low as room temperature. We also describe a complementary Ni-based catalyst system that employs simple unligated Ni(II) salts as an inexpensive alternative to the Pd-based systems for the borylation of aryl (pseudo)halides. Extrapolation of all three systems to a one-pot tandem borylation/Suzuki-Miyaura cross-coupling is also demonstrated on advanced intermediates and drug substances.

An Electrophilic Natural Product Provides a Safe and Robust Odor Neutralization Approach To Counteract Malodorous Organosulfur Metabolites Encountered in Skunk Spray.

The anal secretions of skunks comprise several types of malodorous organosulfur compounds. The pungent metabolites are used defensively by skunks to repel threats posed by predators, and in many parts of the world, those perceived threats include humans and their pets. The extremely low thresholds for detection of the organosulfur metabolites make efforts to "de-skunk" people, animals, and clothing a process fraught with many challenges. The fungal-derived metabolite pericosine A (4) is a promiscuous yet stabile electrophilic compound that we propose is used by some fungi as a novel form of chemical defense. Our investigations have indicated that pericosine A readily reacts with skunk-spray secretions to transform them into odorless products. Mechanistic and computational studies suggested that pericosine A and its synthetic analogues react via SN2'-type mechanisms with thiols and thioacetates under aqueous conditions to generate stable thioethers. Testing revealed that pericosine A did not cause skin or eye irritation and was highly effective at deodorizing skunk anal gland secretions when formulated to include adjunctive cosmetic ingredients.

Cationic Alkynyl Heck Reaction toward Substituted Allenes Using BobCat: A New Hybrid Pd(0)-Catalyst Incorporating a Water-Soluble dba Ligand.

The cationic alkynyl Heck reaction between aryl triflates and alkynes to give substituted allenes is described. Key to the success of this method was the discovery and development of a new hybrid Pd(0)-catalyst, BobCat, that incorporates a water-soluble dba-ligand and biaryl phosphine ligand to provide substituted allenes in good yields under mild reaction conditions.

Direct Lewis Acid Catalyzed Conversion of Enantioenriched N-Acyloxazolidinones to Chiral Esters, Amides, and Acids.

The identification of Yb(OTf)3 through a multivariable high-throughput experimentation strategy has enabled a unified protocol for the direct conversion of enantioenriched N-acyloxazolidinones to the corresponding chiral esters, amides, and carboxylic acids. This straightforward and catalytic method has shown remarkable chemoselectivity for substitution at the acyclic N-acyl carbonyl for a diverse array of N-acyloxazolidinone substrates. The ionic radius of the Lewis acid catalyst was demonstrated as a key driver of catalyst performance that led to the identification of a robust and scalable esterification of a pharmaceutical intermediate using catalytic Y(OTf)3.

Ru(II)-Catalyzed Synthesis of Substituted Furans and Their Conversion to Butenolides.

The synthesis of densely functionalized trisubstituted and tetrasubstituted furans via a novel Ru(II)-catalyzed intramolecular cyclization of vinyl diazoesters is reported. The synthetic utility of these furans is further demonstrated through a simple acid-mediated reaction to access highly substituted Δα,ß-butenolides.

Unified Approach to Substituted Allenoates via Pd-Catalyzed ß-Hydride Elimination of (E)-Enol Triflates.

A robust synthesis of allenoates via a Pd-catalyzed ß-hydride elimination of (E)-enol triflates is presented. Salient features of this method include low catalyst loadings, mild reaction conditions, and the ability to access all four patterns of substituted allenoates from a single substrate class.

Palladium-Catalyzed Synthesis of Alkynes via a Tandem Decarboxylation/Elimination of (E)-Enol Triflates.

A mild catalytic synthesis of alkynes via a tandem Pd-catalyzed decarboxylation/elimination of enol triflates is described. Key attributes of the method include readily available starting materials, broad functional group tolerance, and the ability to access terminal, internal, and halogenated alkynes. The preliminary scope of the reaction is demonstrated on 25 different examples with yields ranging from 63% to 96%.

Iron-Catalyzed Stereoselective Cross-Coupling Reactions of Stereodefined Enol Carbamates with Grignard Reagents.

A practical and highly stereoselective iron-catalyzed cross-coupling reaction of stereodefined enol carbamates and Grignard reagents to yield tri- and tetrasubstituted acrylates is reported. A facile method for the stereoselective generation of these enol carbamates has also been developed.

Development of an ex vivo lymph node explant model for identification of novel molecules active against Leishmania major.

Leishmaniasis is a vector-borne zoonotic infection affecting people in tropical and subtropical regions of the world. Current treatments for cutaneous leishmaniasis are difficult to administer, toxic, expensive, and limited in effectiveness and availability. Here we describe the development and application of a medium-throughput screening approach to identify new drug candidates for cutaneous leishmaniasis using an ex vivo lymph node explant culture (ELEC) derived from the draining lymph nodes of Leishmania major-infected mice. The ELEC supported intracellular amastigote proliferation and contained lymph node cell populations (and their secreted products) that enabled the testing of compounds within a system that mimicked the immunopathological environment of the infected host, which is known to profoundly influence parasite replication, killing, and drug efficacy. The activity of known antileishmanial drugs in the ELEC system was similar to the activity measured in peritoneal macrophages infected in vitro with L. major. Using the ELEC system, we screened a collection of 334 compounds, some of which we had demonstrated previously to be active against L. donovani, and identified 119 hits, 85% of which were confirmed to be active by determination of the 50% effective concentration (EC50). We found 24 compounds (7%) that had an in vitro therapeutic index (IVTI; 50% cytotoxic/effective concentration [CC50]/EC50) > 100; 19 of the compounds had an EC50 below 1 µM. According to PubChem searchs, 17 of those compounds had not previously been reported to be active against Leishmania. We expect that this novel method will help to accelerate discovery of new drug candidates for treatment of cutaneous leishmaniasis.

Genetic and molecular basis of drug resistance and species-specific drug action in schistosome parasites.

Oxamniquine resistance evolved in the human blood fluke (Schistosoma mansoni) in Brazil in the 1970s. We crossed parental parasites differing ~500-fold in drug response, determined drug sensitivity and marker segregation in clonally derived second-generation progeny, and identified a single quantitative trait locus (logarithm of odds = 31) on chromosome 6. A sulfotransferase was identified as the causative gene by using RNA interference knockdown and biochemical complementation assays, and we subsequently demonstrated independent origins of loss-of-function mutations in field-derived and laboratory-selected resistant parasites. These results demonstrate the utility of linkage mapping in a human helminth parasite, while crystallographic analyses of protein-drug interactions illuminate the mode of drug action and provide a framework for rational design of oxamniquine derivatives that kill both S. mansoni and S. haematobium, the two species responsible for >99% of schistosomiasis cases worldwide.

The bat flower: a source of microtubule-destabilizing and -stabilizing compounds with synergistic antiproliferative actions.

The biosynthesis of secondary metabolites provides higher plants with mechanisms of defense against microbes, insects, and herbivores. One common cellular target of these molecules is the highly conserved microtubule cytoskeleton, and microtubule-targeting compounds with insecticidal, antifungal, nematicidal, and anticancer activities have been identified from plants. A new retro-dihydrochalcone, taccabulin A, with microtubule-destabilizing activity has been identified from the roots and rhizomes of Tacca species. This finding is notable because the microtubule-stabilizing taccalonolides are also isolated from these sources. This is the first report of an organism producing compounds with both microtubule-stabilizing and -destabilizing activities. A two-step chemical synthesis of taccabulin A was performed. Mechanistic studies showed that taccabulin A binds within the colchicine site on tubulin and has synergistic antiproliferative effects against cancer cells when combined with a taccalonolide, which binds to a different site on tubulin. Taccabulin A is effective in cells that are resistant to many other plant-derived compounds. The discovery of a natural source that contains both microtubule-stabilizing and -destabilizing small molecules is unprecedented and suggests that the synergistic action of these compounds was exploited by nature long before it was discovered in the laboratory.

A small molecule modulates Jumonji histone demethylase activity and selectively inhibits cancer growth.

The pharmacological inhibition of general transcriptional regulators has the potential to block growth through targeting multiple tumorigenic signalling pathways simultaneously. Here, using an innovative cell-based screen, we identify a structurally unique small molecule (named JIB-04) that specifically inhibits the activity of the Jumonji family of histone demethylases in vitro, in cancer cells, and in tumours in vivo. Unlike known inhibitors, JIB-04 is not a competitive inhibitor of α-ketoglutarate. In cancer, but not in patient-matched normal cells, JIB-04 alters a subset of transcriptional pathways and blocks viability. In mice, JIB-04 reduces tumour burden and prolongs survival. Importantly, we find that patients with breast tumours that overexpress Jumonji demethylases have significantly lower survival. Thus, JIB-04, a novel inhibitor of Jumonji demethylases in vitro and in vivo, constitutes a unique potential therapeutic and research tool against cancer, and validates the use of unbiased cellular screens to discover chemical modulators with disease relevance.

Pd-catalyzed asymmetric ß-hydride elimination en route to chiral allenes.

We wish to report our preliminary results on the discovery and development of a catalytic, asymmetric ß-hydride elimination from vinyl Pd(II)-complexes derived from enol triflates to access chiral allenes. To achieve this, we developed a class of chiral phosphite ligands that demonstrate high enantioselectivity, allow access of either allene enantiomer, and are readily synthesized. The methodology is demonstrated on over 20 substrates, and application to the formal asymmetric total synthesis of the natural product, (+)-epibatidine, is also provided.

Development of inhibitors of the PAS-B domain of the HIF-2α transcription factor.

Hypoxia inducible factors (HIFs) are heterodimeric transcription factors induced in a variety of pathophysiological settings, including cancer. We describe the first detailed structure-activity relationship study of small molecules designed to inhibit HIF-2α-ARNT heterodimerization by binding an internal cavity of the HIF-2α PAS-B domain. Through a series of biophysical characterizations of inhibitor-protein interactions (NMR and X-ray crystallography), we have established the structural requirements for artificial inhibitors of the HIF-2α-ARNT PAS-B interaction. These results may serve as a foundation for discovering therapeutic agents that function by a novel mode of action.

Allosteric inhibition of hypoxia inducible factor-2 with small molecules.

Hypoxia inducible factors (HIFs) are heterodimeric transcription factors induced in many cancers where they frequently promote the expression of protumorigenic pathways. Though transcription factors are typically considered 'undruggable', the PAS-B domain of the HIF-2α subunit contains a large cavity within its hydrophobic core that offers a unique foothold for small-molecule regulation. Here we identify artificial ligands that bind within this pocket and characterize the resulting structural and functional changes caused by binding. Notably, these ligands antagonize HIF-2 heterodimerization and DNA-binding activity in vitro and in cultured cells, reducing HIF-2 target gene expression. Despite the high sequence identity between HIF-2α and HIF-1α, these ligands are highly selective and do not affect HIF-1 function. These chemical tools establish the molecular basis for selective regulation of HIF-2, providing potential therapeutic opportunities to intervene in HIF-2-driven tumors, such as renal cell carcinomas.