An efficient synthesis for a new class antimalarial agent, 7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole

Efficient synthesis is essential for antimalarial therapeutics. A four-step route has been established for the synthesis of 7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole 1 that is a potent new class boron-containing antimalarial agent in preclinical development with IC₅₀ = 26 nM against the malaria parasite Plasmodium falciparum.     

Synthesis, characterization and antitumor activity of 1,2-disubstituted ferrocenes and cyclodextrin inclusion complexes

Seven different ferrocene derivatives have been tested in vitro against Ehrlich ascites tumor cells. Neither ferrocene nor the monosubstituted derivative N,N-dimethylaminomethylferrocene showed cytotoxic activity (IC₅₀ > 1000 μM for 3 h treatments). Better results were obtained with 1,2-disubstituted derivatives. The IC₅₀ values ranged from 376.6 μM for 1,2-diformylferrocene to 71.2 μM for racemic 2-(N,N-dimethylaminomethyl)ferrocenecarboxamide. The latter derivative was also encapsulated in native β-cyclodextrin (CD), heptakis-2,3,6-tri-O-methyl-β-CD (TRIMEB) and 2-hydroxypropyl-β-CD (HPβCD) to give 1:1 (host:guest) inclusion compounds. The existence of true inclusion complexes in the solid state was confirmed by a combination of powder X-ray diffraction, thermogravimetric analysis, FTIR and ¹³C CP MAS NMR spectroscopy. The IC₅₀ value for the β-CD inclusion compound was identical to that obtained for the nonincluded ferrocene derivative. By contrast, the inclusion compounds comprising TRIMEB and HPβCD yielded IC₅₀ values of 25.2 and 20.0 μM, respectively. No obvious relationship could be established between the redox behavior of the compounds determined by cyclic voltammetry and the biochemical data.     

Synthesis of α-trifluoromethyl-α-amino-β-sulfone hydroxamates: novel nanomolar inhibitors of matrix metalloproteinases

The racemic α-trifluoromethyl-α-amino-β-sulfone hydroxamates 1 were synthesized by means of a nucleophilic addition of sulfur-stabilized carbanions to a N-Cbz imine of trifluoropyruvate (4). The free amino derivative 1a was the most potent inhibitor of both MMP-3 (stromelysin-1) and MMP-9 (gelatinase-B), showing an IC₅₀ = 14 nM and 1 nM, respectively, and excellent selectivity versus MMP-1 (>5000-fold difference in inhibitory capacity). The N-Me derivative 1b was the most selective for MMP-3 with respect to MMP-9 (62-fold difference).     

The discovery,synthesis and antimalarial evaluation of natural product-based polyamine alkaloids

Bioassay-guided fractionation of an antimalarial extract derived from the fungus Ramaria subaurantiaca afforded the known polyamine alkaloid, pistillarin. Nine pistillarin analogues were synthesised via EDC-mediated chemistry and these compounds along with the previously reported natural product polyamines, ianthelliformisamines A–C and spermatinamine, were evaluated against Plasmodium falciparum (3D7) parasites and a normal human cell line to determine parasite-specific activity. Spermatinamine (IC₅₀ 0.23 μM) and pistillarin (IC₅₀ 1.9 μM) were the two most potent antimalarials identified during these studies.     

Cytotoxic eremophilane sesquiterpenoids from the saprobic fungus Berkleasmium nigroapicale BCC 8220

Berkleasmins A-E, five new eremophilane sesquiterpenoids were isolated from the saprobic fungus Berkleasmium nigroapicale BCC 8220. The structures of the new compounds were elucidated by analyses of NMR spectroscopic and mass spectrometry data in combination with chemical means. Berkleasmins A and C exhibited cytotoxic activity against cancer cell-lines (NCI-H187, MCF-7, and KB) as well as nonmalignant Vero cells with IC₅₀ values of 1.1-7.5 μg/mL, and these compounds also showed antimalarial activity with respective IC₅₀ of 3.1 and 2.8 μg/mL.     

Four Classes of Structurally Unusual Peptides from Two Marine-Derived Fungi: Structures and Bioactivities

The structures and biological properties of peptides produced by two genera of marine-derived fungi, an atypical Acremonium sp., and a Metarrhizium sp., were explored. The Acremonium strain was isolated from a marine sponge and has previously been shown by our group to produce peptides from the efrapeptin and RHM families. The isolation and structural elucidation of the new linear pentadecapeptides efrapeptins Eα (1) and H (2) and N-methylated octapeptides RHM3 (3) and RHM4 (4) were carried out through a combination of 1D and 2D NMR techniques and tandem MS. Additional known efrapeptins E, F, and G and the known syctalidamides A and B were also isolated. The absolute configurations of 1-4 are proposed to be the same as the original compound families. The marine sponge-derived Metarrhizium sp. was shown to produce destruxin cyclic depsipeptides including A, B, B2, desmethyl B, E chlorohydrin, and E2 chlorohydrin. Efrapeptins Eα (1), F, and G each displayed IC₅₀s of 1.3 nM against H125 cells, and destruxin E2 chlorohydrin displayed an IC₅₀ of 160 nM against HCT-116 cells. An initial therapeutic assessment suggested a continuous (168 h) exposure of at least 2 ng/mL, or a daily (24 h) exposure of at least 300 ng/mL for H125 cells treated with efrapeptin G, and a continuous (168 h) exposure of at least 190 ng/mL for HCT-116 cells treated with destruxin E2 chlorohydrin, will cause 90% tumor cell death in vitro.     

The synthesis, structural characterization and in vitro anti-cancer activity of novel N-(3-ferrocenyl-2-naphthoyl) dipeptide ethyl esters and novel N-(6-ferrocenyl-2-naphthoyl) dipeptide ethyl esters

N-(3-ferrocenyl-2-naphthoyl) dipeptide esters (5-7) and N-(6-ferrocenyl-2-naphthoyl) dipeptide esters (8-10) were prepared by coupling either 3-ferrocenylnaphthalene-2-carboxylic acid 2 or 6-ferrocenylnaphthalene-2-carboxylic acid 4 to the dipeptide ethyl esters GlyAla(OEt) (5, 8), AlaGly(OEt) (6, 9), and AlaAla(OEt) (7, 10) using the standard N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC), 1-hydroxybenzotriazole (HOBt) protocol. All the compounds were fully characterized using a combination of ¹H NMR, ¹³C NMR, DEPT-135 and ¹H-¹³C COSY (HMQC) spectroscopy, electrospray ionization mass spectrometry (ESI-MS) and cyclic voltammetry (CV). In vitro, the cytotoxic effects of compounds 5-10 show improvements over the corresponding N-(ferrocenyl)benzoyl derivatives, with IC₅₀ values against the H1299 lung cancer cells ranging from 1.2 μM to 8.0 μM. N-(6-ferrocenyl-2-naphthoyl)-glycine-l-alanine ethyl ester 8 was found to be the most active derivative of the naphthoyl series so far, displaying an IC₅₀ value of 1.3 ± 0.1 μM. This value is slightly lower than that found for the clinically employed anti-cancer drug cisplatin (IC₅₀ = 1.5 ± 0.1 μM against H1299).     

Modiolin and phthalide derivatives from the endophytic fungus Microsphaeropsis arundinis PSU-G18

One new modiolin, microsphaerodiolin (1), and seven new phthalides, microsphaerophthalides A-G (2-8), together with 12 known compounds were isolated from the endophytic fungus Microsphaeropsis arundinis PSU-G18. Their structures were elucidated by spectroscopic methods. The new 3-oxygenated phthalides are rare natural products. The known 1-(2,5-dihydroxyphenyl)-2-buten-1-one exhibited significant antifungal activity against Microsporum gypseum SH-MU-4 with an MIC value of 8 μg/mL, moderate antimalarial activity with an IC₅₀ value of 9.63 μg/mL and strong radical scavenging potency with the IC₅₀ value of 0.018 mg/mL. The new compounds 2 and 6 showed moderately antifungal activity against M. gypseum SH-MU-4 and Cryptococcus neoformans, respectively, with equal MIC values of 64 μg/mL.     

Fascioquinols A-F: bioactive meroterpenes from a deep-water southern Australian marine sponge, Fasciospongia sp.

Chemical investigation of a southern Australian deep-water marine sponge, Fasciospongia sp., returned the new meroterpene sulfate fascioquinol A (1) together with a series of acid mediated hydrolysis/cyclization products, fascioquinols B (2), C (3) and D (4), and strongylophorine-22 (5). Additional co-metabolites include the new meroterpenes fascioquinol E (6) and fascioquinol F (8), together with the known sponge metabolite geranylgeranyl 1,4-hydroquinone (7). Structures were assigned to 1-8 on the basis of detailed spectroscopic analysis, chemical interconversion, mechanistic and biosynthetic considerations, and literature comparisons. The known 1,4-hydroquinone 7 was identified as the dominant cytotoxic principle in the Fasciospongia sp. extract, with selective inhibitory activity against gastric adenocarcinoma (AGS, IC₅₀ 8 μM) and neuroblastoma (SH-SY5Y, IC₅₀ 4 μM) cell lines. By contrast, while the fascioquinols displayed little or no inhibitory activity towards human cell lines, 1 and 2 displayed promising Gram-positive selective antibacterial activity towards Staphylococcus aureus (IC₅₀ 0.9-2.5 μM) and Bacillus subtilis (IC₅₀ 0.3-7.0 μM).     

A cell-microelectronic sensing technique for profiling cytotoxicity of chemicals

A cell-microelectronic sensing technique is developed for profiling chemical cytotoxicity and is used to study different cytotoxic effects of the same class chemicals using nitrosamines as examples. This technique uses three human cell lines (T24 bladder, HepG2 liver, and A549 lung carcinoma cells) and Chinese hamster ovary (CHO-K1) cells in parallel as the living components of the sensors of a real-time cell electronic sensing (RT-CES) method for dynamic monitoring of chemical toxicity. The RT-CES technique measures changes in the impedance of individual microelectronic wells that is correlated linearly with changes in cell numbers during t log phase of cell growth, thus allowing determination of cytotoxicity. Four nitrosamines, N-nitrosodimethylamine (NDMA), N-nitrosodiphenylamine (NDPhA), N-nitrosopiperidine (NPip), and N-nitrosopyrrolidine (NPyr), were examined and unique cytotoxicity profiles were detected for each nitrosamine. In vitro cytotoxicity values (IC₅₀) for NDPhA (ranging from 0.6 to 1.9 mM) were significantly lower than the IC₅₀ values for the well-known carcinogen NDMA (15-95 mM) in all four cell lines. T24 cells were the most sensitive to nitrosamine exposure among the four cell lines tested (T24 > CHO > A549 > HepG2), suggesting that T24 may serve as a new sensitive model for cytotoxicity screening. Cell staining results confirmed that administration of the IC₅₀ concentration from the RT-CES experiments inhibited cell growth by 50% compared to the controls, indicating that the RT-CES method provides reliable measures of IC₅₀. Staining and cell-cycle analysis confirmed that NDPhA caused cell-cycle arrest at the G0/G1 phase, whereas NDMA did not disrupt the cell cycle but induced cell death, thus explaining the different cytotoxicity profiles detected by the RT-CES method. The parallel cytotoxicity profiling of nitrosamines on the four cell lines by the RT-CES method led to the discovery of the unique cytotoxicity of NDPhA causing cell-cycle arrest. This study demonstrates a new approach to comprehensive testing of chemical toxicity.     

N-Hydroxypyridone alkaloids,chromone derivatives,and tetrahydroxanthones from the scale-insect pathogenic fungus Orbiocrella sp. BCC 33248

Six new compounds, an N-hydroxypyridone glucoside, orbiocrellin A (1), its aglycone orbiocrellin B (2), chromone glucosides 3 and 4, a dihydrochromone 5a/5b, and a chromone 6, were isolated from the scale-insect pathogenic fungus Orbiocrella sp. BCC 33248. Orbiocrellin A (1) exhibited antimalarial activity against Plasmodium falciparum K1 (IC₅₀ 3.1 μg/mL) while it was non-cytotoxic. In contrast, orbiocrellin B (2) showed both antimalarial (IC₅₀ 2.1 μg/mL) and cytotoxic (NCI-H187 cells, IC₅₀ 0.70 μg/mL) activities.     

Substituted Aryl Benzylamines as Potent and Selective Inhibitors of 17β-Hydroxysteroid Dehydrogenase Type 3

17β-Hydroxysteroid dehydrogenase type 3 (17β-HSD3) is expressed at high levels in testes and seminal vesicles; it is also present in prostate tissue and involved in gonadal and non-gonadal testosterone biosynthesis. The enzyme is membrane-bound, and a crystal structure is not yet available. Selective aryl benzylamine-based inhibitors were designed and synthesised as potential agents for prostate cancer therapeutics through structure-based design, using a previously built homology model with docking studies. Potent, selective, low nanomolar IC₅₀ 17β-HSD3 inhibitors were discovered using N-(2-([2-(4-chlorophenoxy)phenylamino]methyl)phenyl)acetamide (1). The most potent compounds have IC₅₀ values of approximately 75 nM. Compound 29, N-[2-(1-Acetylpiperidin-4-ylamino)benzyl]-N-[2-(4-chlorophenoxy)phenyl]acetamide, has an IC₅₀ of 76 nM, while compound 30, N-(2-(1-[2-(4-chlorophenoxy)-phenylamino]ethyl)phenyl)acetamide, has an IC₅₀ of 74 nM. Racemic C-allyl derivative 26 (IC₅₀ of 520 nM) was easily formed from 1 in good yield and, to determine binding directionality, its enantiomers were separated by chiral chromatography. Absolute configuration was determined using single crystal X-ray crystallography. Only the S-(+)-enantiomer (32) was active with an IC₅₀ of 370 nM. Binding directionality was predictable through our in silico docking studies, giving confidence to our model. Importantly, all novel inhibitors are selective over the type 2 isozyme of 17β-HSD2 and show <20% inhibition when tested at 10 µM. Lead compounds from this series are worthy of further optimisation and development as inhibitors of testosterone production by 17β-HSD3 and as inhibitors of prostate cancer cell growth.     

Versatile synthesis of chiral 2-substituted-5-oxo-1,2,3,4-tetrahydro-5H-1,4-benzodiazepines as novel scaffolds for peptidomimetic building

The stereocontrolled synthesis of phenylalanine and tryptophan derived 5-oxo-1,2,3,4-tetrahydro-5H-1,4-benzodiazepine derivatives is described. This new methodology involves a modified Strecker reaction of N-Boc protected amino aldehydes and methyl anthranilate, reduction of the resulting α-amino nitriles, and lactamization. The resulting 2-substituted-5-oxo-1,2,3,4-tetrahydro-5H-1,4-benzodiazepines were further functionalized at position 4 by alkylation or acylation reactions. One of these new tryptophan-derived 1,4-benzodiazepines showed significant selective binding affinity at cholecystokinin CCK₁ receptors (IC₅₀=156.5±33.2 nM).     

N-Demethyl-sauroxine,a novel Lycodine Group alkaloid from Huperzia saururus

The present study describes the isolation and identification of N-demethyl-sauroxine, a novel Lycopodium alkaloid obtained from Huperzia saururus (Lam.) Trevis. (Lycopodiaceae). Its structure and relative stereochemistry were elucidated on the basis of its spectral data and chemical correlations. Additionally, acetylcholinesterase inhibitory activity was evaluated (IC₅₀ = 209.6 ± 1.1 μM). The structure of the already identified alkaloid sauroxine was also re-validated through two dimensional NMR data.     

Novel highly potent, structurally simple γ-trifluoromethyl γ-sulfone hydroxamate inhibitor of stromelysin-1 (MMP-3)

The γ-trifluoromethyl γ-sulfone hydroxamate 1 was synthesized both in racemic and enantiomerically pure forms by means of a thia-Michael reaction of p-methoxythiophenol on achiral and chiral 3,3,3-trifluorocrotonoyl Michael acceptors. The (R)-1 enantiomer was the most potent inhibitor of MMP-3 (stromelysin-1), showing an IC₅₀ = 3.2 nM, as well as the most selective with respect to MMP-9 (65-fold).     

Gold nanoparticle-based electrochemical detection of protein phosphorylation

In this report, we demonstrate the application of Au nanoparticles in the electrochemical detection of protein phosphorylation. The method is based on the labeling of a specific phosphorylation event with Au nanoparticles, followed by electrochemical detection. The phosphorylation reaction is coupled with the biotinylation of the kinase substrate using a biotin-modified adenosine 5′-triphosphate [γ]-biotinyl-3,6,9-trioxaundecanediamine (ATP) as the co-substrate. When the phosphorylated and biotinylated kinase substrate is exposed to streptavidin-coated Au nanoparticles, the high affinity between the streptavidin and biotin resulted in the attachment of Au nanoparticles on the kinase substrate. The electrochemical response obtained from Au nanoparticles enables monitoring the activity of the kinase and its substrate, as well as the inhibition of small molecule inhibitors on protein phosphorylation. We determined the activity of Src non-receptor protein tyrosine kinase, p60^c-Src and protein kinase A in combination with their highly specific substrate peptides Raytide™ EL and Kemptide, respectively. The detection limits for Raytide™ EL and Kemptide were determined as 5 and 10 μM, (S/N = 3), and the detection limits for the kinase activity of p60^c-Src and protein kinase A (PKA) were determined as 5 and 10 U mL⁻¹, (S/N = 3), respectively. Tyrosine kinase reactions were also performed in the presence of a well-defined inhibitor, 4-amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazolo[3,4-d]pyrimidine (PP2), and its negative control molecule, 4-amino-7-phenylpyrazol[3,4-d] pyrimidine (PP3), which had no inhibition effect. Based on the dependency of Au nanoparticle signal on inhibitor concentration, IC₅₀ value, half-maximal inhibition of the inhibitors was estimated. IC₅₀ values of PP2, genistein and herbimycin A to p60^c-Src were detected as 5 nM, 25 μM and 900 nM, respectively. The inhibition of PKA activity on Kemptide using ellagic acid was monitored with an IC₅₀ of 3.5 μM. The performance of the biosensor was optimized including the kinase reaction, incubation with streptavidin-coated Au nanoparticles, and the small molecule inhibitors. Kinase peptide-modified electrochemical biosensors are promising candidates for cost-effective kinase activity and inhibitor screening assays.     

Photocytotoxic and anaerobic DNA cleavage activity of binuclear 3,3′-dithiodipropionic acid cobalt(II) complexes having phenanthroline bases

Dicobalt(II) complexes [{(B)Co^II}₂(μ-dtdp)₂] (1–3) of 3,3′-dithiodipropionic acid (dtdp) and phenanthroline bases (B), viz. 1,10-phenanthroline (phen in 1), dipyrido[3,2-d:2′,3′-f]quinoxaline (dpq in 2) and dipyrido[3,2-a:2′,3′-c]phenazine (dppz in 3), have been prepared, characterized and their photo-induced anaerobic DNA cleavage activity studied. The elemental analysis and mass spectral data suggest binuclear formulation of the complexes. The redox inactive complexes have magnetically non-interacting dicobalt(II) core showing magnetic moment of ∼3.9 μ_B per cobalt(II) center. The complexes show good binding propensity to calf thymus DNA giving K_b values within 4.3 × 10⁵–4.0 × 10⁶ M⁻¹. Thermal melting and viscosity data predict DNA groove binding and/or partial intercalative nature of the complexes. The complexes show significant anaerobic DNA cleavage activity in green light under argon atmosphere possibly involving radical species generated from the disulfide moiety in a type-I pathway. The DNA cleavage reaction under aerobic medium in green light is found to involve hydroxyl radical species. The dppz complex 3 exhibits significant photocytotoxicity in HeLa cervical cancer cells with an IC₅₀ value of 2.3 μM in UV-A light of 365 nm, while it is essentially non-toxic in dark giving an IC₅₀ value of >200 μM. A significant reduction of the dark toxicity of the organic dppz base (IC₅₀ = 8.3 μM in dark) is observed on binding to the cobalt(II) center while essentially retaining its photocytotoxicity in UV-A light (IC₅₀ = 0.4 μM).     

Synthesis and cytotoxicity studies of new dimethylamino-functionalised and heteroaryl-substituted titanocene anti-cancer drugs

From the carbolithiation of N,N-dimethylamino fulvene (3a) and different ortho-lithiated heterocycles (furan, thiophene and N-methylpyrrole), the corresponding lithium cyclopentadienide intermediate (4a-c) was formed. These three lithiated intermediates underwent a transmetallation reaction with TiCl₄ resulting in dimethylamino-functionalised titanocenes 5a-c. When these titanocenes were tested against LLC-PK cells, the IC₅₀ values obtained were of 240, and 28 μM for titanocenes 5a and 5b, respectively. The most cytotoxic titanocene 5c with an IC₅₀ value of 5.5 μM is found to be almost as cytotoxic as cis-platin, which showed an IC₅₀ value of 3.3 μM, when tested on the LLC-PK cell line, and titanocene 5c is approximately 400 times better than titanocene dichloride itself.     

Insights into Antimalarial Activity of N-Phenyl-Substituted Cinnamanilides

Due to the urgent need of innovation in the antimalarial therapeutic arsenal, a series of thirty-seven ring-substituted N-arylcinnamanilides prepared by microwave-assisted synthesis were subjected to primary screening against the chloroquine-sensitive strain of P. falciparum 3D7/MRA-102. The lipophilicity of all compounds was experimentally determined as the logarithm of the capacity factor k, and these data were subsequently used in the discussion of structure-activity relationships. Among the screened compounds, fourteen derivatives exhibited IC₅₀ from 0.58 to 31 µM, whereas (2E)-N-(4-bromo-2-chlorophenyl)-3-phenylprop-2-enamide (24) was the most effective agent (IC₅₀ = 0.58 µM). In addition, (2E)-N-[2,6-dibromo-4-(trifluoromethyl)- phenyl]-3-phenylprop-2-enamide (36), (2E)-N-[4-nitro-3-(trifluoromethyl)phenyl]-3-phenylprop- 2-enamide (18), (2E)-N-(2-bromo-5-fluorophenyl)-3-phenylprop-2-enamide (23), and (2E)-3-phenyl-N-(3,4,5-trichlorophenyl)prop-2-enamide (33) demonstrated efficacy in the IC₅₀ range from 2.0 to 4.3 µM, comparable to the clinically used standard chloroquine. The results of a cell viability screening performed using THP1-Blue™ NF-κB cells showed that none of these highly active compounds displayed any significant cytotoxic effect up to 20 μM, which makes them promising Plasmodium selective substances for further investigations.     

Calorimetric investigation of the effect of hydroxyanthraquinones in Rheum officinale Baill on Staphylococcus aureus growth

The inhibitory effects of five hydroxyanthraquinones (HAQs) from root and rhizoma of Rheum officinale Baill, a traditional Chinese medicinal (TCM) herb, on Staphylococcus aureus growth were investigated by calorimetry. The power-time curves of S.aureus with and without HAQ were acquired and the extent and duration of inhibitory effects on the metabolism evaluated by growth rate constants (k₁, k₂), half inhibitory ratio (IC₅₀), maximum heat output (P_max) and peak time (t_p). The value of k₁ and k₂ of S. aureus in the presence of the five HAQs decreased with the increasing concentrations of HAQs. Moreover, P_max was reduced and the value of t_p increased with increasing concentrations of the five drugs. The inhibitory activity varied for different drugs. IC₅₀ of the five HAQs was 4 μg ml⁻¹ for emodin, 3.5 μg ml⁻¹ for rhein, 10 μg ml⁻¹ for aloe-emodin, 1000 μg ml⁻¹ for chrysophanol, 1600 μg ml⁻¹ for physcion. The sequence of antimicrobial activity of the five HAQs: rhein > emodin > aloe-emodin > chrysophanol > physicion.