Identification of new 3-phenyl-1H-indole-2-carbohydrazide derivatives and their structure-activity relationships as potent tubulin inhibitors and anticancer agents: A combined in silico, in vitro and synthetic study.

Virtual screening of commercially available molecular entities by using CDRUG, structure-based virtual screening, and similarity identified eight new derivatives of 3-phenyl-1H-indole-2-carbohydrazide with anti-proliferative activities. The molecules were tested experimentally for inhibition of tubulin polymerisation, which revealed furan-3-ylmethylene-3-phenyl-1H-indole-2-carbohydrazide (27a) as the most potent candidate. Molecule 27a was able to induce G2/M phase arrest in A549 cell line, similar to other tubulin inhibitors. Synthetic modifications of 27a were focussed on small substitutions on the furan ring, halogenation at R1 position and alteration of furyl connectivity. Derivatives 27b, 27d and 27i exhibited the strongest tubulin inhibition activities and were comparable to 27a. Bromine substitution at R1 position showed most prominent anticancer activities; derivatives 27b-27d displayed the strongest activities against HuCCA-1 cell line and were more potent than doxorubicin and the parent molecule 27a with IC50 values <0.5 µM. Notably, 27b with a 5-methoxy substitution on furan displayed the strongest activity against HepG2 cell line (IC50 = 0.34 µM), while 27d displayed stronger activity against A549 cell line (IC50 = 0.43 µM) compared to doxorubicin and 27a. Fluorine substitutions at the R1 position tended to show more modest anti-tubulin and anticancer activities, and change of 2-furyl to 3-furyl was tolerable. The new derivatives, thiophenyl 26, displayed the strongest activity against A549 cell line (IC50 = 0.19 µM), while 1-phenylethylidene 21b and 21c exhibited more modest anticancer activities with unclear mechanisms of action; 26 and 21c demonstrated G2/M phase arrest, but showed weak tubulin inhibitory properties. Molecular docking suggests the series inhibit tubulin at the colchicine site, in agreement with the experimental findings. The calculated molecular descriptors indicated that the molecules obey Lipinski's rule which suggests the molecules are drug-like structures.

Synthesis, Characterization, and Biologic Activity of New Acyl Hydrazides and 1,3,4-Oxadiazole Derivatives.

Starting from isoniazid and carboxylic acids as precursors, thirteen new hydrazides and 1,3,4-oxadiazoles of 2-(4-substituted-phenoxymethyl)-benzoic acids were synthesized and characterized by appropriate means. Their biological properties were evaluated in terms of apoptosis, cell cycle blocking, and drug metabolism gene expression on HCT-8 and HT-29 cell lines. In vitro antimicrobial tests were performed by the microplate Alamar Blue assay for the anti-mycobacterial activities and an adapted agar disk diffusion technique for other non-tubercular bacterial strains. The best antibacterial activity (anti-Mycobacterium tuberculosis effects) was proved by 9. Compounds 7, 8, and 9 determined blocking of G1 phase. Compound 7 proved to be toxic, inducing apoptosis in 54% of cells after 72 h, an effect that can be predicted by the increased expression of mRNA caspases 3 and 7 after 24 h. The influence of compounds on gene expression of enzymes implicated in drug metabolism indicates that synthesized compounds could be metabolized via other pathways than NAT2, spanning adverse effects of isoniazid. Compound 9 had the best antibacterial activity, being used as a disinfectant agent. Compounds 7, 8, and 9, seemed to have antitumor potential. Further studies on the action mechanism of these compounds on the cell cycle may bring new information regarding their biological activity.

3D Bioprinting of Carbohydrazide-Modified Gelatin into Microparticle-Suspended Oxidized Alginate for the Fabrication of Complex-Shaped Tissue Constructs.

Extrusion-based bioprinting of hydrogels in a granular secondary gel enables the fabrication of cell-laden three-dimensional (3D) constructs in an anatomically accurate manner, which is challenging using conventional extrusion-based bioprinting processes. In this study, carbohydrazide-modified gelatin (Gel-CDH) was synthesized and deposited into a new multifunctional support bath consisting of gelatin microparticles suspended in an oxidized alginate (OAlg) solution. During extrusion, Gel-CDH and OAlg were rapidly cross-linked because of the Schiff base formation between aldehyde groups of OAlg and amino groups of Gel-CDH, which has not been demonstrated in the domain of 3D bioprinting before. Rheological results indicated that hydrogels with lower OAlg to Gel-CDH ratios possessed superior mechanical rigidity. Different 3D geometrically intricate constructs were successfully created upon the determination of optimal bioprinting parameters. Human mesenchymal stem cells and human umbilical vein endothelial cells were also bioprinted at physiologically relevant cell densities. The presented study has offered a novel strategy for bioprinting of natural polymer-based hydrogels into 3D complex-shaped biomimetic constructs, which eliminated the need for cytotoxic supplements as external cross-linkers or additional cross-linking processes, therefore expanding the availability of bioinks.

Synthesis, in vitro, and in vivo (Zebra fish) antitubercular activity of 7,8-dihydroquinolin-5(6H)-ylidenehydrazinecarbothioamides.

We, herein, describe the synthesis of a series of novel aryl tethered 7,8-dihydroquinolin-5(6H)-ylidenehydrazinecarbothioamides 4a-v, which showed in vitro and in vivo antimycobacterial activity against Mycobacterium tuberculosis (Mtb) H37Rv. The intermediates dihydro-6H-quinolin-5-ones 3a-v were synthesized from ß-enaminones, reacting with cyclochexane-1,3-dione/5,5-dimethylcyclohexane-1,3-dione and ammonium acetate using a modified Bohlmann-Rahtz reaction conditions. They were further reacted with thiosemicarbazide to give the respective hydrazine carbothioamides 4a-v. All the new analogues 4a-v, were characterized by their NMR and mass spectral data analysis. Among the twenty-two compounds screened for in vitro antimycobacterial activity against Mycobacterium tuberculosis H37Rv (ATCC27294), two compounds, 4e and 4j, exhibited the highest inhibition with an MIC of 0.39 µg/mL. Compounds 4a, 4g, and 4k were found to inhibit Mtb at an MIC of 0.78 µg/mL. Hydrazinecarbothioamides 4a-k, exhibited enhanced activity than dihydroquinolinones 3a-k. The observed increase in potency provides a clear evidence that hydrazinecarbothioamide is a potential pharmacophore, collectively imparting synergistic effect in enhancing antitubercular activity of the dihydroquinolinone core. The in vivo (Zebra fish) antimycobacterial screening of the in vitro active molecules led to the identification of a hit compound, 4j, with significant activity in the Mtb nutrient starvation model (2.2-fold reduction). Docking studies of 4j showed a hydrogen bond with the P156 residue of the protein.

Effective Virtual Screening Strategy toward heme-containing proteins: Identification of novel IDO1 inhibitors.

Developing small molecules occupying the heme-binding site using computational approaches remains a challenging task because it is difficult to characterize heme-ligand interaction in heme-containing protein. Indoleamine 2,3-dioxygenase 1 (IDO1) is an intracellular heme-containing dioxygenase which is associated with the immunosuppressive effects in cancer. With IDO1 as an example, herein we report a combined virtual screening (VS) strategy including high-specificity heme-binding group (HmBG)-based pharmacophore screening and cascade molecular docking to identify novel IDO1 inhibitors. A total of four hit compounds were obtained and showed proper binding with the heme iron coordinating site. Further structural optimization led to a promising compound S18-3, which exerted potent anti-tumor efficacy in BALB/c mice bearing established CT26 tumors by activating the host's immune system. These results suggest that S18-3 merits further study to assess its potential for the intervention of cancer. Furthermore, our study also unveils a novel in silico-based strategy for identifying potential regulators for hemeproteins within short timeframe.

Structure-Based Screening of Tetrazolylhydrazide Inhibitors versus KDM4 Histone Demethylases.

Human histone demethylases are known to play an important role in the development of several tumor types. Consequently, they have emerged as important medical targets for the treatment of human cancer. Herein, structural studies on tetrazolylhydrazide inhibitors as a new scaffold for a certain class of histone demethylases, the JmjC proteins, are reported. A series of compounds are structurally described and their respective binding modes to the KDM4D protein, which serves as a high-resolution model to represent the KDM4 subfamily in crystallographic studies, are examined. Similar to previously reported inhibitors, the compounds described herein are competitors for the natural KDM4 cofactor, 2-oxoglutarate. The tetrazolylhydrazide scaffold fills an important gap in KDM4 inhibition and newly described, detailed interactions of inhibitor moieties pave the way to the development of compounds with high target-binding affinity and increased membrane permeability, at the same time.

Dynamic covalent chemistry enables formation of antimicrobial peptide quaternary assemblies in a completely abiotic manner.

Naturally occurring peptides and proteins often use dynamic disulfide bonds to impart defined tertiary/quaternary structures for the formation of binding pockets with uniform size and function. Although peptide synthesis and modification are well established, controlling quaternary structure formation remains a significant challenge. Here, we report the facile incorporation of aryl aldehyde and acyl hydrazide functionalities into peptide oligomers via solid-phase copper-catalysed azide-alkyne cycloaddition (SP-CuAAC) click reactions. When mixed, these complementary functional groups rapidly react in aqueous media at neutral pH to form peptide-peptide intermolecular macrocycles with highly tunable ring sizes. Moreover, sequence-specific figure-of-eight, dumbbell-shaped, zipper-like and multi-loop quaternary structures were formed selectively. Controlling the proportions of reacting peptides with mismatched numbers of complementary reactive groups results in the formation of higher-molecular-weight sequence-defined ladder polymers. This also amplified antimicrobial effectiveness in select cases. This strategy represents a general approach to the creation of complex abiotic peptide quaternary structures.

Synthesis and insecticidal activity of acridone derivatives to Aedes aegypti and Culex quinquefasciatus larvae and non-target aquatic species.

A serious Mosquito borne yellow fever is one of the grave diseases which affect the major population. Since there is no specific treatment for yellow fever, there is a necessity to develop an effective agent. The series of acridinone analogues 3 to 5 were synthesized with help of non-conventional microwave heating and confirmed by respective spectral characterization. 5c and 3b showed highest activity to kill 90% of larvae against A. aegypti and C. quinquefasciatus, respectively. Also the active products were treated to check the mortality of non-target aquatic species. Through the reports of the larvicidal bioassay, compounds 3b against C. quinquefasciatus whereas 5c against A. aegypti were found to be more active. By keeping this as a platform, further extension of the work can be done to find out a valuable drug for controlling disease vectors.

Synthesis and biological evaluation of 4-(2-(dimethylamino)ethoxy)benzohydrazide derivatives as inhibitors of Entamoeba histolyica.

In the quest for potent antiamoebic agents, a series of hydrazone hybrids (H1H30) have been designed and sequentially synthesized. The dimethylaminoethoxy and hydrazone entities incorporated into one molecule proved to be more persuasive and selective approach towards designing of antiamoebic agent. The synthesized compounds exhibited promising results against E. histolytica. The compound N'-(2-chlorobenzylidene)-4-(2-(dimethylamino) ethoxy)benzohydrazide was most impending among the series. Cytotoxicity profile showed better cell viability on lung cancer cell line (A549 cells) by MTT assay.

Investigation of the cytotoxic implications of metal chelators against melanoma and approaches to improve the cytotoxicity profiles of metal coordinating agents.

The cytotoxic activity of thiosemicarbazones (TSC) and thiocarbohydrazones was investigated against the MelRm melanoma cell line. In general, the melanoma line was susceptible to metal coordinating agents, the most useful of which incorporated the dipyridyl ketone hydrazone sub-structure. The impact of copper supplementation on the cytotoxic activity towards the melanoma line (MelRm) of metal coordinating agents when acting as ionophores is less predictable than the general improvement that has been seen in other cancer cells such as breast adenocarcinoma (MCF-7). The bimetallic nature of thiocarbohydrazone complexes with resultant loss of lipophilicity is a limiting factor in usage against MelRm. The cytotoxic activity of TSC against MelRm when used as copper ionophores could be markedly improved through combination with a partner drug capable of disrupting cellular defences to oxidative stress. In the absence of copper supplementation, both TSC and thiocarbohydrazones could be used to initiate cell cycle arrest and this could be employed to improve cytotoxicity profiles of other metallodrugs such as cisplatin.

Reaction parameters for the synthesis of N,N-dimethyl fatty hydrazides from oil.

Hydrazide derivatives have been synthesized from methyl esters, hydrazones and vegetable oils. They are important due to their diverse applications in pharmaceutical products, detergents as well as in oil and gas industries. The chemical synthesis of fatty hydrazides is well-established; however, only a few publications described the synthesis of fatty hydrazide derivatives, particularly, when produced from refined, bleached and deodorized palm olein. Here, the synthesis and characterization of N,N-dimethyl fatty hydrazides are reported. The N,N-dimethyl fatty hydrazides was successfully synthesized from fatty hydrazides and dimethyl sulfate in the presence of potassium hydroxide with the molar ratio of 1:1:1, 6 hours reaction time and 80℃ reaction temperature in ethanol. The product yield and purity were 22% and 89%, respectively. The fatty hydrazides used were synthesized from refined, bleached and deodorized palm olein with hydrazine monohydrate at pH 12 by enzymatic route. Fourier transform infrared, gas chromatography and nuclear magnetic resonance (NMR) spectroscopy techniques were used to determine the chemical composition of N,N-dimethyl fatty hydrazides. Proton NMR confirmed the product obtained were N,N-dimethyl fatty hydrazides.

Copper(ii)-catalyzed coupling reaction: an efficient and regioselective approach to N',N'-diaryl acylhydrazines.

Using N'-aryl acylhydrazines as aryl donors, a novel copper(ii)-catalyzed homo-coupling reaction of N'-aryl acylhydrazines has been developed for the synthesis of N',N'-diaryl acylhydrazines. We also provided a complementary procedure for the preparation of unsymmetrical diaryl acylhydrazines via cross-coupling reaction. These protocols featured mild reaction conditions, wide functional group tolerance and highly regioselective products. Control experiments indicated that this kind of coupling reaction might undergo a transient acyl diazene intermediate.

[Bacteriological substantiation of the antituberculosis effect of immunotropic drug tubosan].

The antituberculosis effect of tubosan registered in the Russian Federation as an immunostimulant drug has been studied on a series of 53 clinical strains of Mycobacterium tuberculosis (MBT). It is established that tubosan produces a bactericidal effect on drug-sensitive MBT strains and a bacteriostatic effect on resistant MBT strains. Use of tubosan for the treatment of 102 patients with drug-resistant destructive tuberculosis showed evident clinical-radiological involution process, with prospects of clinical recovery for 75% of patients. Results of this study show that tubosan possesses both antituberculosis and immunostimulant properties. It is recommended to use tubosan for complex treatment of patients with drug-resistant tuberculosis.

Design, synthesis, structural characterization by IR, (1) H, (13) C, (15) N, 2D-NMR, X-ray diffraction and evaluation of a new class of phenylaminoacetic acid benzylidene hydrazines as pfENR inhibitors.

Recent studies have revealed that plasmodial enoyl-ACP reductase (pfENR, FabI), one of the crucial enzymes in the plasmodial type II fatty acid synthesis II (FAS II) pathway, is a promising target for liver stage malaria infections. Hence, pfENR inhibitors have the potential to be used as causal malarial prophylactic agents. In this study, we report the design, synthesis, structural characterization and evaluation of a new class of pfENR inhibitors. The search for inhibitors began with a virtual screen of the iResearch database by molecular docking. Hits obtained from the virtual screen were ranked according to their Glide score. One hit was selected as a lead and modified to improve its binding to pfENR; from this, a series of phenylamino acetic acid benzylidene hydrazides were designed and synthesized. These molecules were thoroughly characterized by IR, (1) H, (13) C, (15) N, 2D-NMR (COSY, NOESY, (1) H-(13) C, (1) H-(15) N HSQC and HMBC), and X-ray diffraction. NMR studies revealed the existence of conformational/configurational isomers around the amide and imine functionalities. The major species in DMSO solution is the E, E form, which is in dynamic equilibrium with the Z, E isomer. In the solid state, the molecule has a completely extended conformation and forms helical structures that are stabilized by strong hydrogen bond interactions, forming a helical structure stabilized by N-H…O interactions, a feature unique to this class of compounds. Furthermore, detailed investigation of the NMR spectra indicated the presence of a minor impurity in most compounds. The structure of this impurity was deduced as an imidazoline-4-one derivative based on (1) H-(13) C and (1) H-(15) H HMBC spectra and was confirmed from the NOESY spectra. The molecules were screened for in vitro activity against recombinant pfENR enzyme by a spectrophotometric assay. Four molecules, viz. 17, 7, 10, and 12 were found to be active at 7, 8, 10, and 12 µm concentration, respectively, showing promising pfENR inhibitory potential. A classification model was derived based on a binary QSAR approach termed recursive partitioning (RP) to highlight structural characteristics that could be tuned to improve activity.

"True" antiandrogens-selective non-ligand-binding pocket disruptors of androgen receptor-coactivator interactions: novel tools for prostate cancer.

Prostate cancer (PCa) therapy typically involves administration of "classical" antiandrogens, competitive inhibitors of androgen receptor (AR) ligands, dihydrotestosterone (DHT) and testosterone (tes), for the ligand-binding pocket (LBP) in the ligand-binding domain (LBD) of AR. Prolonged LBP-targeting leads to resistance, and alternative therapies are urgently required. We report the identification and characterization of a novel series of diarylhydrazides as selective disruptors of AR interaction with coactivators through application of structure and ligand-based virtual screening. Compounds demonstrate full ("true") antagonism in AR with low micromolar potency, selectivity over estrogen receptors α and ß and glucocorticoid receptor, and partial antagonism of the progesterone receptor. MDG506 (5) demonstrates low cellular toxicity in PCa models and dose responsive reduction of classical antiandrogen-induced prostate specific antigen expression. These data provide compelling evidence for such non-LBP intervention as an alternative approach or in combination with classical PCa therapy.

Novel furfurylidene N-acylhydrazones derived from natural safrole: discovery of LASSBio-1215, a new potent antiplatelet prototype.

We describe herein the discovery of (E)-N-methyl-N'-((5-nitrofuran-2-yl)methylene)benzo[d]( 1 , 3 ) dioxole-5-carbohydrazide (9e), named LASSBio-1215, as a novel antiplatelet agent belonging to the N-methyl-N-acylhydrazone class, which exert their antiaggregating actions on human and rabbit platelets induced by different agonists, through cyclooxygenase-1 (COX-1) or thromboxane synthase inhibition. This compound was elected after screening of a series of functionalized furyl N-acylhydrazone derivatives, synthesized from natural safrole 10. In vitro assays showed that compound 9e presents platelet-aggregating activity in rabbit platelet-rich plasma (PRP) induced by arachidonic acid (IC(50) = 0.7 µM) and collagen (IC(50) = 4.5 µM). Moreover, LASSBio-1215 also inhibited almost completely the second wave of adenosine diphosphate-induced platelet aggregation in human PRP, and this effect was correlated with their ability to block the production of pro-aggregating autacoid thromboxane A(2).

Synthesis, antihypertensive activity, and 3D-QSAR studies of some new p-hydroxybenzohydrazide derivatives.

p-Hydroxybenzohydrazide 2 on treatment with aromatic/aliphatic aldehyde followed by cyclization with carbon disulphide afforded compounds 4a-4n. Also, compound 2 by treatment of substituted isothiocyanate followed by the treatment of chloroacetic acid yields the corresponding compounds 6a-6i. All the test compounds were assayed for antihypertensive activity by non-invasive blood pressure measurement and invasive blood pressure measurement methods. The test compounds showed significant antihypertensive activity. The intact compounds were subjected to 3D-QSAR studies. The 3D-QSAR analysis was carried out by PHASE program and a statistically reliable model with good predictive power (r(2) = 0.98, q(2) = 0.74) was achieved. The 3D-QSAR plots illustrated insights into the structure-activity relationship of these compounds which may aid in the design of potent p-hydroxybenzohydrazide derivatives as antihypertensive agents.

Characterisation of hydrazides and hydrazine derivatives as novel aspartic protease inhibitors.

Virtual screening of an in-house virtual library of synthetic compounds using FlexX, followed by enzyme inhibition, identified hydrazide and hydrazine derivatives as novel aspartic protease inhibitors. These compounds inhibited human cathepsin D and Plasmodium falciparum plasmepsin-II with low micromolar concentrations (IC(50) = 1-2.5 microM). Modelling studies with plasmepsin-II predicted binding of ligands at the centre of the extended substrate-binding cleft, where hydrazide/hydrazine parts of the inhibitors acted as the transition state mimic by forming electrostatic interactions with catalytic aspartates.

Synthesis and biological evaluation of novel hydrazide based cytotoxic agents.

Recently, we designed and synthesized a series of pyrroloquinoxaline compounds with hydrazine moiety linking a nitrogen-containing polycyclic group to a heteroaroyl system. Several derivatives, with attractive drug-like properties, were identified as promising cytotoxic agents, showing excellent potency in a panel of cancer cell lines. In the current study, we synthesized a further 19 new analogues to optimize their physicochemical properties and assess a coherent mechanism of action. Several chemical modifications were made to the reference compounds by varying the fused-ring system and/or the heteroacyl moiety. To evaluate their in vitro activity, we tested these compounds in six human cancer cell lines derived from different origins. Among them, two compounds ( 9 and 12 ) showed similar potency as the reference compounds with IC(50) values in the sub-micromolar range in all cell lines tested. Furthermore, compound 12 showed excellent in vivo efficacy in our preliminary human ovarian cancer mouse xenograft studies. Flow cytometric studies indicated that both derivatives interrupted cell cycle progression in colorectal cancer HCT116 cell lines and ovarian cancer SKOV-3 cells. Further mechanistic studies revealed that 9 and 12 were able to induce reactive oxygen species in SKOV-3 cells with apparently different kinetic patterns. Considering their cytotoxicity profiles in a variety of in vitro and in vivo cancer models, these hydrazide based compounds seem to have considerable potentials as novel chemotherapeutics.