Synthesis and biological evaluation of 2-(3',4',5'-trimethoxybenzoyl)-3-N,N-dimethylamino benzo[b]furan derivatives as inhibitors of tubulin polymerization

Molecules that target microtubules have an important role in the treatment of cancer. A new class of inhibitors of tubulin polymerization based on the 2-(3,4,5-trimethoxybenzoyl)-2-dimethylamino-benzo[b]furan molecular skeleton was synthesized and evaluated for antiproliferative activity, inhibition of tubulin polymerization, and cell cycle effects. The most promising compound in this series was 2-(3,4,5-trimethoxybenzoyl)-3-dimethylamino-6-methoxy-benzo[b]furan, which inhibits cancer cell growth at nanomolar concentrations and interacts strongly with tubulin by binding to the colchicine site.     

3- and 6-Substituted 2-amino-4,5,6,7-tetrahydrothieno[2,3-c]pyridines as A1 adenosine receptor allosteric modulators and antagonists

A series of 2-amino-4,5,6,7-tetrahydrothieno[2,3-c]pyridines were prepared and evaluated as potential allosteric modulators at the A₁ adenosine receptor. The structure–activity relationships of the 3- and 6-positions of a series of 2-amino-4,5,6,7-tetrahydrothieno[2,3-c]pyridines were explored. Despite finding that 3- and 6-substituted 2-amino-4,5,6,7-tetrahydrothieno[2,3-c]pyridines possess the ability to recognize an allosteric site on the agonist-occupied A₁AR at relatively high concentrations, the structural modifications we have performed on this scaffold favor the expression of orthosteric antagonist properties over allosteric properties. This research has identified 2-amino-4,5,6,7-tetrahydrothieno[2,3-c]pyridines as novel class of orthosteric antagonist of the A₁AR and highlighted the close relationship between structural elements governing allosteric modulation and orthosteric antagonism of agonist function at the A₁AR.     

Design,synthesis and structure–activity relationship of 2-(3′,4′,5′-trimethoxybenzoyl)-benzo[b]furan derivatives as a novel class of inhibitors of tubulin polymerization

The biological importance of microtubules in mitosis and cell division makes them an interesting target for the development of anticancer agents. Small molecules such as benzo[b]furans are attractive as inhibitors of tubulin polymerization. Thus, a new class of inhibitors of tubulin polymerization based on the 2-(3′,4′,5′-trimethoxybenzoyl)-benzo[b]furan molecular skeleton, with electron-donating (Me, OMe or OH) or electron-withdrawing (F, Cl and Br) substituents on the benzene ring, was synthesized and evaluated for antiproliferative activity, inhibition of tubulin polymerization and cell cycle effects. Adding a methyl group at the C-3 position resulted in increased activity. The most promising compound in this series was 2-(3′,4′,5′-trimethoxybenzoyl)-3-methyl-6-ethoxy-benzo[b]furan, which inhibits cancer cell growth at nanomolar concentrations and interacts strongly with tubulin by binding to the colchicine site.     

Substituted 2-(3′,4′,5′-trimethoxybenzoyl)-benzo[b]thiophene derivatives as potent tubulin polymerization inhibitors

The central role of microtubules in cell division and mitosis makes them a particularly important target for anticancer agents. On our early publication, we found that a series of 2-(3′,4′,5′-trimethoxybenzoyl)-3-aminobenzo[b]thiophenes exhibited strong antiproliferative activity in the submicromolar range and significantly arrested cells in the G2–M phase of the cell cycle and induced apoptosis.In order to investigate the importance of the amino group at the 3-position of the benzo[b]thiophene skeleton, the corresponding 3-unsubstituted and methyl derivatives were prepared. A novel series of inhibitors of tubulin polymerization, based on the 2-(3,4,5-trimethoxybenzoyl)-benzo[b]thiophene molecular skeleton with a methoxy substituent at the C-4, C-5, C-6 or C-7 position on the benzene ring, was evaluated for antiproliferative activity against a panel of five cancer cell lines, for inhibition of tubulin polymerization and for cell cycle effects. Replacing the methyl group at the C-3 position resulted in increased activity compared with the corresponding 3-unsubstituted counterpart. The structure–activity relationship established that the best activities were obtained with the methoxy group placed at the C-4, C-6 or C-7 position. Most of these compounds exhibited good growth inhibition activity and arrest K562 cells in the G2–M phase via microtubule depolymerization.     

Glucose-6-phosphatase catalytic enzyme inhibitors: synthesis and in vitro evaluation of novel 4,5,6,7-tetrahydrothieno[3,2-c]- and -[2,3-c]pyridines

The discovery of the first class of potent glucose-6-phosphatase catalytic site inhibitors, substituted 4,5,6,7-tetrahydrothieno[3,2-c]- and -[2,3-c]pyridines, is described. Optimisation of this series involved solution phase combinatorial synthesis and very potent compounds were prepared with IC50 values down to 140 nM. The structure activity relationship (SAR) of these compounds indicates that: a tetrahydrothieno[3,2-c]pyridine core ring system and the isomeric [2,3-c] system are equipotent and much better than the corresponding benzo analogue, 1,2,3,4-tetrahydro-isoquinoline. The 4-substituent of the tetrahydrothieno[3,2-c]pyridine ring has to be a phenyl group, optionally substituted with a lipophilic 4-substituent, such as trifluoromethoxy or chloro. The 5-substituent of the tetrahydrothieno[3,2-c]pyridine ring has to be a substituted benzoyl; anisoyl and (E)-3-furan-3-ylacryloyl are the best of the investigated groups. Substitution in the benzoyl ortho position seems to be forbidden, whereas substitution in the meta position is tolerated only if a methoxy para substituent is present. These SAR findings were parallel to those obtained in the 4,5,6,7-tetrahydrothieno[2,3-c]pyridine system. Enantioselectivity in enzyme recognition was observed and the activity resided in all cases only in one of the enantiomers.     

Synthesis of novel antimitotic agents based on 2-amino-3-aroyl-5-(hetero)arylethynyl thiophene derivatives

Microtubules are dynamic structures that play a crucial role in cellular division and are recognized as an important target for cancer therapy. In search of new compounds with strong antiproliferative activity and simple molecular structure, a new series of 2-amino-3-(3′,4′,5′-trimethoxybenzoyl)-5-(hetero)aryl ethynyl thiophene derivatives was prepared by the Sonogashira coupling reaction of the corresponding 5-bromothiophenes with several (hetero)aryl acetylenes. When these compounds were analyzed in vitro for their inhibition of cell proliferation, the 2- and 3-thiophenyl acetylene derivatives were the most powerful compounds, both of which exerted cytostatic effects at submicromolar concentrations. In contrast, the presence of a more flexible ethyl chain between the (hetero)aryl and the 5-position of the thiophene ring resulted in significant reduction in activity relative to the 5-(hetero)aryl acetylene substituted derivatives. The effects of a selected series of compounds on cell cycle progression correlated well with their strong antiproliferative activity and inhibition of tubulin polymerization. We found that the antiproliferative effects of the most active compounds were associated with increase of the proportion of cells in the G₂/M and sub-G₁ phases of the cell cycle.     

Synthesis and bioactivities of novel 4,5,6,7-tetrahydrothieno[2,3-c]pyridines as inhibitors of tumor necrosis factor-alpha (TNF-alpha) production

Novel 4,5,6,7-tetrahydrothieno[2,3-c]pyridine derivatives were synthesized and evaluated for their abilities to inhibit lipopolysaccharide (LPS)-stimulated production of TNF-alpha in rat whole blood. Several of these compounds exhibited potent inhibitory activity.     

A novel tubulin inhibitor, 6h,suppresses tumor-associated angiogenesis and shows potent antitumor activity against non–small cell lung cancers

Tumor angiogenesis is closely associated with the metastasis and progression of non–small cell lung cancer (NSCLC), a highly vascularized solid tumor. However, novel therapeutics are lacking for the treatment of this cancer. Here, we developed a series of 2-aryl-4-(3,4,5-trimethoxy-benzoyl)-5-substituted-1,2,3-triazol analogs (6a–6x) as tubulin colchicine-binding site inhibitors, aiming to find a novel promising drug candidate for NSCLC treatment. We first identified 2-(2-fluorophenyl)-3-(3,4,5-trimethoxybenzoyl)-5-(3-hydroxyazetidin-1-yl)-2H-1,2,3-triazole (6h) as a hit compound, which inhibited angiogenesis induced by NSCLC cells both in vivo and in vitro. In addition, our data showed that 6h could tightly bind to the colchicine-binding site of tubulin and inhibit tubulin polymerization. We also found that 6h could effectively induce G2/M cell cycle arrest of A549 and H460 cells, inhibit cell proliferation, and induce apoptosis. Furthermore, we showed 6h had the potential to inhibit the migration and invasion of NSCLC cells, two basic characteristics of tumor metastasis. Finally, we found 6h could effectively inhibit tumor progression in A549 xenograft mouse models with minimal toxicity. Taken together, these findings provide strong evidence for the development of 6h as a promising microtubule colchicine-binding site inhibitor for NSCLC treatment.     

A new anti-tubulin agent containing the benzo[b]thiophene ring system.

A new type of inhibitor of tubulin polymerization was discovered based on the 3-aroyl-2-arylbenzo[b]thiophene molecular skeleton. The lead compound in this series, 2-(4'-methoxyphenyl)-3-(3',4',5'-trimethoxybenzoyl)-6-methoxybe nzo[b]thiophene 1, inhibited tubulin polymerization, caused an increase in the mitotic index of CA46 Burkitt lymphoma cells, and inhibited the growth of several human cancer cell lines.     

Synthesis and bioactivities of novel bicyclic thiophenes and 4,5,6,7-tetrahydrothieno[2,3-c]pyridines as inhibitors of tumor necrosis factor-alpha (TNF-alpha) production

We synthesized bicyclic thiophenes and 4,5,6,7-tetrahydrothieno[2,3-c]pyridine derivatives, and evaluated for their ability to inhibit LPS-stimulated production of TNF-alpha. Several compounds revealed excellent in vivo activity. Furthermore, an effective compound was found in adjuvant-induced arthritic model (AIA) of rat.     

1,5-Disubstituted 1,2,3-triazoles as cis-restricted analogues of combretastatin A-4: Synthesis, molecular modeling and evaluation as cytotoxic agents and inhibitors of tubulin

A series of cis-restricted 1,5-disubstituted 1,2,3-triazole analogues of combretastatin A-4 (1) have been prepared. The triazole 12f, 2-methoxy-5-(1-(3,4,5-trimethoxyphenyl)-1H-1,2,3-triazol-5-yl)aniline, displayed potent cytotoxic activity against several cancer cell lines with IC₅₀ values in the nanomolar range. The ability of triazoles to inhibit tubulin polymerization has been evaluated, and 12f inhibited tubulin polymerization with IC₅₀ = 4.8 μM. Molecular modeling experiments involving 12f and the colchicine binding site of ,β-tubulin showed that the triazole moiety interacts with β-tubulin via hydrogen bonding with several amino acids.     

Novel [1,2,4]triazolo[1,5-a]pyrimidine derivatives as potent antitubulin agents: Design,multicomponent synthesis and antiproliferative activities

As restricted CA-4 analogues, a novel series of [1,2,4]triazolo[1,5-a]pyrimidines possessing 3,4,5-trimethoxylphenyl groups has been achieved successfully via an efficient one-pot three-component reaction of 3-(3,4,5-trimethoxyphenyl)-1H-1,2,4-triazol-5-amine, 1,3-dicarbonyl compounds and aldehydes. Initial biological evaluation demonstrated some of target compounds displayed potent antitumor activity in vitro against three cancer cell lines. Among them, the most highly active analogue 26 inhibited the growth of HeLa, and A549 cell lines with IC₅₀ values at 0.75, and 1.02 μM, respectively, indicating excellent selectivity over non-tumoural cell line HEK-293 (IC₅₀ = 29.94 μM) which suggested that the target compounds might possess a high safety index. Moreover, cell cycle analysis illustrated that the analogue 26 significantly induced HeLa cells arrest in G2/M phase, meanwhile the compound could dramatically affect cell morphology and microtubule networks. In addition, compound 28 exhibited potent anti-tubulin activity with IC₅₀ values of 9.90 μM, and molecular docking studies revealed the analogue occupied the colchicine-binding site of tubulin. These observations suggest that [1,2,4]triazolo[1,5-a]pyrimidines represent a new class of tubulin polymerization inhibitors and well worth further investigation aiming to generate potential anticancer agents.     

Design,synthesis and bioactivities of novel diarylthiophenes: inhibitors of tumor necrosis factor-alpha (TNF-alpha) production

The design, synthesis and SAR of novel diarylthiophene derivatives were performed. These compounds were designed by structural hybridization of TNF-alpha production inhibitors bearing 4-fluorophenyl and 4-pyridyl groups such as FR133605, FR167653 and SB210313, and 6-acetyl-3-ethoxycarbonyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridine (1) found previously by us. As a result, several compounds were more potent in vitro than FR133605 against TNF-alpha production stimulated with lipopolysaccharide (LPS).     

Absence of mutagenic activity of ticlopidine in the Ames Salmonella/microsome test

Ticlopidine hydrochloride, 5-(o-chlorobenzyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine hydrochloride, a platelet aggregation inhibitor, was tested for mutagenic activity in the Ames Salmonella/mammalian microsome test. Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were employed. Two of these strains (TA1535 and TA100) are sensitive to base-pair substitution mutagens, and the remaining 3 are sensitive to frame-shift mutagens. There was no evidence that ticlopidine hydrochloride had any mutagenic activity either in the presence or absence of a liver microsomal supplement.     

2-Aminothiophene-3-carboxylates and carboxamides as adenosine A1 receptor allosteric enhancers

Three series of 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene and 2-amino-5,6,7,8-tetrahydrocyclohepta[b]thiophenes with 3-carboxylates and carboxamides have been prepared using the Gewald synthesis and evaluated as A(1)AR allosteric enhancers. The structure-activity relationships of these classes of compound are described. A number of compounds, notably 7b, are more potent and efficacious than PD81,723 (1).     

Cytotoxic and antivascular 1-methyl-4-(3-fluoro-4-methoxyphenyl)-5-(halophenyl)-imidazoles

A series of 1-methyl-4,5-diphenylimidazoles 6 with various patterns of m-halogen substitution at the 5-phenyl ring were tested for cytotoxicity in cancer and nonmalignant cell lines and for their capacity to prevent tube formation in HUVEC cultures. Unlike the monofluoro and difluoro derivatives 6a and 6e, the monobromo and diiodo analogs 6c and 6h were strongly cytotoxic and inhibited the polymerization of tubulin and the tube formation by HUVEC. The dibromo derivative 6g displayed a unique selectivity for KB-3-1 cervix and PC-3 prostate cancer cells. It also inhibited the tube formation by HUVEC and the polymerization of tubulin which is indicative of its potential antiangiogenic activity in solid tumors.     

Anti-cancer activity of m-carborane-containing trimethoxyphenyl derivatives through tubulin polymerization inhibition

m-Carborane-containing compound 1a was identified as a cell growth inhibitor from a random screening of a boron compound library. As 1a is a mixture of diastereomers due to the presence of two chiral carbons, we designed achiral derivatives 2–4 and studied the structure-activity relationships of the methoxy groups on the benzene ring. 3,4,5-Trimethoxybenzyl derivative 2a and 3,4,5-trimethoxybenzoyl derivative 3a showed more potent anti-cancer activity against the human breast cancer cell line MDA-MB-453 than lead compound 1a. Compound 3a inhibited tubulin polymerization in a dose-dependent manner.     

3-(3-Methoxyphenyl)-6-(3-amino-4-methoxyphenyl)-7H-[1,2,4] triazolo [3,4-b][1,3,4] thiadiazine,a novel tubulin inhibitor,evokes G2/M cell cycle arrest and apoptosis in SGC-7901 and HeLa cells

Gastric cancer and cervical cancer are two major malignant tumors that threaten human health. The novel chemotherapeutic drugs are needed urgently to treat gastric cancer and cervical cancer with high anticancer activity and metabolic stability. Previously we have reported the synthesis, characterization and identification of a novel combretastatin A-4 analog, 3-(3-methoxyphenyl)-6-(3-amino-4- methoxyphenyl) -7H-[1,2,4]triazolo[3,4-b][1,3,4] thiadiazine (XSD-7). In this study, we sought to investigate its anticancer mechanisms in a human gastric cancer cell line (SGC-7901 cells) and human cervical carcinoma cell line (HeLa cells). The 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay showed that XSD-7 induced cytotoxicity in SGC-7901 and HeLa cells with inhibitory concentration 50 values of 0.11 ± 0.03 and 0.12 ± 0.05 µM, respectively. Immunofluorescence studies proved that XSD-7 inhibited microtubule polymerization during cell division in SGC-7901 and HeLa cells. Then, these cells were arrested at G2/M cell cycle and subsequently progressed into apoptosis. In further study, mitochondrial membrane potential analysis and Western blot analysis demonstrated that XSD-7 treatment-induced SGC-7901 cell apoptosis via both the mitochondria-mediated pathway and the death receptor-mediated pathway. In contrast, XSD-7 induced apoptosis in HeLa cells mainly via the mitochondria-mediated pathway. Hence, our data indicate that XSD-7 exerted antiproliferative activity by disrupting microtubule dynamics, leading to cell cycle arrest, and eventually inducing cell apoptosis. XSD-7 with novel structure has the potential to be developed for therapeutic treatment of gastric cancer and cervical cancer.     

Discovery of 1-benzoyl-3-cyanopyrrolo[1,2-a]quinolines as a new series of apoptosis inducers using a cell- and caspase-based high-throughput screening assay. Part 1: Structure-activity relationships of the 1- and 3-positions

1-Benzoyl-3-cyanopyrrolo[1,2-a]quinoline (2a) was identified as a novel apoptosis inducer through our caspase- and cell-based high-throughput screening assay. Compound 2a had good activity against several breast cancer cell lines but was much less active against several other cancer cell lines. SAR studies of 2a found that substitution at the 4-position of the 1-benzoyl group was important for activity. Replacing the 3-cyano group by an ester or ketone group led to inactive compounds. Interestingly, 4-substituted analogs such as 1-(4-(1H-imidazol-1-yl)benzoyl)-3-cyanopyrrolo[1,2-a]quinoline (2k) were found to be broadly and highly active in the caspase activation assay as well as in the cell growth inhibition assay with low nM EC(50) and GI(50) values in human breast cancer cells T47D, human colon cancer cells HCT116, and hepatocellular carcinoma cancer cells SNU398. Compound 2a was found not to inhibit tubulin polymerization up to 50 microM, while 2k was found to inhibit tubulin polymerization with an IC(50) value of 5 microM, indicating that certain substituents at the 4-position of the 1-benzoyl group can change the mechanism of action.     

Design and synthesis of 3-(4,5,6,7-tetrahydro-3H-imidazo[4,5-c]pyridin-2-yl)-1H-quinolin-2-ones as VEGFR-2 kinase inhibitors

A series of 3-(4,5,6,7-tetrahydro-3H-imidazo[4,5-c]pyridin-2-yl)-1H-quinolin-2-ones have been identified as a new class of VEGFR-2 kinase inhibitors. A variety of (4,5,6,7-tetrahydro-imidazo[5,4-c]pyridin-2-yl)-acetic acid ethyl esters were synthesized, and their VEGFR-2 inhibitory activity was evaluated. Described herein are the preparation of the series and the effects of the compounds on VEGFR-2 kinase activity.