Synthesis of α3ß4 Nicotinic Acetylcholine Receptor Modulators Derived from Aristoquinoline That Reduce Reinstatement of Cocaine-Seeking Behavior.

Growing evidence suggests that inhibition of the α3ß4 nicotinic acetylcholine receptor (nAChR) represents a promising therapeutic strategy to treat cocaine use disorder. Recently, aristoquinoline (1), an alkaloid from Aristotelia chilensis, was identified as an α3ß4-selective nAChR inhibitor. Here, we prepared 22 derivatives of 1 and evaluated their ability to inhibit the α3ß4 nAChR. These studies revealed structure-activity trends and several compounds with increased potency compared to 1 with few off-target liabilities. Additional mechanistic studies indicated that these compounds inhibit the α3ß4 nAChR noncompetitively, but do not act as channel blockers, suggesting they are negative allosteric modulators. Finally, using a cocaine-primed reinstatement paradigm, we demonstrated that 1 significantly attenuates drug-seeking behavior in an animal model of cocaine relapse. The results from these studies further support a role for the α3ß4 nAChR in the addictive properties of cocaine and highlight the possible utility of aristoquinoline derivatives in treating cocaine use disorder.

Modified Akuamma Alkaloids with Increased Potency at the Mu-opioid Receptor.

Akuammine (1) and pseudoakuammigine (2) are indole alkaloids found in the seeds of the akuamma tree (Picralima nitida). Both alkaloids are weak agonists of the mu opioid receptor (µOR); however, they produce minimal effects in animal models of antinociception. To probe the interactions of 1 and 2 at the opioid receptors, we have prepared a collection of 22 semisynthetic derivatives. Evaluation of this collection at the µOR and kappa opioid receptor (κOR) revealed structural-activity relationship trends and derivatives with improved potency at the µOR. Most notably, the introduction of a phenethyl moiety to the N1 of 2 produces a 70-fold increase in potency and a 7-fold increase in selectivity for the µOR. The in vitro potency of this compound resulted in increased efficacy in the tail-flick and hot-plate assays of antinociception. The improved potency of these derivatives highlights the promise of exploring natural product scaffolds to probe the opioid receptors.

What We Have Gained from Ibogaine: α3ß4 Nicotinic Acetylcholine Receptor Inhibitors as Treatments for Substance Use Disorders.

For decades, ibogaine─the main psychoactive alkaloid found in Tabernanthe iboga─has been investigated as a possible treatment for substance use disorders (SUDs) due to its purported ability to interrupt the addictive properties of multiple drugs of abuse. Of the numerous pharmacological actions of ibogaine and its derivatives, the inhibition of α3ß4 nicotinic acetylcholine receptors (nAChRs), represents a probable mechanism of action for their apparent anti-addictive activity. In this Perspective, we examine several classes of compounds that have been discovered and developed to target α3ß4 nAChRs. Specifically, by focusing on compounds that have proven efficacious in pre-clinical models of drug abuse and have been evaluated clinically, we highlight the promising potential of the α3ß4 nAChRs as viable targets to treat a wide array of SUDs. Additionally, we discuss the challenges faced by the existing classes of α3ß4 nAChR ligands that must be overcome to develop them into therapeutic treatments.

Rational inhibitor design for Pseudomonas aeruginosa salicylate adenylation enzyme PchD.

Pseudomonas aeruginosa is an increasingly antibiotic-resistant pathogen that causes severe lung infections, burn wound infections, and diabetic foot infections. P. aeruginosa produces the siderophore pyochelin through the use of a non-ribosomal peptide synthetase (NRPS) biosynthetic pathway. Targeting members of siderophore NRPS proteins is one avenue currently under investigation for the development of new antibiotics against antibiotic-resistant organisms. Here, the crystal structure of the pyochelin adenylation domain PchD is reported. The structure was solved to 2.11 Å when co-crystallized with the adenylation inhibitor 5'-O-(N-salicylsulfamoyl)adenosine (salicyl-AMS) and to 1.69 Å with a modified version of salicyl-AMS designed to target an active site cysteine (4-cyano-salicyl-AMS). In the structures, PchD adopts the adenylation conformation, similar to that reported for AB3403 from Acinetobacter baumannii.

Syntheses of Aristotelia Alkaloids: Reflections in the Chiral Pool.

The Aristotelia alkaloids are a family of monoterpene indole alkaloids possessing a characteristic azabicyclononane scaffold, which has been assembled by several synthetic methods. Herein we review those approaches that have adopted a biomimetic approach to unite heterocyclic synthons with chiral pool monoterpenes. Throughout this discussion, the tendency of monoterpenes like α-pinene and limonene to undergo racemization is highlighted, revealing the challenges in developing stereospecific syntheses of these alkaloids. Finally, we provide a brief discussion of how these synthetic efforts have enabled the structural confirmation and elucidation of the Aristotelia alkaloids' absolute configurations, including our own recent efforts to employ bioactivity data to deduce the naturally occurring configuration of the quinoline alkaloid aristoquinoline.

Synthesis of Aristoquinoline Enantiomers and Their Evaluation at the α3ß4 Nicotinic Acetylcholine Receptor.

The first synthesis of aristoquinoline (1), a naturally occurring nicotinic acetylcholine receptor (nAChR) antagonist, was accomplished using two different approaches. Comparison of the synthetic material's spectroscopic data to that of the isolated alkaloid identified a previously misassigned stereogenic center. An evaluation of each enantiomer's activity at the α3ß4 nAChR revealed that (+)-1 is significantly more potent than (-)-1. This unexpected finding suggests that naturally occurring 1 possesses the opposite absolute configuration from indole-containing Aristotelia alkaloids.

Isolation and Pharmacological Characterization of Six Opioidergic Picralima nitida Alkaloids.

The seeds of the akuamma tree (Picralima nitida) have been used as a traditional treatment for pain and fever. Previous studies have attributed these effects to a series of indole alkaloids found within the seed extracts; however, these pharmacological studies were significantly limited in scope. Herein, an isolation protocol employing pH-zone-refining countercurrent chromatography was developed to provide six of the akuamma alkaloids in high purity and quantities sufficient for more extensive biological evaluation. Five of these alkaloids, akuammine (1), pseudo-akuammigine (3), akuammicine (4), akuammiline (5), and picraline (6), were evaluated against a panel of >40 central nervous system receptors to identify that their primary targets are the opioid receptors. Detailed in vitro investigations revealed 4 to be a potent kappa opioid receptor agonist, and three alkaloids (1-3) were shown to have micromolar activity at the mu opioid receptor. The mu opioid receptor agonists were further evaluated for analgesic properties but demonstrated limited efficacy in assays of thermal nociception. These findings contradict previous reports of the antinociceptive properties of the P. nitida alkaloids and the traditional use of akuamma seeds as analgesics. Nevertheless, their opioid-preferring activity does suggest the akuamma alkaloids provide distinct scaffolds from which novel opioids with unique pharmacologic properties and therapeutic utility can be developed.

Synthetic Studies of Neoclerodane Diterpenes from Salvia divinorum: Design, Synthesis, and Evaluation of Analogues with Improved Potency and G-protein Activation Bias at the µ-Opioid Receptor.

Previous structure-activity relationship (SAR) studies identified the first centrally acting, non-nitrogenous µ-opioid receptor (MOR) agonist, kurkinorin (1), derived from salvinorin A. In an effort to further probe the physiological effects induced upon activation of MORs with this nonmorphine scaffold, a variety of analogues were synthesized and evaluated in vitro for their ability to activate G-proteins and recruit ß-arrestin-2 upon MOR activation. Through these studies, compounds that are potent agonists at MORs and either biased toward ß-arrestin-2 recruitment or biased toward G-protein activation have been identified. One such compound, 25, has potent activity and selectivity at the MOR over KOR with bias for G-protein activation. Impressively, 25 is over 100× more potent than morphine and over 5× more potent than fentanyl in vitro and elicits antinociception with limited tolerance development in vivo. This is especially significant given that 25 lacks a basic nitrogen and other ionizable groups present in other opioid ligand classes.

Predictive compound accumulation rules yield a broad-spectrum antibiotic.

Most small molecules are unable to rapidly traverse the outer membrane of Gram-negative bacteria and accumulate inside these cells, making the discovery of much-needed drugs against these pathogens challenging. Current understanding of the physicochemical properties that dictate small-molecule accumulation in Gram-negative bacteria is largely based on retrospective analyses of antibacterial agents, which suggest that polarity and molecular weight are key factors. Here we assess the ability of over 180 diverse compounds to accumulate in Escherichia coli. Computational analysis of the results reveals major differences from the retrospective studies, namely that the small molecules that are most likely to accumulate contain an amine, are amphiphilic and rigid, and have low globularity. These guidelines were then applied to convert deoxynybomycin, a natural product that is active only against Gram-positive organisms, into an antibiotic with activity against a diverse panel of multi-drug-resistant Gram-negative pathogens. We anticipate that these findings will aid in the discovery and development of antibiotics against Gram-negative bacteria.

Synthetic Studies of Neoclerodane Diterpenes from Salvia divinorum: Identification of a Potent and Centrally Acting µ Opioid Analgesic with Reduced Abuse Liability.

Opioids are widely used to treat millions suffering from pain, but their analgesic utility is limited due to associated side effects. Herein we report the development and evaluation of a chemical probe exhibiting analgesia and reduced opioid-induced side effects. This compound, kurkinorin (5), is a potent and selective µ-opioid receptor (MOR) agonist (EC50 = 1.2 nM, >8000 µ/κ selectivity). 5 is a biased activator of MOR-induced G-protein signaling over ß-arrestin-2 recruitment. Metadynamics simulations of 5's binding to a MOR crystal structure suggest energetically preferred binding modes that differ from crystallographic ligands. In vivo studies with 5 demonstrate centrally mediated antinociception, significantly reduced rewarding effects, tolerance, and sedation. We propose that this novel MOR agonist may represent a valuable tool in distinguishing the pathways involved in MOR-induced analgesia from its side effects.

Holo Structure and Steady State Kinetics of the Thiazolinyl Imine Reductases for Siderophore Biosynthesis.

Thiazolinyl imine reductases catalyze the NADPH-dependent reduction of a thiazoline to a thiazolidine, a required step in the formation of the siderophores yersiniabactin (Yersinia spp.) and pyochelin (Pseudomonas aeruginosa). These stand-alone nonribosomal peptide tailoring domains are structural homologues of sugar oxidoreductases. Two closed structures of the thiazolinyl imine reductase from Yersinia enterocolitica (Irp3) are presented here: an NADP(+)-bound structure to 1.45 Å resolution and a holo structure to 1.28 Å resolution with NADP(+) and a substrate analogue bound. Michaelis-Menten kinetics were measured using the same substrate analogue and the homologue from P. aeruginosa, PchG. The data presented here support the hypothesis that tyrosine 128 is the likely general acid residue for catalysis and also highlight the phosphopantetheine tunnel for tethering of the substrate to the nonribosomal peptide synthetase module during assembly line biosynthesis of the siderophore.

Azaphilones inhibit tau aggregation and dissolve tau aggregates in vitro.

The aggregation of the microtubule-associated protein tau is a seminal event in many neurodegenerative diseases, including Alzheimer's disease. The inhibition or reversal of tau aggregation is therefore a potential therapeutic strategy for these diseases. Fungal natural products have proven to be a rich source of useful compounds having wide varieties of biological activities. We have previously screened Aspergillus nidulans secondary metabolites for their ability to inhibit tau aggregation in vitro using an arachidonic acid polymerization protocol. One aggregation inhibitor identified was asperbenzaldehyde, an intermediate in azaphilone biosynthesis. We therefore tested 11 azaphilone derivatives to determine their tau assembly inhibition properties in vitro. All compounds tested inhibited tau filament assembly to some extent, and four of the 11 compounds had the advantageous property of disassembling preformed tau aggregates in a dose-dependent fashion. The addition of these compounds to the tau aggregates reduced both the total length and number of tau polymers. The most potent compounds were tested in in vitro reactions to determine whether they interfere with tau's normal function of stabilizing microtubules (MTs). We found that they did not completely inhibit MT assembly in the presence of tau. These derivatives are very promising lead compounds for tau aggregation inhibitors and, more excitingly, for compounds that can disassemble pre-existing tau filaments. They also represent a new class of anti-tau aggregation compounds with a novel structural scaffold.

Synthesis and κ-opioid receptor activity of furan-substituted salvinorin A analogues.

The neoclerodane diterpene salvinorin A, found in the leaves of Salvia divinorum, is a potent κ-opioid receptor agonist, making it an attractive scaffold for development into a treatment for substance abuse. Although several successful semisynthetic studies have been performed to elucidate structure-activity relationships, the lack of analogues with substitutions to the furan ring of salvinorin A has prevented a thorough understanding of its role in binding to the κ-opioid receptor. Herein we report the synthesis of several salvinorin A derivatives with modified furan rings. Evaluation of these compounds in a functional assay indicated that sterically less demanding substitutions are preferred, suggesting the furan ring is bound in a congested portion of the binding pocket. The most potent of the analogues successfully reduced drug-seeking behavior in an animal model of drug-relapse without producing the sedation observed with other κ-opioid agonists.

Expanding the results of a high throughput screen against an isochorismate-pyruvate lyase to enzymes of a similar scaffold or mechanism.

Antibiotic resistance is a growing health concern, and new avenues of antimicrobial drug design are being actively sought. One suggested pathway to be targeted for inhibitor design is that of iron scavenging through siderophores. Here we present a high throughput screen to the isochorismate-pyruvate lyase of Pseudomonas aeruginosa, an enzyme required for the production of the siderophore pyochelin. Compounds identified in the screen are high nanomolar to low micromolar inhibitors of the enzyme and produce growth inhibition in PAO1 P. aeruginosa in the millimolar range under iron-limiting conditions. The identified compounds were also tested for enzymatic inhibition of Escherichia coli chorismate mutase, a protein of similar fold and similar chemistry, and of Yersinia enterocolitica salicylate synthase, a protein of differing fold but catalyzing the same lyase reaction. In both cases, subsets of the inhibitors from the screen were found to be inhibitory to enzymatic activity (mutase or synthase) in the micromolar range and capable of growth inhibition in their respective organisms (E. coli or Y. enterocolitica).

Palladium-catalyzed transformations of salvinorin A, a neoclerodane diterpene from Salvia divinorum.

Transformations that selectively modify the furan ring present in a variety of naturals products would be useful in the synthesis of biological probes but remain largely underexplored. The neoclerodane diterpene salvinorin A, isolated from Salvia divinorum, is an example of a furan-containing natural product. Following selective bromination of salvinorin A, Suzuki-Miyaura and Sonogashira couplings were accomplished in moderate to good yields without hydrolyzing the labile C-2 acetate or altering the stereochemistry of the epimerizable centers.