Synthesis,molecular properties prediction and biological evaluation of indole-vinyl sulfone derivatives as novel tubulin polymerization inhibitors targeting the colchicine binding site

Twenty-two novel indole-vinyl sulfone derivatives were designed, synthesized and evaluated as tubulin polymerization inhibitors. The physicochemical and drug-likeness properties of all target compounds were predicted by Osiris calculations. All compounds were evaluated for their antiproliferative activities, among them, compound 7f exhibited the most potent activity against a panel of cancer cell lines, which was 2–7 folds more potent than our previously reported compound 4. Especially, 7f displayed about 8-fold improvement of selective index as compared with compound 4, indicating that 7f might have lower toxicity. Besides, 7f inhibited the microtubule polymerization by binding to the colchicine site of tubulin. Further investigations showed that compound 7f effectively disrupted microtubule network, caused cell cycle arrest at G2/M phase and induced cell apoptosis in K562 cells. Moreover, 7f reduced the cell migration and disrupted capillary-like tube formation in HUVEC cells. Importantly, the in vivo anti-tumor activity of 7f was validated in H22 liver cancer xenograft mouse model without apparent toxicity, suggesting that 7f is a promising anti-tubulin agent for cancer therapy.     

Design,synthesis and molecular modeling of new 4-phenylcoumarin derivatives as tubulin polymerization inhibitors targeting MCF-7 breast cancer cells

A new set of 4-phenylcoumarin derivatives was designed and synthesized aiming to introduce new tubulin polymerization inhibitors as anti-breast cancer candidates. All the target compounds were evaluated for their cytotoxic effects against MCF-7 cell line, where compounds 2f, 3a, 3b, 3f, 7a and 7b, showed higher cytotoxic effect (IC₅₀?=?4.3–21.2?μg/mL) than the reference drug doxorubicin (IC₅₀?=?26.1?μg/mL), additionally, compounds 1 and 6b exhibited the same potency as doxorubicin (IC₅₀?=?25.2 and 28.0?μg/mL, respectively). The thiazolidinone derivatives 3a, 3b and 3f with potent and selective anticancer effects towards MCF-7 cells (IC₅₀?=?11.1, 16.7 and 21.2?μg/mL) were further assessed for tubulin polymerization inhibition effects which showed that the three compounds were potent tubulin polymerization suppressors with IC₅₀ values of 9.37, 2.89 and 6.13?μM, respectively, compared to the reference drug colchicine (IC₅₀?=?6.93?μM). The mechanistic effects on cell cycle progression and induction of apoptosis in MCF-7 cells were determined for compound 3a due to its potent and selective cytotoxic effects in addition to its promising tubulin polymerization inhibition potency. The results revealed that compound 3a induced cell cycle cessation at G2/M phase and accumulation of cells in pre-G1 phase and prevented its mitotic cycle, in addition to its activation of caspase-7 mediating apoptosis of MCF-7 cells. Molecular modeling studies for compounds 3a, 3b and 3f were carried out on tubulin crystallography, the results indicated that the compounds showed binding mode similar to the co-crystalized ligand; colchicine. Moreover, pharmacophore constructed models and docking studies revealed that thiazolidinone, acetamide and coumarin moieties are crucial for the activity. Molecular dynamics (MD) studies were carried out for the three compounds over 100?ps. MD results of compound 3a showed that it reached the stable state after 30?ps which was in agreement with the calculated potential and kinetic energy of compound 3a.     

Synthesis and biological evaluation of cinnamido linked benzophenone hybrids as tubulin polymerization inhibitors and apoptosis inducing agents

A new class of hybrid molecules containing cinnamide subunit linked to benzophenone as inhibitors of tubulin polymerization were synthesized and evaluated for their anticancer potential. These hybrids exhibit anticancer activity with IC₅₀ values ranging from 0.06 to 16.3 μM. Compounds 4f and 4g possessing fluoro and trifluoromethyl on the cinnamido subunit showed significant cytotoxic activity with IC₅₀ values 0.06 and 0.09 μM against HeLa cell line, respectively. These compounds showed cell cycle arrest at G2/M phase of the cell cycle and inhibited tubulin polymerization followed by activation of caspase-3 activity and apoptotic cell death. Further in vitro tubulin polymerization assay showed that the level of tubulin inhibition was comparable to that of 2a for the compounds 4f and 4g. Moreover, Hoechst 33258 staining and DNA fragmentation assay suggested that these compounds induce cell death by apoptosis. Overall, the current study demonstrates that the synthesis of benzophenone linked cinnamide subunit conjugates as promising anticancer agents with G2/M arrest and apoptotic-inducing ability via targeting tubulin.     

Synthesis,cytotoxic activity,and tubulin polymerization inhibitory activity of new pyrrol-2(3H)-ones and pyridazin-3(2H)-ones

A series of new pyrrol-2(3H)-ones 4a-f and pyridazin-3(2H)-ones 7a-f were synthesized and characterized using different spectroscopic tools. Some of the tested compounds revealed moderate activity against 60 cell lines. The E form of the pyrrolones 4 showed good cytotoxic activity than both the Z form and the corresponding open amide form. Furthermore, the in vitro cytotoxic activity against HepG2 and MCF-7 cell lines revealed that compounds (E)4b, 6f and 7f showed good cytotoxic activity against HepG2 with IC₅₀ values of 11.47, 7.11 and 14.80 μM, respectively. Compounds (E)4b, 6f, 7d and 7f showed a pronounced inhibitory effect against cellular localization of tubulin. Flow cytometric analysis indicated that HepG2 cells treated with (E)4b showed a predominated growth arrest at the S-phase compared to that of G2/M-phase. Molecular modeling study using MOE® program indicated that most of the target compounds showed good binding of β-subunit of tubulin with the binding free energy (dG) values about −10 kcal/mole.     

Synthesis,biological evaluation and molecular modeling of 1,3,4-thiadiazol-2-amide derivatives as novel antitubulin agents

A series of 1,3,4-thiadiazol-2-amide derivatives (6a–w) were designed and synthesized as potential inhibitors of tubulin polymerization and as anticancer agents. The in vitro anticancer activities of these compounds were evaluated against three cancer cell lines by the MTT method. Among all the designed compounds, compound 6f exhibited the most potent anticancer activity against A549, MCF-7 and HepG2 cancer cell lines with IC₅₀ values of 0.03 μM, 0.06 μM and 0.05 μM, respectively. Compound 6f also exhibited significant tubulin polymerization inhibitory activity (IC₅₀ = 1.73 μM), which was superior to the positive control. The obtained results, along with a 3D-QSAR study and molecular docking that were used for investigating the probable binding mode, could provide an important basis for further optimization of compound 6f as a novel anticancer agent.     

Discovery of noscapine derivatives as potential β-tubulin inhibitors

Twenty novel 1,2,3-triazole noscapine derivatives were synthesized starting from noscapine by consecutive N-demethylation, reduction of lactone ring, N-propargylation and Huisgen 1,3-dipolar cycloaddition reaction. In order to select the most promising molecules to subject to further biophysical and biological evaluation, a molecular docking analysis round was performed using noscapine as reference compound. The molecules featuring docking predicted binding affinity better than that of noscapine were then subjected to MTT assay against MCF7 cell line. The obtained results disclosed that all the selected triazole derivatives exhibited a remarkably lower cell viability compared to noscapine in the range of 20 μM in 48 h. In an attempt to correlate the biological activity with the ability to bind tubulin, the surface plasmon resonance (SPR) assay was employed. Compounds 8a, 8h, 9c, 9f and 9j were able to bind tubulin with affinity constant values in the nanomolar range and higher if compared to noscapine. Integrating computational predictions and experimental evaluation, two promising compounds (8h and 9c) were identified, whose relevant cytotoxicity was supposed to be correlated with tubulin binding affinity. These findings shed lights onto structural modifications of noscapine toward the identification of more potent cytotoxic agents targeting tubulin.     

Synthesis and biological evaluation of 1-benzyl-N-(2-(phenylamino)pyridin-3-yl)-1H-1,2,3-triazole-4-carboxamides as antimitotic agents

A library of 1-benzyl-N-(2-(phenylamino)pyridin-3-yl)-1H-1,2,3-triazole-4-carboxamides (7a–al) have been designed, synthesized and screened for their anti-proliferative activity against some selected human cancer cell lines namely DU-145, A-549, MCF-7 and HeLa. Most of them have shown promising cytotoxicity against lung cancer cell line (A549), amongst them 7f was found to be the most potent anti-proliferative congener. Furthermore, 7f exhibited comparable tubulin polymerization inhibition (IC₅₀ value 2.04 µM) to the standard E7010 (IC₅₀ value 2.15 µM). Moreover, flow cytometric analysis revealed that this compound induced apoptosis via cell cycle arrest at G₂/M phase in A549 cells. Induction of apoptosis was further observed by examining the mitochondrial membrane potential and was also confirmed by Hoechst staining as well as Annexin V-FITC assays. Furthermore, molecular docking studies indicated that compound 7f binds to the colchicine binding site of the β-tubulin. Thus, 7f exhibits anti-proliferative properties by inhibiting the tubulin polymerization through the binding at the colchicine active site and by induction of apoptosis.     

Synthesis and biological evaluation of podophyllotoxin congeners as tubulin polymerization inhibitors

A series of new podophyllotoxin derivatives containing structural modifications at C-7, C-8, and C-9 were synthesized and evaluated for their cytotoxic activity against three human cancer cell lines. All the synthesized compounds showed significant growth inhibition with GI₅₀ values in micromolar levels while some of the compounds were several times more potent against MCF-7 and HeLa cell lines than MIAPACA cell line. Three compounds (12a, 12d and 12e) emerged as potent compounds with broad spectrum of cytotoxic activity against all the tested cell lines with GI₅₀ values in the range of 0.01–2.1 μM. These compounds induce microtubule depolymerization and arrests cells at the G2/M phase of the cell cycle. Moreover, compounds 12d and 12e disrupted microtubule network and accumulated tubulin in the soluble fraction in a similar manner to their parent podophyllotoxin scaffold. In addition, structure activity relationship studies within the series were also discussed. Molecular docking studies of these compounds into the colchicine-binding site of tubulin, revealed possible mode of inhibition by these compounds.     

Synthesis,biological evaluation and molecular docking study of 1-amino-2-aroylnaphthalenes against prostate cancer

A series of functionalized naphthalene was synthesized and screened against human prostate cancer cell line (PC-3). The in vitro antiproliferative activity of the synthesized compounds was evaluated by monitoring their cytotoxic effects against PC-3 cells by using MTT assay. We observed that compound 5f resulted in more than 50% cell death at 14?µM. Treatment of PC-3 cells with 5f provides apoptosis by flow cytometry. Western blotting showed decreased expression of pro-caspase 8 and 9. Our study shows that cancer cell treated with 5f has higher concentration of reactive oxygen species as compare to untreated sample, which facilitate cancerous cell to enter apoptosis. Exact mechanism by which ROS is generated after 5f treatment is still under study. Molecular docking study further strengthens the results obtained from in vitro experiments. Compound 5f can be considered as a promising leads for anticancer agent against prostate cancer cells due to its potent cytotoxic activity and apoptotic effect.     

Design,synthesis and biological evaluation of millepachine derivatives as a new class of tubulin polymerization inhibitors

A series of novel tubulin polymerization inhibitors (9a–9p) have been synthesized and evaluated for their in vitro and in vivo biological activities. Among these compounds, 9e displayed strong antiproliferative activity against several tumor cell lines (IC₅₀ = 0.15–0.62 μM). Compound 9e was also shown to arrest cells in the G2/M phase of the cell cycle and inhibit the polymerization of tubulin. Molecular docking studies suggested that 9e binds into the colchicine binding site of tubulin. In xenograft experiments, 9e exerted more potent anticancer effect than anticancer drug taxol against the H460 Human lung carcinoma in BALB/c nude mice. In summary, these findings suggest that 9e is a promising new antimitotic compound for the potential treatment of cancer.     

Design,synthesis, biological and in silico evaluation of coumarin-hydrazone derivatives as tubulin targeted antiproliferative agents

Coumarin-based different series of hydrazone derivatives were synthesized and evaluated for anticancer activity against four different human cancer cell lines. The activity of the compounds were compared with doxorubicin as a standard drug and all the compounds exhibited good to moderate cytotoxicity with IC₅₀ values ranging from 6.07 to 60.45 µM against all the examined cancer cell lines. Based on the screening results, it was concluded that the compounds 12a and 18a were the most promising medicinal entities. In vitro tubulin polymerisation inhibition assay was performed for the compounds 12a and 18a and these two compounds displayed good potency when compared with colchicine as the standard drug. The interaction of these compounds with tubulin protein was also studied with the help of molecular docking technique using Discovery studio software. Furthermore, the molecular and ADMET properties of the compounds were computed with Osiris property software and PreADMET server. The compounds exhibited exciting in vitro and in silico results. Hence we propose that the compounds 12a and 18a could be developed as tubulin targeted potential antiproliferative agents.     

Synthesis and evaluation of N-((1-benzyl-1H-1,2,3-triazol-4-yl)methyl)nicotinamides as potential anticancer agents that inhibit tubulin polymerization

A series of N-((1-benzyl-1H-1,2,3-triazol-4-yl)methyl)nicotinamides (4) was synthesized and tested for their anticancer activity against a panel of 60 human cancer cell lines. Some of the representative compounds such as 4a, 4b, 4f, 4g, 4i and 4t were selected for the five dose study and amongst them 4g and 4i displayed significant anticancer activity with GI₅₀ values ranging from 0.25 to 8.34 and 1.42 to 5.86 μM, respectively. Cell cycle analysis revealed that these compounds induced cell cycle arrest at G2/M phase in MCF-7 cells. The most active compound in this series 4g also inhibited tubulin polymerization with IC₅₀ value 1.93 μM superior to that of E7010. Moreover, assay to investigate the effect on caspase-9, Hoechst staining and DNA fragmentation analysis suggested that these compounds induced cell death by apoptosis. Docking experiments showed that they interact and bind efficiently with tubulin protein. Overall, the results demonstrate that N-((1-benzyl-1H-1,2,3-triazol-4-yl)methyl)nicotinamide scaffold possess anticancer property by inhibiting the tubulin polymerization.     

Benzopyridooxathiazepine derivatives as novel potent antimitotic agents

Herein, we describe the structure–activity relationship study of a new 1-(arylalkyl)-11H-benzo[f]-1,2-dihydropyrido[3,2,c][1,2,5]oxathiazepine 5,5-dioxide series of antimitotic agents. The pharmacological results obtained from previous works allowed us to identify compound 1 as a new cytotoxic agent inhibiting tubulin polymerization. We have undertaken the synthesis of its non-methylated analogue 7 and have extended our investigations to a novel, structurally related benzopyridooxathiazepine dioxide series. Among all analogues synthesized in this study, compound 10b was the most promising, being 12-fold more potent than compound 1. Its activity over a panel of five tumoral cell lines was in the nanomolar range for all of the histological types tested and flow cytometric studies performed on L1210 cells showed an accumulation of the cells in the G2/M phases of the cell cycle with a significant percentage of tetraploid cells (8N DNA content). This interesting pharmacological profile, resulting from inhibition of tubulin polymerization, encouraged us to perform preliminary in vivo studies.     

Dual-targeting antitumor conjugates derived from platinum(IV) prodrugs and microtubule inhibitor CA-4 significantly exhibited potent ability to overcome cisplatin resistance

Here we report that three platinum(IV) prodrugs containing a tubulin inhibitor CA-4, as dual-targeting platinum(IV) prodrug, were synthesized and evaluated for antitumor activity using MTT assay. Among them, complex 9 exhibited the most potent antitumor activity against the tested cancer lines including cisplatin resistance cancer cells, and simultaneously displayed lower toxicity compared to cisplatin, respectively. Moreover, complex 9, in which was conjugated to an inhibitor of tubulin at one axial position of platinum(IV) complex, could effectively enter the cancer cells, and significantly induce cell apoptosis and arrest the cell cycle in A549 cells at G2/M stage, and dramatically disrupt the microtubule organization. In addition, mechanism studies suggested that complex 9 significantly induced reactive oxygen species (ROS) generation and decreased mitochondrial trans-membrane potential (MMP) in A549 cells, and effectively induced activation of caspases triggering apoptotic signaling through mitochondrial dependent apoptosis pathways.     

Discovery of substituted 4-anilino-2-arylpyrimidines as a new series of apoptosis inducers using a cell- and caspase-based high throughput screening assay. 2. Structure–activity relationships of the 2-aryl group

As a continuation of our efforts to discover and develop the apoptosis inducing 4-anilino-2-(2-pyridyl)pyrimidines as potential anticancer agents, we explored replacing the 2-pyridyl group by other aryl groups. SAR studies showed that the 2-pyridyl group can be replaced by a 3-pyridyl, 4-pyridyl and 2-pyrazinyl group, and that the SAR for the anilino group was similar to that of the 2-pyridyl series. However, replacement of the 2-pyridyl group by a phenyl group, a 3,5-dichloro-4-pyridyl group, or a saturated ring led to inactive compounds. Several potent compounds, including 2f, 3d, 3j and 4a, with EC₅₀ values of 0.048–0.024 μM in the apoptosis induction assay against T47D cells, were identified through the SAR studies. In a tubulin polymerization assay, compound 2f, which was active against all the three cell lines tested (T47D, HTC116 and SNU398), inhibited tubulin polymerization with an IC₅₀ value of 0.5 μM, while compound 2a, which was active against T47D cells but not active against HTC116 and SNU398 cells, was not active in the tubulin assay at up to 50 μM.     

Solvent free,catalyst free,microwave or grinding assisted synthesis of bis-cyclic imide derivatives and their evaluation for anticancer activity

Cyclic imides are well known to be very important antitumor agents such as mitonafide and amonafide etc. Based on this fact, we have synthesized two series of cyclic imide derivatives containing two cyclic imide moiety in their structures (bis-cyclic imides) and screened them for in vitro anticancer activity against five human cancer cell lines i.e. breast (T47D), lung (NCl H-522), colon (HCT-15), ovary (PA-1) and liver (Hep G2). One series of bis-cyclic imide derivatives (3a–h) have been synthesized by condensation of acid anhydrides (1a–b) with diamines (2a–d) and another series (9a–f, 10a–f, 11a–f and 12a–f) by condensation of various diamines (4a–f) with diacids (5–8) in good yields. Structures assigned to 3a–h, 9a–f, 10a–f, 11a–f and 12a–f were fully characterized by spectroscopic means and elemental analysis. On screening for in vitro anticancer activity, compounds 3a (breast T47D), 3d (breast T47D, liver Hep G2), 3e (breast T47D, liver Hep G2), 3h (colon HCT-15), 10f (liver Hep G2) and 11a (colon HCT-15, ovary PA-1) exhibited good anticancer activities with IC₅₀ values range from 12.41 ± 3.2 to 17.9 ± 2.5 μM.     

Discovery and optimization of 3,4,5-trimethoxyphenyl substituted triazolylthioacetamides as potent tubulin polymerization inhibitors

Based on our previous research, three series of new triazolylthioacetamides possessing 3,4,5-trimethoxyphenyl moiety were synthesized, and evaluated for antiproliferative activities and inhibition of tubulin polymerization. The most promising compounds 8b and 8j demonstrated more significant antiproliferative activities against MCF-7, HeLa, and HT-29 cell lines than our lead compound 6. Moreover, analogues 8f, 8j, and 8o manifested more potent antiproliferative activities against HeLa cell line with IC₅₀ values of 0.04, 0.05 and 0.16?μM, respectively, representing 100-, 82-, and 25-fold improvements of the activity compared to compound 6. Furthermore, the representative compound, 8j, was found to induce significant cell cycle arrest at the G₂/M phase in HeLa cell lines via a concentration-dependent manner. Meanwhile, compound 8b exhibited the most potent tubulin polymerization inhibitory activity with an IC₅₀ value of 5.9?μM, which was almost as active as that of CA-4 (IC₅₀?=?4.2?μM). Additionally, molecular docking analysis suggested that 8b formed stable interactions in the colchicine-binding site of tubulin.     

(E)-N-Aryl-2-oxo-2-(3,4,5-trimethoxyphenyl)acetohydrazonoyl cyanides as tubulin polymerization inhibitors: Structure-based bioisosterism design,synthesis, biological evaluation,molecular docking and in silico ADME prediction

A series of (E)-N-Aryl-2-oxo-2-(3,4,5-trimethoxyphenyl)acetohydrazonoyl cyanides have been synthesized and evaluated for their anticancer activity in human hepatocellular liver carcinoma HepG2 and breast adenocarcinoma MCF-7?cell lines. Among all the tested compounds, compound 3a, 3e and 3n displayed more activity than lead compound with IC₅₀ value of 0.26–0.61?μM. Meanwhile, these compounds (3a, 3e and 3n) showed potent antiproliferative activity against a panel of cancer cells and the HCT-8/T multidrug resistant cell line with IC₅₀ values in the range of 0.077– 7.44?μM. Flow cytometric analyses revealed that compound 3n induced cell cycle arrest in G2/M phases in a dose dependent manner. The compound 3n also displayed potent tubulin polymerization inhibition with an IC₅₀ value of 0.9?µM, with ten folds more active than colchicine (IC₅₀?=?9?μM). Molecular docking studies revealed that compound 3n efficiently interacted with the colchicine binding site of tubulin through hydrophobic, cation-π and hydrogen bond interaction. Furthermore, in silico pharmacokinetic prediction shown that these compounds have a good ADME-related physicochemical parameters. These results demonstrate that 3n exhibits potent cytotoxicity in cancer cells by targeting the colchicine binding site of tubulin and potentially acts as a therapeutic lead compound for the development of anticancer drugs.     

Tumor-associated carbonic anhydrase isoform IX and XII inhibitory properties of certain isatin-bearing sulfonamides endowed with in vitro antitumor activity towards colon cancer

Three series of indolinone-based sulfonamides (3a–f, 6a–f and 9a–f) were in vitro evaluated as inhibitors of the tumor-associated carbonic anhydrase (CA, EC 4.2.1.1) isoforms hCA IX and XII, using a stopped-flow CO₂ hydrase assay. All the investigated sulfonamides displayed single- or double-digit nanomolar inhibitory activities towards both hCA IX (K_Is: 6.2–64.8 nM) and XII (K_Is: 7.1–55.6 nM) isoforms. All sulfonamides (3a–f, 6a–f and 9a–f) were in vitro examined for their potential anticancer activity against colorectal cancer HCT-116 and breast cancer MCF-7 cell lines. Sulfonamide 9e was found to be the most potent counterpart against HCT-116 (IC₅₀ = 3.67 ± 0.33 µM). Sulfonamide 9e displayed good selectivity profile for inhibition of the tumor-associated isoforms (CAs IX & XII) over the off-target cytosolic CAs I and II. 9e was screened for cell cycle disturbance and apoptosis induction in HCT-116 cells. It was found to persuade cell cycle arrest at G₂-M stage as well as alter the Sub-G₁ phase. Also, 9e induced the intrinsic apoptotic mitochondrial pathway in HCT-116 cells via down-regulation of the anti-apoptotic protein Bcl-2 level with concurrent boosting the pro-apoptotic Bax, caspase-9, caspase-3, cytochrome C and p53 levels.     

Click chemistry-assisted synthesis of novel aminonaphthoquinone-1,2,3-triazole hybrids and investigation of their cytotoxicity and cancer cell cycle alterations

A series of 12 novel 1,4-naphthoquinone-1,2,3-triazole hybrids were designed and synthesized through copper-catalyzed click reaction of 2-(prop-2-ynylamino)naphthalene-1,4-dione (3) and different azidomethyl-benzene derivatives. The synthesized compounds were assessed for their anticancer activity against three cancer cell lines (MCF-7, HT-29 and MOLT-4) by MTT assay. The results showed that the majority of the synthesized compounds displayed cytotoxic activity. Derivatives 6f and 6h, bearing 4-trifluoromethyl-benzyl and 4-tert-butyl-benzyl groups, respectively, as well as intermediate 3 demonstrated good cytotoxic potential against all tested cancer cell lines, among which compound 6f showed the highest activity. Flow cytometric analysis revealed that compounds 3, 6f and 6h arrested cell cycle at G0/G1 phase in MCF-7 cells. Therefore, synthesized aminonaphthoquinone-1,2,3-triazole derivatives can be introduced as promising molecules for further development as potential anticancer agents.