Unambiguous Identification of β‐Tubulin as the Direct Cellular Target Responsible for the Cytotoxicity of Chalcone by Photoaffinity Labeling

Chalcone is a simple and potentially privileged structure in medicinal chemistry with a diverse repertoire of biological activities, among which cytotoxicity is of particular interest. The sharp structure–activity relationship (SAR) for chalcone's cytotoxicity suggests structure‐specific target interactions. Despite the numerous putative targets proposed, evidence for direct target interactions in cells is unavailable. In this study, guided by the sharp cytotoxic SAR, we developed a cytotoxic chalcone‐based photoaffinity labeling (PAL) probe, (E)‐3‐(3‐azidophenyl)‐1‐[3,5‐dimethoxy‐4‐(prop‐2‐yn‐1‐yloxy)phenyl]‐2‐methylprop‐2‐en‐1‐one (C95; IC₅₀: 0.38±0.01 μm ), along with two structurally similar non‐cytotoxic probes. These probes were used to search for the direct cellular target responsible for chalcone's cytotoxicity through intact cell‐based PAL experiments, in which β‐tubulin was identified to specifically interact with the cytotoxic probe (i.e., C95) but not the non‐cytotoxic probes. A set of phenotypical and biochemical assays further reinforced β‐tubulin as the cytotoxic target of chalcones. Peptide mass quantitation by mass spectrometric analysis revealed one peptide potentially labeled by C95, providing information on chalcone's binding site on β‐tubulin.     

Unusual η2‐Allene Osmacycle with Apoptotic Properties

Screening of a library of structurally unusual osmacyclic complexes for their antiproliferate properties in HeLa cells led to the discovery of a highly cytotoxic η²‐allene osmacycle. In this remarkably stable complex, osmium constitutes part of a metallacycle through the formation of a σ‐bond to a carbon in combination with coordination to an allene moiety. The osmacycle strongly induces apoptosis in Burkitt‐like lymphoma cells at submicromolar concentrations. The reduction of the mitochondrial membrane potential, the induction of DNA fragmentation, and the activation of caspases‐9 and ‐3 reveal that programmed cell death occurs through the intrinsic mitochondrial pathway. From the lipophilic and cationic nature of the osmacycle, in addition to a low oxidation potential (E_1/2=+0.27 V vs. Fc/Fc⁺, Fc=ferrocene) it is proposed that mitochondria are the cellular target where oxidative decomposition initiates apoptosis.     

Synthesis of a Fluorogenic Analogue of Sphingosine‐1‐Phosphate and Its Use to Determine Sphingosine‐1‐Phosphate Lyase Activity

Illuminating an ER enzyme : We report on the design and synthesis of a fluorogenic chemical sensor ( 1 ) to measure sphingosine‐1‐phosphate lyase activity in high‐throughput screening formats, as well as its validation using lyase knockout (Sgpl1?/?) cells.

     

Synthesis and Properties of Alkoxyethyl β‐d‐Xylopyranoside

In order to improve the water solubility of sugar‐based surfactants, alkyl β‐d‐ xylopyranosides, novel sugar‐based surfactants, 1,2‐trans alkoxyethyl β‐d‐ xylopyranosides, with alkyl chain length n = 6–12 were stereoselectively prepared by the trichloroacetimidate method. Their properties including hydrophilic–lipophilic balance (HLB) number, water solubility, surface tension, emulsification, foamability, thermotropic liquid crystal, and hygroscopicity were investigated. The results indicated that their HLB number decreased with increase of alkyl chain, the water solubility improved since the hydrophilic oxyethene (─OCH₂CH₂─) fragment was introduced. The dissolution process was entropy driven at 25–45 °C for alkyl chain length n = 6–10. Octyloxyethyl β‐d‐ xylopyranoside had the best foaming ability. Nonyloxyethyl β‐d‐ xylopyranoside had the best foam stability and the emulsifying ability was better in toluene/water system than in rapeseed oil/water system. The surface tension of in aqueous solution dropped to 27.8 mN m^?1 at the critical micelle concentration, and it also showed the most distinct thermotropic liquid phases with cross pattern texture upon heating and the fan schlieren texture on cooling. Hexyloxyethyl β‐d‐ xylopyranoside possessed the strongest hygroscopicity. Based on the effective improvement of water solubility, the prepared alkoxyethyl β‐d‐ xylopyranosides showed excellent surface activity and are expected to develop their practical application as a class of novel sugar‐based surfactants.     

A Ring‐Closing Metathesis Approach to Cyclic α,β‐Dehydroamino Acids

A comprehensive study on the synthesis and ring‐closing metathesis (RCM) of α,β‐dehydroamino acids is described. This sequence has led to the formation of a range of biologically relevant functionalized nitrogen heterocycles. The incorporation of chiral building blocks in the RCM precursors eventually resulted in the formation of optically active 4‐substituted cyclic dehydroamino acids. In addition, olefin isomerization under metathesis conditions was observed for a number of compounds, which could be successfully inhibited either by the introduction of allylic substituents or by the addition of a ruthenium hydride scavenger.     

Design of a Highly Selective and Potent Class of Non‐planar Estrogen Receptor β Agonists

Selective activation of the estrogen receptor β (ERβ) could be a safe approach to hormone replacement therapy for both women and men, in contrast to the estrogens currently used for women which activate both ERβ and ERα, occasionally causing severe side effects. The selective ERβ agonist AC‐131 has shown efficacy in animal models of Parkinson’s disease and neuropathic pain. With the use of AC‐131 as template, herein we report the discovery, synthesis, and structure–activity relationship (SAR) study of a new class of dihydrobenzofurans as potent and selective ERβ agonists. The SAR was established by enantioselective synthesis, molecular modeling, and whole‐cell‐based functional assays. The most potent diastereomer, cis‐ 10 ‐SR, was shown to have an EC₅₀ value of <1 nM , potency 100‐fold higher than that of AC‐131. Even more interestingly, compound trans‐ 10 ‐SS exhibited 1000‐fold ERβ/ERα selectivity while still maintaining good potency (～10 nM ). In addition, trans‐ 10 ‐SS showed only partial agonist activity (30–60 % Eff.) toward ERα at 10 μM . This unprecedented selectivity could be rationalized by molecular modeling. Compound trans‐ 10 ‐SS appears to be the first molecule to take advantage of both conservative amino acid differences found in the α‐ and β‐faces of the binding cavities of ERα and ERβ.     

Synthesis of poly(β‐pinene)‐g‐polystyrene from allylic brominated poly(β‐pinene)

Poly(β‐pinene) was brominated by N‐bromosuccinimide on the allylic carbons. Then the brominated product was activated by AlEt₂Cl to initiate the polymerization of styrene to give a β‐pinene/styrene graft copolymer. AlEt₂Cl was selected because it alone could not initiate the polymerization of styrene. The obtained graft copolymer was characterized by GPC, ¹H‐NMR, and DSC measurements, respectively. © 2000 John Wiley & Sons, Inc. J Appl Polym Sci 75: 599–603, 2000     

Synthesis of an Enantiocomplementary Catalyst of β‐Isocupreidine (β‐ICD) from Quinine

Compound 20 , a pseudoenantiomer of β‐isocupreidine (β‐ICD), was synthesized from quinine employing a Barton reaction of nitrosyl ester 13 and acid‐catalyzed cyclization of carbinol 18 as key steps. The Baylis–Hillman reaction of benzaldehyde, p‐nitrobenzaldehyde, and hydrocinnamaldehyde with 1,1,1,3,3,3‐hexafluoroisopropyl acrylate (HFIPA) using 20 as a chiral amine catalyst was found to give the corresponding S‐enriched adducts in high optical purity (>91% ee) in contrast to the β‐ICD‐catalyzed reaction which affords R‐enriched adducts. This result suggests that compound 20 can serve as an enantiocomplementary catalyst of β‐ICD in the asymmetric Baylis–Hillman reaction of aldehydes with HFIPA.     

Chemistry and Pharmacology of Nicotinic Ligands Based on 6‐[5‐(Azetidin‐2‐ylmethoxy)pyridin‐3‐yl]hex‐5‐yn‐1‐ol (AMOP‐H‐OH) for Possible Use in Depression

AMOP‐H‐OH (sazetidine‐A; 6‐[5‐(azetidin‐2‐ylmethoxy)pyridin‐3‐yl]hex‐5‐yn‐1‐ol) and some sulfur‐bearing analogues were tested for their activities in vitro against human α4β2‐, α4β4‐, α3β4*‐ and α1*‐nicotinic acetylcholine receptors (nAChRs). AMOP‐H‐OH was also assessed in an antidepressant efficacy model. AMOP‐H‐OH and some of its analogues have high potency and selectivity for α4β2‐nAChRs over other nAChR subtypes. Effects are manifested as partial agonism, perhaps reflecting selectivity for high sensitivity (α4)₃(β2)₂‐nAChRs. More prolonged exposure to AMOP‐H‐OH and its analogues produces inhibition of subsequent responses to acute challenges with full nicotinic agonists, again selectively for α4β2‐nAChRs over other nAChR subtypes. The inhibition is mediated either via antagonism or desensitization of nAChR function, but the degree of inhibition of α4β2‐nAChRs is limited by the partial agonist activity of the drugs. Certain aspects of the in vitro pharmacology suggest that AMOP‐H‐OH and some of its analogues have a set of binding sites on α4β2‐nAChRs that are distinct from those for full agonists. The in vitro pharmacological profile suggests that peripheral side effects of AMOP‐H‐OH or its analogues would be minimal and that their behavioral effects would be dominated by central nAChR actions. AMOP‐H‐OH also has profound and high potency antidepressant‐like effects in the forced swim test. The net action of prolonged exposure to AMOP‐H‐OH or its analogues, as for nicotine, seems to be a selective decrease in α4β2‐nAChR function. Inactivation of nAChRs may be a common neurochemical endpoint for nicotine dependence, its treatment, and some of its manifestations, including relief from depression.     

Structural characterization of α‐ and β‐nucleated isotactic polypropylene

Modification of isotactic polypropylene (iPP) with two nucleation agents, namely 1,3:24‐bis(3,4‐dimethylobenzylideno) sorbitol (DMDBS) (α‐nucleator) and N, N′‐dicyclohexylo‐2,6‐naphthaleno dicarboxy amide (NJ) (β‐nucleator), leads to significant changes of the structure, morphology and properties. Both nucleating agents cause an increase in the crystallization temperature. The efficiency determined in a self‐nucleation test is 73.4 % for DMDBS and 55.9 % for NJ. The modification with NJ induces the creation of the hexagonal β‐form of iPP. The addition of DMDBS lowers the haze of iPP while the presence of NJ increases the haze. Copyright © 2004 Society of Chemical Industry     

Copper(I)‐Catalyzed Stereoselective Synthesis of (E)‐α‐Alkynyl‐α,β‐unsaturated Esters from a Terminal Alkyne,Diazoesters and Aldehydes

A new synthetic method for α‐alkynyl‐α,β‐unsaturated esters is presented herein. The method is based on a copper(I)‐catalyzed three‐component reaction of a terminal alkyne, diazoesters and aldehydes. The reaction is featured by mild conditions, high yields and excellent stereoselectivity. Cu(I) carbene migratory insertion is proposed as the key step in the transformation.

     

Synthesis of S‐Adenosyl‐L‐homocysteine Capture Compounds for Selective Photoinduced Isolation of Methyltransferases

Understanding the interplay of different cellular proteins and their substrates is of major interest in the postgenomic era. For this purpose, selective isolation and identification of proteins from complex biological samples is necessary and targeted isolation of enzyme families is a challenging task. Over the last years, methods like activity‐based protein profiling (ABPP) and capture compound mass spectrometry (CCMS) have been developed to reduce the complexity of the proteome by means of protein function in contrast to standard approaches, which utilize differences in physical properties for protein separation. To isolate and identify the subproteome consisting of S‐adenosyl‐L ‐methionine (SAM or AdoMet)‐dependent methyltransferases (methylome), we developed and synthesized trifunctional capture compounds containing the chemically stable cofactor product S‐adenosyl‐L ‐homocysteine (SAH or AdoHcy) as selectivity function. SAH analogues with amino linkers at the N6 or C8 positions were synthesized and attached to scaffolds containing different photocrosslinking groups for covalent protein modification and biotin for affinity isolation. The utility of these SAH capture compounds for selective photoinduced protein isolation is demonstrated for various methyltransferases (MTases) acting on DNA, RNA and proteins as well as with Escherichia coli cell lysate. In addition, they can be used to determine dissociation constants for MTase–cofactor complexes.     

Synthesis and Derivatization of Substituted (R)‐ and (S)‐ C‐Allylglycines

Various (R)‐ and (S)‐C‐allylglycine derivatives were synthesized by means of an auxiliary controlled diastereoselective aza‐Claisen rearrangement. Starting from (S)‐configured auxiliaries derived from optically active proline, an aza‐Claisen rearrangement enabled us to synthesize α(R)‐configured γ,δ‐unsaturated amides. Since (R)‐allylglycine derivatives could be directly generated by reacting N‐allylproline derivatives and various protected glycine fluorides, the corresponding (S)‐enantiomers were built‐up via an initial α‐chloroacetyl chloride rearrangement and a subsequent chloride azide substitution with complete inversion of the configuration. High diastereoselectivities were obtained (>15 : 1). The auxiliary could be efficiently removed by organolithium reactions of the amides furnishing α‐amino ketones. Another allyllithium addition allowed us to introduce a second allyl chain with high diastereoselectivity. Final ring closures by means of metatheses using Grubbs' (I) catalyst gave raise to the formation of enantiopure phenanthridines and cyclohexenes displaying defined substitution patterns ready for alkaloid total syntheses.     

(R)‐α‐Lipoyl‐Glycyl‐L‐Prolyl‐L‐Glutamyl Dimethyl Ester Codrug as a Multifunctional Agent with Potential Neuroprotective Activities

The (R)‐α‐lipoyl‐glycyl‐L ‐prolyl‐L ‐glutamyl dimethyl ester codrug (LA‐GPE, 1 ) was synthesized as a new multifunctional drug candidate with antioxidant and neuroprotective properties for the treatment of neurodegenerative diseases. Physicochemical properties, chemical and enzymatic stabilities were evaluated, along with the capacity of LA‐GPE to penetrate the blood–brain barrier (BBB) according to an in vitro parallel artificial membrane permeability assay for the BBB. We also investigated the potential effectiveness of LA‐GPE against the cytotoxicity induced by 6‐hydroxydopamine (6‐OHDA) and H₂O₂ on the human neuroblastoma cell line SH‐SY5Y by using the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) reduction assay. Our results show that codrug 1 is stable at both pH 1.3 and 7.4, exhibits good lipophilicity (log P=1.51) and a pH‐dependent permeability profile. Furthermore, LA‐GPE was demonstrated to be significantly neuroprotective and to act as an antioxidant against H₂O₂‐ and 6‐OHDA‐induced neurotoxicity in SH‐SY5Y cells.     

Efficient Synthesis of β,γ‐Alkynyl α‐Amino Acid Derivatives by Ag(I)‐Catalyzed Alkynylation of α‐Imino Esters

The first catalytic synthesis of β,γ‐alkynyl α‐amino acid derivatives was achieved by direct addition of terminal alkynes to α‐imino esters in the presence of an Ag(I) salt under mild reaction conditions.     

Cationic copolymers of α,β‐poly‐(N‐2‐hydroxyethyl)‐DL‐aspartamide (PHEA) and α,β‐polyasparthylhydrazide (PAHy): synthesis and characterization

In the present study the derivatization of two water‐soluble synthetic polymers, α,β‐poly(N‐2‐hydroxyethyl)‐DL ‐aspartamide (PHEA) and α,β‐polyasparthylhydrazide (PAHy), with glycidyltrimethylammonium chloride (GTA) is described. This reaction permits the introduction of positive charges in the macromolecular chains of PHEA and PAHy in order to make easier the electrostatic interaction with DNA. Different parameters affect the reaction of derivatization, such as GTA concentration and reaction time. PHEA reacts partially and slowly with GTA; on the contrary the reaction of PAHy with GTA is more rapid and extensive. The derivatization of PHEA and PAHy with GTA is a convenient method to introduce positive groups in their chains and it permits the preparation of interpolyelectrolyte complexes with DNA. © 2000 Society of Chemical Industry     

Ruthenium(II)‐Catalyzed Protocol for Preparation of Diverse α,β‐ and β,β‐Dihaloenones from Diazodicarbonyls

Efficient one‐step syntheses of α,β‐ and β,β‐dihaloenones were achieved by ruthenium(II)‐catalyzed reactions between cyclic or acyclic diazodicarbonyl compounds and oxalyl chloride or oxalyl bromide in moderate to good yields. This methodology offers several significant advantages, which include ease of handling, mild reaction conditions, one‐step reaction, and the use of an effective and non‐toxic catalyst. The synthesized compounds were further transformed into highly functionalized novel molecules bearing aromatic rings on the enone moiety using the Suzuki reaction.

     

Alcohol Dehydrogenase‐Catalyzed Synthesis of Enantiomerically Pure δ‐Lactones as Versatile Intermediates for Natural Product Synthesis

Starting from ethyl 4‐bromobutyrate, the chemoenzymatic synthesis of 6‐vinyl‐tetrahydro‐pyran‐2‐one has been accomplished. A screening of a number of available alcohol dehydrogenases and intense optimization of reaction parameters enabled us to establish an efficient synthesis of either enantiomer of the vinyllactone with excellent enantiomeric excess (>99%). The scope of possible applications of enantiopure vinyllactones has been verified by subjection to cross‐metathesis resulting in the total synthesis of the insect pheromone (S)‐5‐hexadecanolide and the cytotoxic styryllactone goniothalamine as well as derivatives thereof.     

Synthesis of γ‐Fluoro‐α, β‐unsaturated Carboxylic Esters from Saturated α‐Fluoro Aldehydes

γ‐Fluoro‐α, β‐unsaturated carboxylic esters 7a, 7b and 7d and 4‐fluoro‐4‐phenylbut‐3‐enoic ester ( 8 ) are obtained by two alternative pathways from 2‐fluoro aldehydes 5a—d , either by Horner—Wadsworth—Emmons reaction or by Wittig reaction. The aldehydes 5a—d are prepared by Swern oxidation of the corresponding fluorohydrins 4a—d . These are available from α‐olefins by bromofluorination, bromineby‐acetate replacement and subsequent hydrolysis.