Weige Zhang

3-(3-Hydroxy-4-methoxyphenyl)-4-(3,4,5-trimethoxyphenyl)-1,2,5-selenadiazole (G-1103), a novel combretastatin A-4 analog, induces G2/M arrest and apoptosis by disrupting tubulin polymerization in human cervical HeLa cells and fibrosarcoma HT-1080 cells.

Microtubule is a popular target for anticancer drugs. In this study, we describe the effect 3-(3-hydroxy-4-methoxyphenyl)-4-(3,4,5-trimethoxyphenyl)-1,2,5-selenadiazole (G-1103), a newly synthesized analog of combretastatin A-4 (CA-4), showing a strong time- and dose-dependent anti-proliferative effect on human cervical cancer HeLa cells and human fibrosarcoma HT-1080 cells. We demonstrated that the growth inhibitory effects of G-1103 in HeLa and HT-1080 cells were associated with microtubule depolymerization and proved that G-1103 acted as microtubule destabilizing agent. Furthermore, cell cycle analysis revealed that G-1103 treatment resulted in cell cycle arrest at the G2/M phase in a time-dependent manner with subsequent apoptosis induction. Western blot analysis revealed that down-regulation of cdc25c and up-regulation of cyclin B1 was related with G2/M arrest in HeLa and HT-1080 cells treatment with G-1103. In addition, G-1103 induced HeLa cell apoptosis by up-regulating cleaved caspase-3, Fas, cleaved caspase-8 expression, which indicated that G-1103 induced HeLa cell apoptosis was mainly associated with death receptor pathway. However, G-1103 induced HT-1080 cell apoptosis by up-regulating cleaved caspase-3, Fas, cleaved caspase-8, Bax and cleaved caspase-9 expression and down-regulating anti-apoptotic protein Bcl-2 expression, which indicated that G-1103 induced HT-1080 cell apoptosis was associated with both mitochondrial and death receptor pathway. Taken together, all the data demonstrated that G-1103 exhibited its antitumor activity through disrupting the microtubule assembly, causing cell cycle arrest and consequently inducing apoptosis in HeLa and HT-1080 cells. Therefore, the novel compound G-1103 is a promising microtubule inhibitor that has great potentials for therapeutic treatment of various malignancies.

Ultrasound-promoted two-step synthesis of 3-arylselenylindoles and 3-arylthioindoles as novel combretastatin A-4 analogues

A series of 3-(3′-hydroxy-4′-methoxyphenyl)selenyl-5,6,7-trimethoxy-1H-indoles and 3-(3′-hydroxy-4′-methoxyphenyl)thio-5,6,7-trimethoxy-1H-indoles were obtained as a new class of combretastatin A-4 (CA-4) analogues via a convenient ultrasound (US)-assisted two-step process involving 3-selenenylation/sulfenylation followed by O-deallylation. With the assistance of US irradiation, both the reaction rates and yields of selenenylation, sulfenylation and O-deallylation could be significantly improved. A comparison of the reaction rates of O-deallylation and ester reduction demonstrated that O-deallylation was more sensitive to US irradiation. Finally, these products were evaluated for their antiproliferative activities, and most of them showed moderate to potent activities against three human cancer cell lines in vitro.

Involvement of mitochondria and caspase pathways in N-demethyl-clarithromycin-induced apoptosis in human cervical cancer HeLa cell

AbstractAim:To study the mechanisms by which N-demethyl-clarithromycin (NDC) induces human cervical cancer HeLa cell apoptosis in vitro.Methods:The viability of N-demethyl-clarithromycin-induced HeLa cells was measured by MTT assay. Apoptotic cells with condensed nuclei were visualized by phase contrast microscopy. Nucleosomal DNA fragmentation was assayed by agarose gel electrophoresis. Measurement of mitochondrial transmembrane potential was analyzed by a FACScan flowcytometer. Caspase-3, poly-(ADP-ribose) polymerase (PARP), caspase-activated DNase (ICAD), Bcl-2, Bax, p53, and SIRT1 protein expression and the release of cytochrome c were detected by Western blot analysis.Results:N-demethyl-clarithromycin, an anti-inflammatory substance, inhibited HeLa cell growth in a dose- and time-dependent manner. N-demethyl-clarithromycin induced HeLa cell death through the apoptotic pathways. The pancaspase inhibitor (z-VAD-fmk), caspase-3 inhibitor (z-DEVD-fmk) andthecaspase-9 inhibitor (z-LEHD-fmk) partially enhanced cell viability induced by N-demethyl-clarithromycin, but the caspase-8 inhibitor (z-IETD-fmk) had almost no effect. Caspase-3 was activated then followed by the degradation of caspase-3 substrates, the inhibitor of ICAD and PARP. Simultaneously, mitochondrial transmembrane potential was markedly reduced and the release of cytochrome c in the cytosol was increased. N-demethyl-clarithromycin upregulated the expression ratio of mitochondrial Bax/Bcl-2, and significantly increased the expression of the p53 protein. It also downregulated anti-apoptotic protein SIRT1 expression.Conclusion:N-demethyl-clarithromycin induced apoptosis in HeLa cells via the mitochondrial pathway.

BZML, a novel colchicine binding site inhibitor, overcomes multidrug resistance in A549/Taxol cells by inhibiting P-gp function and inducing mitotic catastrophe

Multidrug resistance (MDR) interferes with the efficiency of chemotherapy. Therefore, developing novel anti-cancer agents that can overcome MDR is necessary. Here, we screened a series of colchicine binding site inhibitors (CBSIs) and found that 5-(3, 4, 5-trimethoxybenzoyl)-4-methyl-2-(p-tolyl) imidazol (BZML) displayed potent cytotoxic activity against both A549 and A549/Taxol cells. We further explored the underlying mechanisms and found that BZML caused mitosis phase arrest by inhibiting tubulin polymerization in A549 and A549/Taxol cells. Importantly, BZML was a poor substrate for P-glycoprotein (P-gp) and inhibited P-gp function by decreasing P-gp expression at the protein and mRNA levels. Cell morphology changes and the expression of cycle- or apoptosis-related proteins indicated that BZML mainly drove A549/Taxol cells to die by mitotic catastrophe (MC), a p53-independent apoptotic-like cell death, whereas induced A549 cells to die by apoptosis. Taken together, our data suggest that BZML is a novel colchicine binding site inhibitor and overcomes MDR in A549/Taxol cells by inhibiting P-gp function and inducing MC. Our study also offers a new strategy to solve the problem of apoptosis-resistance.

2-Methoxy-5((3,4,5-trimethosyphenyl)seleninyl) phenol (SQ0814061), a novel microtubule inhibitor, evokes G2/M cell cycle arrest and apoptosis in human breast cancer cells

Breast cancer is the leading cause of cancer death in women worldwide, and novel chemotherapeutic drugs with high activity and no drug resistance for treating breast cancer are needed urgently. In this study, we investigated the antitumor effect of 2-methoxy-5((3,4,5-trimethosyphenyl)seleninyl) phenol (SQ0814061), which has a strong inhibition of cell growth in MCF-7 and MDA-MB-231 cells. We demonstrated that SQ0814061 (SQ) time-dependently induced cell cycle arrest at G2/M phase and subsequently progressed into apoptosis, which is associated with microtubule depolymerization. Western blot analysis revealed that up-regulation of cyclin B1 and Aurora A was related with G2/M phase arrest in MCF-7 and MDA-MB-231 cells treatment with SQ. However, the formation of multinucleated cells after a long time exposed to SQ of MCF-7 cells delayed the cell death. In addition, apoptosis induced by SQ is correlated with the down-regulation of the PI3K-Akt-MDM2 pathway in MCF-7 and MDA-MB-231 cells. Treatment with the PI3K specific inhibitor, LY294002, increased SQ-induced cell growth inhibitory rate and apoptosis rate of MCF-7 and MDA-MB-231 cells. Moreover, SQ induced MCF-7 and MDA-MB-231 cells to generate reactive oxygen species (ROS), and the SQ-induced cell death was ROS dependent. In conclusion, all the data demonstrated that SQ exhibited its antitumor activity through disrupting the microtubule assembly, inducing cell cycle arrest and eventually apoptosis which is associated with PI3K-Akt-MDM2 pathway in MCF-7 and MDA-MB-231 cells. Therefore, the novel compound SQ is a promising microtubule inhibitor that has tremendous potentials for therapeutic treatment of human mastocarcinoma.

Activated carbon for aerobic oxidation: Benign approach toward 2-benzoylbenzimidazoles and 2-benzoylbenzoxazoles synthesis.

A general strategy involving a novel and highly efficient aerobic benzylic oxidation promoted by cheap, reusable activated carbon in water is developed. Application of this method has been demonstrated in the benign synthesis of bioactive 2-benzoylbenzimidazoles and 2-benzoylbenzoxazoles derivatives. Furthermore, the activated carbon catalyst could be recovered and reused at least three times without significantly losing its activity. Preliminary research suggests that the oxidation mechanism may involve intermediate hydroperoxidation and that a portion of the final carbonyl product is obtained through a secondary benzylic alcohol intermediate. Finally, theoretical calculations reveal that the oxidation yield is closely associated with the electric density at the benzylic position of the substrate.

Methyl 5-[(1H-indol-3-yl)selanyl]-1H-benzoimidazol-2-ylcarbamate (M-24), a novel tubulin inhibitor, causes G2/M arrest and cell apoptosis by disrupting tubulin polymerization in human cervical and breast cancer cells

Methyl 5-[(1H-indol-3-yl)selanyl]-1H-benzoimidazol-2-ylcarbamate (M-24) is a newly synthesized analogue of nocodazole by our group and has been found to be active for some cancer cells. However, its sensitivity to different cell lines and the underlying anticancer mechanism are still unclear. In this study, we proved that M-24 had strong time- and dose-dependent anti-proliferative effects on human cervical cancer HeLa cells and human breast carcinoma MCF-7 cells. We demonstrated that the growth inhibitory effects of M-24 in both cell lines were associated with microtubule depolymerization. Furthermore, M-24 treatment resulted in cell cycle arrest at the G2/M phase in a dose-dependent manner with subsequent apoptosis induction. Western blotting analysis revealed that up-regulation of cyclin B1 and cdc2 was related with G2/M arrest in both cell lines. In addition, M-24-induced HeLa cell apoptosis was mainly associated with mitochondria-dependent intrinsic pathway. However, M-24-induced MCF-7 cell apoptosis was associated with both mitochondrial and death receptor pathway. In conclusion, M-24 caused apoptosis through disrupting microtubule assembly and inducing cell cycle arrest in HeLa and MCF-7 cells. Therefore, the novel compound M-24 is a promising microtubule-destabilizing agent that has great potential for the therapy of various malignancies especially human cervical and breast cancers.

Design, synthesis and structure-activity relationship of 3,6-diaryl-7 H -[1,2,4]triazolo[3,4-b][1,3,4]thiadiazines as novel tubulin inhibitors

A novel series of 3,6-diaryl-7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazines were designed, synthesized and biologically evaluated as vinylogous CA-4 analogues, which involved a rigid [1,2,4]triazolo[3,4-b][1,3,4]thiadiazine scaffold to fix the configuration of (Z,E)-butadiene linker of A-ring and B-ring. Among these rigidly vinylogous CA-4 analogues, compounds 4d, 5b, 5i, 6c, 6e, 6g, 6i and 6k showed excellent antiproliferative activities against SGC-7901, A549 and HT-1080 cell lines with IC50 values at the nanomolar level. Compound 6i showed the most highly active antiproliferative activity against the three human cancer cell lines with an IC50 values of 0.011–0.015 µM, which are comparable to those of CA-4 (IC50 = 0.009–0.013 µM). Interestingly, SAR studies revealed that 3,4-methylenedioxyphenyl, 3,4-dimethoxyphenyl, 3-methoxyphenyl and 4-methoxyphenyl could replace the classic 3,4,5-trimethoxyphenyl in CA-4 structure and keep antiproliferative activity in this series of designed compounds. Tubulin polymerization experiments showed that 6i could effectively inhibit tubulin polymerization, which was corresponded with CA-4, and immunostaining experiments suggested that 6i significantly disrupted microtubule/tubulin dynamics. Furthermore, 6i potently induced cell cycle arrest at G2/M phase in SGC-7901 cells. Competitive binding assays and docking studies suggested that compound 6i binds to the tubulin perfectly at the colchicine binding site. Taken together, these results revealed that 6i may become a promising lead compound for new anticancer drugs discovery.

Synthesis and antiproliferative activity of 2-aryl-4-(3,4,5-trimethoxybenzoyl)-1,2,3-triazol derivatives as microtubule-destabilizing agents

A series of 2-aryl-4-(3,4,5-trimethoxybenzoyl)-1,2,3-triazols were designed as analogs of substituted methoxybenzoyl-aryl-thiazole (SMART) under the consideration of geometric features. The target compounds were synthesized via concise and efficient processes including microwave-assisted cyclization, and were evaluated for their antiproliferative activity against three human cancer cell lines. Most compounds exhibited moderate antiproliferative activity with IC50 values in the micromolar to sub-micromolar range. Tubulin polymerization and immunofluorescence studies demonstrated that (Z)-9a was a potent microtubule-destabilizing agent and disrupted the polymerization dynamics. Moreover, (Z)-9a significantly induced accumulation of cells in the G2/M phase and caused microtubule destabilization. Molecular modeling studies showed that (Z)-9a probably binds to the colchicine site of tubulin.

Synthesis and bioevaluation of N,4-diaryl-1,3-thiazole-2-amines as tubulin inhibitors with potent antiproliferative activity

A series of N,4-diaryl-1,3-thiazole-2-amines containing three aromatic rings with an amino linker were designed and synthesized as tubulin inhibitors and evaluated for their antiproliferative activity in three human cancer cell lines. Most of the target compounds displayed moderate antiproliferative activity, and N-(2,4-dimethoxyphenyl)-4-(4-methoxyphenyl)-1,3-thiazol-2-amine (10s) was determined to be the most potent compound. Tubulin polymerization and immunostaining experiments revealed that 10s potently inhibited tubulin polymerization and disrupted tubulin microtubule dynamics in a manner similar to CA-4. Moreover, 10s effectively induced SGC-7901 cell cycle arrest at the G2/M phase in both concentration- and time-dependent manners. The molecular docking results revealed that 10s could bind to the colchicine binding site of tubulin.