Regioselective approach to colchiceine tropolone ring functionalization at C(9) and C(10) yielding new anticancer hybrid derivatives containing heterocyclic structural motifs.

The influence of base type, temperature, and solvent on regioselective C(9)/C(10) "click" modifications within the tropolone ring of colchiceine (2) is investigated. New ether derivatives of 2, bearing alkyne, azide, vinyl, or halide aryl groups enable assembly of the alkaloid part with heterocycles or important biomolecules such as saccharides, geldanamycin or AZT into hybrid scaffolds by dipolar cycloaddition (CuAAC) or Heck reaction. Compared to colchicine (1) or colchiceine (2), ether congeners, as e.g. 3e [IC50s(3e) ∼ 0.9 nM], show improved or similar anticancer effects, whereby the bulkiness of the substituents and the substitution pattern of the tropolone proved to be essential. Biological studies reveal that expanding the ether arms by terminal basic heterocycles as quinoline or pyridine, decreases the toxicity in HDF cells at high anticancer potency (IC50s ∼ 1-2 nM). Docking of ether and hybrid derivatives into the colchicine pocket of αGTP/ß tubulin dimers reveals a relationship between the favourable binding mode and the attractive anticancer potency.

Discoveries and Syntheses of Highly Potent Antimalarial Troponoids.

Novel derivatives of puberulic acid were synthesized and their antimalarial properties were evaluated in vitro against the Plasmodium falciparum K1 parasite strain, cytotoxicity against a human diploid embryonic cell line MRC-5, and in vivo efficacy using a Plasmodium berghei-infected mouse model. From previous information that three hydroxy groups on the tropone framework were essential for antimalarial activity, we converted the carboxylic acid moiety into the corresponding esters, amides, and ketones. These derivatives showed antimalarial activity against chloroquine-resistant Plasmodium in vitro equivalent to puberulic acid. We identified that the pentane-3-yl ester, cyclohexyl ester, iso-butyl ketone, cyclohexyl methyl ketone all show an especially potent antiparasitic effect in vivo at an oral dose of 15 mg/kg without any apparent toxicity. These esters were more effective than the existing commonly used antimalarial drug, artesunate.

Total Synthesis and Computational Investigations of Sesquiterpene-Tropolones Ameliorate Stereochemical Inconsistencies and Resolve an Ambiguous Biosynthetic Relationship.

The sesquiterpene-tropolones belong to a distinctive structural class of meroterpene natural products with impressive biological activities, including anticancer, antifungal, antimalarial, and antibacterial. In this article, we describe a concise, modular, and cycloaddition-based approach to a series of sesquiterpene mono- and bistropolones, including (-)-epolone B, (+)-isoepolone B, (±)-dehydroxypycnidione, and (-)-10-epi-pycnidione. Alongside the development of a general strategy to access this unique family of metabolites were computational modeling studies that justified the diastereoselectivity observed during key cycloadditions. Ultimately, these studies prompted stereochemical reassignments of the pycnidione subclass and shed additional light on the biosynthesis of these remarkable natural products.

Thallium(I) Tropolonates: Synthesis, Structure, Spectral Characteristics, and Antimicrobial Activity Compared to Lead(II) and Bismuth(III) Analogues.

Synthesis, single-crystal X-ray determination diffraction and FT-IR, NMR (1H, 13C, 19F and 205Tl), UV-vis, and luminescence spectra characteristics were described for series of thallium(I) compounds: thallium(I) triflate (Tl(OTf)), 1:1 co-crystals of thallium(I) triflate and tropolone (Htrop), Tl(OTf)·Htrop, as well as simple thallium(I) chelates: Tl(trop) (1), Tl(5-metrop) (2), Tl(hino) (3), with Htrop, 5-methyltropolone (5-meHtrop), 4-isopropyltropolone (hinokitiol, Hhino), respectively, and additionally more complex {Tl@[Tl(hino)]6}(OTf) (4) compound. Comparison of their antimicrobial activity with selected lead(II) and bismuth(III) analogs and free ligands showed that only bismuth(III) complexes demonstrated significant antimicrobial activity, from two- to fivefold larger than the free ligands.

Amide-containing α-hydroxytropolones as inhibitors of hepatitis B virus replication.

The Hepatitis B Virus (HBV) ribonuclease H (RNaseH) is a promising but unexploited drug target. Here, we synthesized and analyzed a library of 57 amide-containing α-hydroxytropolones (αHTs) as potential leads for HBV drug development. Fifty percent effective concentrations ranged from 0.31 to 54 µM, with selectivity indexes in cell culture of up to 80. Activity against the HBV RNaseH was confirmed in semi-quantitative enzymatic assays with recombinant HBV RNaseH. The compounds were overall poorly active against human ribonuclease H1, with 50% inhibitory concentrations of 5.1 to >1,000 µM. The αHTs had modest activity against growth of the fungal pathogen Cryptococcus neoformans, but had very limited activity against growth of the Gram - bacterium Escherichia coli and the Gram + bacterium Staphylococcus aureus, indicating substantial selectivity for HBV. A molecular model of the HBV RNaseH templated against the Ty3 RNaseH was generated. Docking the compounds to the RNaseH revealed the anticipated binding pose with the divalent cation coordinating motif on the compounds chelating the two Mn++ ions modeled into the active site. These studies reveal that that amide αHTs can be strong, specific HBV inhibitors that merit further assessment toward becoming anti-HBV drugs.

Structure and anticancer activities of four Cu(ii) complexes bearing tropolone.

Agents inducing apoptosis and autophagic death could be effective chemotherapeutic drugs. In this work, four novel Cu(ii) complexes formulated as [CuL2] (1), [Cu(phen)LCl]·0.5H2O (2), [Cu2(MQ)2L2] (3) and [Cu(2,2'-bpy)LCl]·H2O (4) (phen = 1,10-Phenanthroline, HMQ = 8-hydroxy-2-methylquinoline, 2,2'-bpy = 2,2'-bipyridine) were prepared from the reactions of copper(ii) chloride with tropolone (HL) in the absence or presence of different ancillary ligands. The solution state structures of 1, 2 and 4 agree well with their solid state structures. Complex 3 presents a dimer structure in the solid state, however, a monomer structure in the solution state. It was shown that all of these complexes are stable under experimental conditions and bind to DNA in an intercalative mode with the binding constant Kb values of 1.05 × 103, 2.57 × 103, 2.53 × 103 and 2.26 × 103 L mol-1 for complexes 1, 2, 3 and 4, respectively. The anti-proliferative tests against cultured human cancer cell lines (A549, Bel-7402, MGC80-3, T24, SK-OV-3, and NCI-H460) in vitro revealed cytotoxic activities for these complexes, which are much better than those for all ligands in these complexes, as well as that for cis-platin. After a careful comparison, the cytotoxic activity of complex 2 against MGC80-3 cells in vitro (IC50 = 3.5 ± 0.9 µM for 2 and 18.0 ± 1.2 for cis-platin) was further investigated in detail as an example. 2 induces the apoptosis of MGC80-3 through a caspase-dependent mitochondrion pathway and can also induce autophagy, which revealed a certain anticancer activity for complex 2.

Maltol- and Allomaltol-Derived Oxidopyrylium Ylides: Methyl Substitution Pattern Kinetically Influences [5 + 3] Dimerization versus [5 + 2] Cycloaddition Reactions.

Oxidopyrylium ylides are useful intermediates in synthetic organic chemistry because of their capability of forming structurally complex cycloadducts. They can also self-dimerize via [5 + 3] cycloaddition, which is an oft-reported side reaction that can negatively impact [5 + 2] cycloadduct yields and efficiency. In select instances, these dimers can be synthesized and used as the source of oxidopyrylium ylide, although the generality of this process remains unclear. Thus, how the substitution pattern governs both dimerization and cycloaddition reactions is of fundamental interest to probe factors to regulate them. The following manuscript details our findings that maltol-derived oxidopyrylium ylides (i.e., with ortho methyl substitution relative to oxide) can be trapped prior to dimerization more efficiently than the regioisomeric allomaltol-derived ylide (i.e., with a para methyl substitution relative to oxide). Density functional theory studies provide evidence in support of a sterically (kinetically) controlled mechanism, whereby gauche interactions between appendages of the approaching maltol-derived ylides are privileged by higher barriers for dimerization and thus are readily intercepted by dipolarophiles via [5 + 2] cycloadditions.

Preparation of 6-aminocyclohepta-2,4-dien-1-one Derivatives via Tricarbonyl(tropone)iron.

aza-Michael adducts of tricarbonyl(tropone)iron are synthesized by two different methods. Primary aliphatic amines and cyclic secondary amines participate in a direct aza-Michael reaction with tricarbonyl(tropone)iron under solvent-free conditions. Less nucleophilic aniline derivatives and more hindered secondary amines add efficiently to the cationic tropone complex formed by protonation of tricarbonyl(tropone)iron. While the protocol utilizing the cationic complex is less efficient overall for accessing the aza-Michael adducts than the direct, solvent-free addition to the neutral complex, it allows the use of a broader range of amine nucleophiles. Following protection of the amine of the aza-Michael adduct as a tert-butyl carbamate, the diene is decomplexed from the iron tricarbonyl fragment upon treatment with cerium(IV) ammonium nitrate to provide derivatives of 6-aminocyclohepta-2,4-dien-1-one. These products can serve as precursors to diverse compounds containing a seven-membered carbocyclic ring. Because the demetallation requires protection of the amine as a carbamate, the aza-Michael adducts of secondary amines cannot be decomplexed using the protocol described here.

Troponoid Atropisomerism: Studies on the Configurational Stability of Tropone-Amide Chiral Axes.

Configurationally stable, atropisomeric motifs are an important structural element in a number of molecules, including chiral ligands, catalysts, and molecular devices. Thus, understanding features that stabilize chiral axes is of fundamental interest throughout the chemical sciences. The following details the high rotational barriers about the Ar-C(O) bond of tropone amides, which significantly exceed those of analogous benzamides. These studies are supported by both experimental and computational rotational barrier measurements. We also report the resolution of an axially chiral α-hydroxytropolone amide into its individual atropisomers, and demonstrate its configurational stability at physiological pH and temperatures over 24 h.

Tropolone-Conjugated DNA: Fluorescence Enhancement in the Duplex.

Tropolone (2-hydroxycyclohepta-2,4,6-triene-1-one and tautomer) is a non-benzenoid bioactive natural chromophore with pH-dependent fluorescence character and extraordinary metal binding affinities, especially with transition-metal ions Cu2+ /Zn2+ /Ni2+ . This report describes the syntheses and biophysical studies of a new tropolonyl thymidine [(4(5)-hydroxy-5(4)-oxo-5(4)H-cyclohepta-1,3,6-trienyl)thymidine] (tr-T) nucleoside and of corresponding tropolone-conjugated DNA oligonucleotides that form B-form DNA duplex structures with a complementary DNA strand, although their duplex structures are less stable than that of the control. Furthermore, the stabilities of those DNA duplex structures are lowered by the presence of increasing numbers of tr-T residue or by decreasing pH of their environments. Most importantly, these duplex structures are made fluorescent because of the presence of the tropolone moieties conjugated to the thymidine residues. The fluorescence behavior of those duplex structures exhibits pH dependence, with stronger fluorescence at lower pH and weaker fluorescence at high pH. Importantly, the fluorescence characters of tr-DNA oligonucleotides are significantly enhanced by nearly threefold after duplex structure formation with their complementary control DNA oligonucleotide. Further, the fluorescence behavior of these tr-DNA duplex structures is also dependent on the pH conditions. Hence, tropolonyl-conjugated DNA represents a class of new fluorescent analogues that might be be employed for sensing DNA duplex formation and provide opportunities to improve fluorescence properties further.

Design, synthesis, and biologic evaluation of novel galloyl derivatives as HIV-1 RNase H inhibitors.

Human immunodeficiency virus (HIV) reverse transcriptase (RT)-associated ribonuclease H (RNase H) remains as the only enzyme encoded within the viral genome not targeted by current antiviral drugs. In this work, we report the design, synthesis, and biologic evaluation of a novel series of galloyl derivatives with HIV-1 RNase H inhibitory activity. Most of them showed IC50 s at sub- to low-micromolar concentrations in enzymatic assays. The most potent compound was II-25 that showed an IC50 of 0.72 ± 0.07 µM in RNase H inhibition assays carried out with the HIV-1BH10 RT. II-25 was 2.8 times more potent than ß-thujaplicinol in these assays. Interestingly, II-25 and other galloyl derivatives were also found to inhibit the HIV IN strand transfer activity in vitro. Structure-activity relationships (SAR) studies and molecular modeling analysis predict key interactions with RT residues His539 and Arg557, while providing helpful insight for further optimization of selected compounds.

Fluorous-Phase Approach to α-Hydroxytropolone Synthesis.

α-Hydroxytropolones (αHTs) are troponoids that demonstrate inhibition against an array of therapeutically significant targets, making them potential drug leads for several human diseases. We have utilized a recently discovered one-pot three-component oxidopyrylium cycloaddition in a solid-supported synthesis of αHTs. Though the procedure is time efficient and generates assay-ready molecules, the system suffers from low yields and an inability to perform reaction modifications on resin-bound intermediates. In order to combat these issues with the solid-phase platform, we incorporated fluorous tags into our synthetic route. Through the implementation of fluorous phase chemistry, we demonstrate a substantial increase in the overall yield of αHTs, as well as an ability to execute metal-catalyzed cross coupling and amide coupling on fluorous tagged intermediates. We also show that tagged molecules can be separated from nonfluorous impurities, and vice versa, by utilizing fluorous liquid-liquid and solid-phase extractions. Hence, these proof-of-principle investigations describe the viability of a fluorous phase approach to αHT synthesis and its potential to serve as a combinatorial technique to produce structurally diverse substrates.

Synthesis, structure-activity relationships, and mechanistic studies of 5-arylazo-tropolone derivatives as novel xanthine oxidase (XO) inhibitors.

Xanthine oxidase (XO) is an enzyme that contains molybdenum at the active site and catalyzes the oxidation of purine bases to uric acid. Even though XO inhibitors are widely used for the treatment of hyperuricemia and gout, only very few such compounds are clinically used as drugs for the treatment of these diseases. Given the unique physicochemical properties of tropolone, i.e., its chelating effect and the pKa value that is similar to that of carboxylic acid, we have synthesized 22 5-arylazotropolone derivatives as potential XO inhibitors. In vitro enzyme-inhibitory assays for XO revealed that 3-nitro derivative 1j showed the most potent XO inhibitory activity, which is by one order of magnitude more potent than allopurinol. An enzyme-kinetic study revealed that 1j inhibited the production of uric acid by XO both competitively and non-competitively. A docking-simulation study of 1j with XO suggested that the carbonyl and hydroxyl groups of the tropolone ring interact with the hydroxy group that acts as a ligand for molybdenum and the amino acid residues around the active site of XO.

Synthesis and biological assessment of 3,7-dihydroxytropolones.

3,7-Dihydroxytropolones (3,7-dHTs) are highly oxygenated troponoids that have been identified as lead compounds for several human diseases. To date, structure-function studies on these molecules have been limited due to a scarcity of synthetic methods for their preparation. New synthetic strategies towards structurally novel 3,7-dHTs would be valuable in further studying their therapeutic potential. Here we describe the successful adaptation of a [5 + 2] oxidopyrilium cycloaddition/ring-opening for 3,7-dHT synthesis, which we apply in the synthesis of a plausible biosynthetic intermediate to the natural products puberulic and puberulonic acid. We have also tested these new compounds in several biological assays related to human immunodeficiency virus (HIV), hepatitis B virus (HBV) and herpes simplex virus (HSV) in order to gain insight into structure-functional analysis related to antiviral troponoid development.

Antimalarial troponoids, puberulic acid and viticolins; divergent synthesis and structure-activity relationship studies.

Divergent synthesis of antimalarial troponoids, including naturally occurring compounds, some of which were identified and isolated by our group, has been achieved utilizing the total synthetic route of puberulic acid. Structure-activity relationships of natural products and simple troponoids inspired us to explore more detailed properties of this class of compounds. Access to new derivatives was facilitated through intermediate compounds generated during the total synthesis of puberulic acid by a stepwise oxidation-aromatization sequence to provide 7-hydroxytropolones and bromination for conversion of the carboxylic acid moiety. The first total synthesis of viticolin A, as well as the synthesis of different methyl-substituted derivatives, has also been achieved. In vitro antimalarial activity and cytotoxicity of novel derivatives were evaluated and fundamental information to facilitate the discovery of more promising antimalarials was obtained.

Gold-Catalyzed Synthesis of Tropone and Its Analogues via Oxidative Ring Expansion of Alkynyl Quinols.

A new and convenient strategy for the synthesis of functionalized tropone derivatives based on the gold-catalyzed oxidative ring expansion of alkynyl quinols has been developed. The reaction proceeds via gold-catalyzed highly regioselective oxidation followed by 1,2-migration of a vinyl or phenyl group. Extension of this chemistry allows ready access to various seven- or six-membered ring systems such as benzotropones, benzooxepines, phenanthrenes, and quinolin-2(1H)-ones.

The discovery of a novel route to highly substituted α-tropolones enables expedient entry to the core of the gukulenins.

A simple and general method for the synthesis of highly substituted α-tropolone ethers that allows rapid access to the bis(tropolone) core of the antiproliferative metabolites (-)-gukulenins A and F (3, 4) is described. The reaction proceeds by thermolytic opening of gem-dibromobicyclo[4.1.0]heptane intermediates, which are readily accessed from simple starting materials. Mechanistic studies suggest the reaction proceeds via an autocatalytic process mediated by methyl hypobromite. This synthetic sequence allows access to a broad array of highly substituted α-tropolones.

Ni(NHC)]-catalyzed cycloaddition of diynes and tropone: apparent enone cycloaddition involving an 8π insertion.

A Ni/N-heterocyclic carbene catalyst couples diynes to the C(α)-C(ß) double bond of tropone, a type of reaction that is unprecedented for metal-catalyzed cycloadditions with aromatic tropone. Many different diynes were efficiently coupled to afford [5-6-7] fused tricyclic products, while [5-7-6] fused tricyclic compounds were obtained as minor byproducts in a few cases. The reaction has broad substrate scope and tolerates a wide range of functional groups, and excellent regioselectivity is found with unsymmetrical diynes. Theoretical calculations show that the apparent enone cycloaddition occurs through a distinctive 8π insertion of tropone. The initial intramolecular oxidative cyclization of diyne produces the nickelacyclopentadiene intermediate. This intermediate undergoes an 8π insertion of tropone, and subsequent reductive elimination generates the [5-6-7] fused tricyclic product. This initial product undergoes two competing isomerizations, leading to the observed [5-6-7] and [5-7-6] fused tricyclic products.

A concise total synthesis of puberulic acid, a potent antimalarial agent.

Efficient and practical total synthesis of puberulic acid has been accomplished via 8 steps, with 54% overall yield, and only two C-C bond formations, without the introduction of oxygen atoms into the core skeleton. Construction of the tropolone framework as the key transformation was achieved by multi-tandem oxidation of the aliphatic-triol, from D-(+)-galactose as the starting material.

Oxidation of cyanobenzocycloheptatrienes: Synthesis, photooxygenation reaction and carbonic anhydrase isoenzymes inhibition properties of some new benzotropone derivatives.

The oxidation of some cyanocycloheptatrienes with CrO3 and pyridine was investigated and a few new nitrile functionalised benzotropone derivatives were obtained. Photooxygenation reaction of these products was also studied. The structures of the formed products were determined on the basis of NMR spectroscopy and the formation mechanism of unusual products was discussed. Human carbonic anhydrase isoenzymes I, and II (hCA I and hCA II) inhibition properties of nitrile functionalized new benzotropone derivatives were also studied. Both CA isozymes were inhibited in the low micromolar range by these nitrile functionalized benzotropone analogues. The newly synthesized benzotropone derivatives showed inhibition constants in the sub-micromolar range (2.51-4.06µM). The best hCA I inhibition was observed in 5H-benzocycloheptene-7-carbonitrile (Ki: 2.88±0.86µM). On the other hand, 5-oxo-5H-benzocycloheptatriene-7-carbonitrile showed the powerful inhibitory effect against hCA II (Ki: 2.51±0.34µM).