Chiral resolution of a caged xanthone and evaluation across a broad spectrum of breast cancer subtypes.

Racemic resolution of (+/-)-MAD28, a representative caged xanthone, was accomplished using (1S, 4R)-(-)-camphanic chloride as the chiral agent. Selective crystallization of the resulting diastereomers in acetonitrile produced, after hydrolysis, the pure enantiomers. Screening of racemic MAD28 and both enantiomers across a broad spectrum of breast cancer cell lines revealed that they: (a) are equipotent in each of the breast cancer subtypes examined; and (b) exhibit a higher degree of cytotoxicity against breast cancer cell lines of basal-like subtype and triple negative receptor status. The results support the notion that MAD28 and related caged xanthones are promising drug leads against chemoresistant and metastatic cancers.

Marine Spirotetronates: Biosynthetic Edifices That Inspire Drug Discovery.

Spirotetronates are actinomyces-derived polyketides that possess complex structures and exhibit potent and unexplored bioactivities. Due to their anticancer and antimicrobial properties, they have potential as drug hits and deserve further study. In particular, abyssomicin C and tetrocarcin A have shown significant promise against antibiotic-resistant S. aureus and tuberculosis, as well as for the treatment of various lymphomas and solid tumors. Improved synthetic routes to these compounds, particularly the class II spirotetronates, are needed to access sufficient quantities for structure optimization and clinical applications.

Gambogic acid identifies an isoform-specific druggable pocket in the middle domain of Hsp90ß.

Because of their importance in maintaining protein homeostasis, molecular chaperones, including heat-shock protein 90 (Hsp90), represent attractive drug targets. Although a number of Hsp90 inhibitors are in preclinical/clinical development, none strongly differentiate between constitutively expressed Hsp90ß and stress-induced Hsp90α, the two cytosolic paralogs of this molecular chaperone. Thus, the importance of inhibiting one or the other paralog in different disease states remains unknown. We show that the natural product, gambogic acid (GBA), binds selectively to a site in the middle domain of Hsp90ß, identifying GBA as an Hsp90ß-specific Hsp90 inhibitor. Furthermore, using computational and medicinal chemistry, we identified a GBA analog, referred to as DAP-19, which binds potently and selectively to Hsp90ß. Because of its unprecedented selectivity for Hsp90ß among all Hsp90 paralogs, GBA thus provides a new chemical tool to study the unique biological role of this abundantly expressed molecular chaperone in health and disease.

The Fe-S cluster-containing NEET proteins mitoNEET and NAF-1 as chemotherapeutic targets in breast cancer.

Identification of novel drug targets and chemotherapeutic agents is a high priority in the fight against cancer. Here, we report that MAD-28, a designed cluvenone (CLV) derivative, binds to and destabilizes two members of a unique class of mitochondrial and endoplasmic reticulum (ER) 2Fe-2S proteins, mitoNEET (mNT) and nutrient-deprivation autophagy factor-1 (NAF-1), recently implicated in cancer cell proliferation. Docking analysis of MAD-28 to mNT/NAF-1 revealed that in contrast to CLV, which formed a hydrogen bond network that stabilized the 2Fe-2S clusters of these proteins, MAD-28 broke the coordinative bond between the His ligand and the cluster's Fe of mNT/NAF-1. Analysis of MAD-28 performed with control (Michigan Cancer Foundation; MCF-10A) and malignant (M.D. Anderson-metastatic breast; MDA-MB-231 or MCF-7) human epithelial breast cells revealed that MAD-28 had a high specificity in the selective killing of cancer cells, without any apparent effects on normal breast cells. MAD-28 was found to target the mitochondria of cancer cells and displayed a surprising similarity in its effects to the effects of mNT/NAF-1 shRNA suppression in cancer cells, causing a decrease in respiration and mitochondrial membrane potential, as well as an increase in mitochondrial iron content and glycolysis. As expected, if the NEET proteins are targets of MAD-28, cancer cells with suppressed levels of NAF-1 or mNT were less susceptible to the drug. Taken together, our results suggest that NEET proteins are a novel class of drug targets in the chemotherapeutic treatment of breast cancer, and that MAD-28 can now be used as a template for rational drug design for NEET Fe-S cluster-destabilizing anticancer drugs.

Caged Garcinia Xanthones, a Novel Chemical Scaffold with Potent Antimalarial Activity.

Caged Garcinia xanthones (CGXs) constitute a family of natural products that are produced by tropical/subtropical trees of the genus Garcinia CGXs have a unique chemical architecture, defined by the presence of a caged scaffold at the C ring of a xanthone moiety, and exhibit a broad range of biological activities. Here we show that synthetic CGXs exhibit antimalarial activity against Plasmodium falciparum, the causative parasite of human malaria, at the intraerythrocytic stages. Their activity can be substantially improved by attaching a triphenylphosphonium group at the A ring of the caged xanthone. Specifically, CR135 and CR142 were found to be highly effective antimalarial inhibitors, with 50% effective concentrations as low as ∼10 nM. CGXs affect malaria parasites at multiple intraerythrocytic stages, with mature stages (trophozoites and schizonts) being more vulnerable than immature rings. Within hours of CGX treatment, malaria parasites display distinct morphological changes, significant reduction of parasitemia (the percentage of infected red blood cells), and aberrant mitochondrial fragmentation. CGXs do not, however, target the mitochondrial electron transport chain, the target of the drug atovaquone and several preclinical candidates. CGXs are cytotoxic to human HEK293 cells at the low micromolar level, which results in a therapeutic window of around 150-fold for the lead compounds. In summary, we show that CGXs are potent antimalarial compounds with structures distinct from those of previously reported antimalarial inhibitors. Our results highlight the potential to further develop Garcinia natural product derivatives as novel antimalarial agents.

Calcium phosphate-bearing matrices induce osteogenic differentiation of stem cells through adenosine signaling.

Synthetic matrices emulating the physicochemical properties of tissue-specific ECMs are being developed at a rapid pace to regulate stem cell fate. Biomaterials containing calcium phosphate (CaP) moieties have been shown to support osteogenic differentiation of stem and progenitor cells and bone tissue formation. By using a mineralized synthetic matrix mimicking a CaP-rich bone microenvironment, we examine a molecular mechanism through which CaP minerals induce osteogenesis of human mesenchymal stem cells with an emphasis on phosphate metabolism. Our studies show that extracellular phosphate uptake through solute carrier family 20 (phosphate transporter), member 1 (SLC20a1) supports osteogenic differentiation of human mesenchymal stem cells via adenosine, an ATP metabolite, which acts as an autocrine/paracrine signaling molecule through A2b adenosine receptor. Perturbation of SLC20a1 abrogates osteogenic differentiation by decreasing intramitochondrial phosphate and ATP synthesis. Collectively, this study offers the demonstration of a previously unknown mechanism for the beneficial role of CaP biomaterials in bone repair and the role of phosphate ions in bone physiology and regeneration. These findings also begin to shed light on the role of ATP metabolism in bone homeostasis, which may be exploited to treat bone metabolic diseases.

Cyclopenta[b]naphthalene cyanoacrylate dyes: synthesis and evaluation as fluorescent molecular rotors.

We describe the design, synthesis and fluorescent profile of a family of environment-sensitive dyes in which a dimethylamino (donor) group is conjugated to a cyanoacrylate (acceptor) unit via a cyclopenta[b]naphthalene ring system. This assembly satisfies the typical D-π-A motif of a fluorescent molecular rotor and exhibits solvatochromic and viscosity-sensitive fluorescence emission. The central naphthalene ring system of these dyes was synthesized via a novel intramolecular dehydrogenative dehydro-Diels-Alder (IDDDA) reaction that permits incorporation of the donor and acceptor groups in variable positions around the aromatic core. A bathochromic shift of excitation and emission peaks was observed with increasing solvent polarity but the dyes exhibited a complex emission pattern with a second red emission band when dissolved in nonpolar solvents. Consistent with other known molecular rotors, the emission intensity increased with increasing viscosity. Interestingly, closer spatial proximity between the donor and the acceptor groups led to decreased viscosity sensitivity combined with an increased quantum yield. This observation indicates that structural hindrance of intramolecular rotation dominates when the donor and acceptor groups are in close proximity. The examined compounds give insight into how excited state intramolecular rotation can be influenced by both the solvent and the chemical structure.

Spirotetronate polyketides as leads in drug discovery.

The discovery of chlorothricin (1) defined a new family of microbial metabolites with potent antitumor antibiotic properties collectively referred to as spirotetronate polyketides. These microbial metabolites are structurally distinguished by the presence of a spirotetronate motif embedded within a macrocyclic core. Glycosylation at the periphery of this core contributes to the structural complexity and bioactivity of this motif. The spirotetronate family displays impressive chemical structures, potent bioactivities, and significant pharmacological potential. This review groups the family members based on structural and biosynthetic considerations and summarizes synthetic and biological studies that aim to elucidate their mode of action and explore their pharmacological potential.

Total Synthesis and Structural Revision of Antibiotic CJ-16,264.

The total synthesis and structural revision of antibiotic CJ-16,264 is described. Starting with citronellal, the quest for the target molecule featured a novel bis-transannular Diels-Alder reaction that casted stereoselectively the decalin system and included the synthesis of six isomers before demystification of its true structure.

Neurotrophic natural products: chemistry and biology.

Neurodegenerative diseases and spinal cord injury affect approximately 50 million people worldwide, bringing the total healthcare cost to over 600 billion dollars per year. Nervous system growth factors, that is, neurotrophins, are a potential solution to these disorders, since they could promote nerve regeneration. An average of 500 publications per year attests to the significance of neurotrophins in biomedical sciences and underlines their potential for therapeutic applications. Nonetheless, the poor pharmacokinetic profile of neurotrophins severely restricts their clinical use. On the other hand, small molecules that modulate neurotrophic activity offer a promising therapeutic approach against neurological disorders. Nature has provided an impressive array of natural products that have potent neurotrophic activities. This Review highlights the current synthetic strategies toward these compounds and summarizes their ability to induce neuronal growth and rehabilitation. It is anticipated that neurotrophic natural products could be used not only as starting points in drug design but also as tools to study the next frontier in biomedical sciences: the brain activity map project.

Excited State Rotational Freedom Impacts Viscosity Sensitivity in Arylcyanoamide Fluorescent Molecular Rotor Dyes.

The microviscosity of intracellular environments plays an important role in monitoring cellular function. Thus, the capability of detecting changes in viscosity can be utilized for the detection of different disease states. Viscosity-sensitive fluorescent molecular rotors are potentially excellent probes for these applications; however, the predictable relationships between chemical structural features and viscosity sensitivity are poorly understood. Here, we investigate a set of arylcyanoamide-based fluorescent probes and the effect of small aliphatic substituents on their viscosity sensitivity. We found that the location of the substituents and the type of π-network of the fluorophore can significantly affect the viscosity sensitivity of these fluorophores. Computational analysis supported the notion that the excited state rotational energy barrier plays a dominant role in the relative viscosity sensitivity of these fluorophores. These findings provide valuable insight into the design of molecular rotor-based fluorophores for viscosity measurement.

Illicium sesquiterpenes: divergent synthetic strategy and neurotrophic activity studies.

Majucin-type sesquiterpenes from Illicium sp., such as jiadifenolide (2), jiadifenin (3), and (1R,10S)-2-oxo-3,4-dehydroxyneomajucin (4, ODNM), possess a complex caged chemical architecture and remarkable neurotrophic activities. As such, they represent attractive small-molecule leads against various neurodegenerative diseases. We present an efficient, enantioselective, and unified synthesis of 2, 3, and 4 and designed analogues that diverge from tetracyclic key intermediate 7. The synthesis of 7 is highlighted by the use of an enantioselective Robinson annulation reaction (construction of the AB rings), a Pd-mediated carbomethoxylation reaction (construction of the C ring), and a one-pot oxidative reaction cascade (construction of the D ring). Evaluation of the neurotrophic activity of these compounds led to the identification of several highly potent small molecules that significantly enhanced the activity of nerve growth factor (NGF) in PC-12 cells. Moreover, efforts to define the common pharmacophoric motif suggest that substitution at the C-10 center significantly affects bioactivity, while the hemiketal moiety of 2 and 3 and the C-1 substitution might not be critical to the neurotrophic activity.

Novel selective inhibitors of aminopeptidases that generate antigenic peptides.

Endoplasmic reticulum aminopeptidases, ERAP1 and ERAP2, as well as Insulin regulated aminopeptidase (IRAP) play key roles in antigen processing, and have recently emerged as biologically important targets for manipulation of antigen presentation. Taking advantage of the available structural and substrate-selectivity data for these enzymes, we have rationally designed a new series of inhibitors that display low micromolar activity. The selectivity profile for these three highly homologous aminopeptidases provides a promising avenue for modulating intracellular antigen processing.

A-ring oxygenation modulates the chemistry and bioactivity of caged Garcinia xanthones.

Natural products of the caged Garcinia xanthones (CGX) family are characterized by a unique chemical structure, potent bioactivities and promising pharmacological profiles. We have developed a Claisen/Diels-Alder reaction cascade that, in combination with a Pd(0)-catalyzed reverse prenylation, provides rapid and efficient access to the CGX pharmacophore, represented by the structure of cluvenone. To further explore this pharmacophore, we have synthesized various A-ring oxygenated analogues of cluvenone and have evaluated their bioactivities in terms of growth inhibition, mitochondrial fragmentation, induction of mitochondrial-dependent cell death and Hsp90 client inhibition. We found that installation of an oxygen functionality at various positions of the A-ring influences significantly both the site-selectivity of the Claisen/Diels-Alder reaction and the bioactivity of these compounds, due to remote electronic effects.

A strategy toward the synthesis of C13-oxidized cembrenolides.

An efficient strategy for the construction of C13-oxidized cembrenolides is reported. Central to this strategy is the installation of the C13 hydroxyl group prior to cembrane macrocyclization (via formation of the C1-C2 bond), allowing access to both C13 alcohol epimers. The orientation of the C13 alcohol was found to influence the cyclization mode of the cembranolide scaffold upon furan oxidation, leading to motifs reminiscent to bipinnatolide F, bielschowskysin, and verrillin.

Synthesis of the tetracyclic core of Illicium sesquiterpenes using an organocatalyzed asymmetric Robinson annulation.

An enantioselective synthesis of the core framework of neurotrophic Illicium majucin-type sesquiterpenes is described here. This strategy is based on an organocatalyzed asymmetric Robinson annulation and provides an efficient approach for a diversity-oriented synthesis of Illicium natural products that holds remarkable therapeutic potential for neurodegenerative diseases.

Fluorescent molecular rotors as versatile in situ sensors for protein quantitation.

Accurate protein quantitation is essential for many cellular mechanistic studies. Existing technology relies on extrinsic sample evaluation that requires significant volumes of sample as well as addition of assay-specific reagents and importantly, is a terminal analysis. This study exploits the unique chemical features of a fluorescent molecular rotor that fluctuates between twisted-to-untwisted states, with a subsequent intensity increase in fluorescence depending on environmental conditions (e.g., viscosity). Here we report the development of a rapid, sensitive in situ protein quantitation method using ARCAM-1, a representative fluorescent molecular rotor that can be employed in both non-terminal and terminal assays.

Nature-inspired total synthesis of (-)-fusarisetin A.

A concise, protecting group-free total synthesis of (-)-fusarisetin A (1) was efficiently achieved in nine steps from commercially available (S)-(-)-citronellal. The synthetic approach was inspired by our proposed biosynthesis of 1. Key transformations of our strategy include a facile construction of the decalin moiety that is produced via a stereoselective IMDA reaction and a one-pot TEMPO-induced radical cyclization/aminolysis that forms the C ring of 1. Our route is amenable to analogue synthesis for biological evaluation.

Aminonaphthalene 2-cyanoacrylate (ANCA) probes fluorescently discriminate between amyloid-ß and prion plaques in brain.

A major challenge for diagnosing and monitoring the progression of amyloid-based diseases is the capability to distinguish between amyloid deposits that are associated with related, but distinctly different, diseases. Here, we demonstrate that aminonaphthalenyl 2-cyanoacrylate-based probes can fluorescently discriminate between different types of amyloid deposits in brain. The discriminating capability of these molecular rotors is due to the stabilization of the ground versus excited states of these probes as a function of the polarity of their microenvironment (i.e., within the binding pocket on the amyloid). This property makes it possible, for the first time, to estimate the inherent static relative permittivity (ε(0)) of the binding pocket of each amyloid within tissue. The capability to selectively follow the deposition of specific amyloids in tissue may provide important information for therapeutic development that is not readily accessible from currently available technology.

Subcellular localization and activity of gambogic acid.

The natural product gambogic acid (GA) has shown significant potential as an anticancer agent as it is able to induce apoptosis in multiple tumor cell lines, including multidrug-resistant cell lines, as well as displaying antitumor activity in animal models. Despite the fact that GA has entered phase I clinical trials, the primary cellular target and mode of action of this compound remain unclear, although many proteins have been shown to be affected by it. By thorough analysis of several cellular organelles, at both the morphological and functional levels, we demonstrate that the primary effect of GA is at the mitochondria. We found that GA induces mitochondrial damage within minutes of incubation at low-micromolar concentrations. Moreover, a fluorescent derivative of GA was able to localize specifically to the mitochondria and was displaced from these organelles after competition with unlabeled GA. These findings indicate that GA directly targets the mitochondria to induce the intrinsic pathway of apoptosis, and thus represents a new member of the mitocans.