(4-Methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone inhibits tubulin polymerization,induces G2/M arrest,and triggers apoptosis in human leukemia HL-60 cells

(4-Methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone (PHT) is a known cytotoxic compound belonging to the phenstatin family. However, the exact mechanism of action of PHT-induced cell death remains to be determined. The aim of this study was to investigate the mechanisms underlying PHT-induced cytotoxicity. We found that PHT displayed potent cytotoxicity in different tumor cell lines, showing IC₅₀ values in the nanomolar range. Cell cycle arrest in G₂/M phase along with the augmented metaphase cells was found. Cells treated with PHT also showed typical hallmarks of apoptosis such as cell shrinkage, chromatin condensation, phosphatidylserine exposure, increase of the caspase 3/7 and 8 activation, loss of mitochondrial membrane potential, and internucleosomal DNA fragmentation without affecting membrane integrity. Studies conducted with isolated tubulin and docking models confirmed that PHT binds to the colchicine site and interferes in the polymerization of microtubules. These results demonstrated that PHT inhibits tubulin polymerization, arrests cancer cells in G₂/M phase of the cell cycle, and induces their apoptosis, exhibiting promising anticancer therapeutic potential.     

Novel microtubule-targeted agent 6-chloro-4-(methoxyphenyl) coumarin induces G2-M arrest and apoptosis in HeLa cells

Aim:

To identify a novel coumarin analogue with the highest anticancer activity and to further investigate its anticancer mechanisms.

Methods:

The viability of cancer cells was investigated using the MTT assay. The cell cycle progression was evaluated using both flow cytometric and Western blotting analysis. Microtubule depolymerization was observed with immunocytochemistry in vivo and a tubulin depolymerization assay in vitro. Apoptosis was demonstrated using Annexin V/Propidium Iodide (PI) double-staining and sub-G₁ analysis.

Results:

Among 36 analogues of coumarin, 6-chloro-4-(methoxyphenyl) coumarin showed the best anticancer activity (IC₅₀ value about 200 nmol/L) in HCT116 cells. The compound had a broad spectrum of anticancer activity against 9 cancer cell lines derived from colon cancer, breast cancer, liver cancer, cervical cancer, leukemia, epidermoid cancer with IC₅₀ value of 75 nmol/L–1.57 μmol/L but with low cytotocitity against WI-38 human lung fibroblasts (IC₅₀ value of 12.128 μmol/L). The compound (0.04–10 μmol/L) induced G₂-M phase arrest in HeLa cells in a dose-dependent manner, which was reversible after the compound was removed. The compound (10–300 μmol/L) induced the depolymerization of purified porcine tubulin in vitro. Finally, the compound (0.04–2.5 μmol/L) induced apoptosis of HeLa cells in dose- and time-dependent manners.

Conclusion:

6-Chloro-4-(methoxyphenyl) coumarin is a novel microtubule-targeting agent that induces G₂–M arrest and apoptosis in HeLa cells.     

Fisetin‐induced apoptosis of human oral cancer SCC‐4 cells through reactive oxygen species production,endoplasmic reticulum stress,caspase‐, and mitochondria‐dependent signaling pathways

Oral cancer is one of the cancer‐related diseases in human populations and its incidence rates are rising worldwide. Fisetin, a flavonoid from natural products, has been shown to exhibit anticancer activities in many human cancer cell lines but the molecular mechanism of fisetin‐induced apoptosis in human oral cancer cells is still unclear; thus, in this study, we investigated fisetin‐induced cell death and associated signal pathways on human oral cancer SCC‐4 cells in vitro. We examined cell morphological changes, total viable cells, and cell cycle distribution by phase contrast microscopy and flow cytometry assays. Reactive oxygen species (ROS), Ca²⁺, mitochondria membrane potential (ΔΨ_m), and caspase‐8, ‐9, and ‐3 activities were also measured by flow cytometer. Results indicate that fisetin induced cell death through the cell morphological changes, caused G2/M phase arrest, induction of apoptosis, promoted ROS and Ca²⁺ production, and decreased the level of ΔΨ_m and increased caspase‐3, ‐8, and ‐9 activities in SCC‐4 cells. DAPI staining and DNA gel electrophoresis were also used to confirm fisetin‐induced cell apoptosis in SCC‐4 cells. Western blotting also found out that Fisetin increased the proapoptotic proteins such as Bax and Bid and decreased the antiapoptotic proteins such as Bcl‐2. Furthermore, results also showed that Fisetin increased the cytochrome c, AIF, and Endo G release from mitochondria in SCC‐4 cells. We also used ATF‐6α, ATF‐6β, GADD153, and GRP78 which indicated that fisetin induced cell death through ER stress. Based on those observations, we suggest that fisetin induced cell apoptosis through ER stress, mitochondria‐, and caspase‐dependent pathways.     

Determination of anticancer potential of a novel pharmacologically active thiosemicarbazone derivative against colorectal cancer cell lines

Thiosemicarbazones have received noteworthy attention due to their numerous pharmacological activities. Various thiosemicarbazone derivatives have been reported to play a key role as potential chemotherapeutic agents for the management of cancer. Herein, we aimed to establish the anticancer efficacy of novel thiosemicarbazone derivative C4 against colon cancer in vitro. The MTT viability assay identified C4 as a promising anticancer compound in a panel of cancer cell lines with the most potent activity against colon cancer cells. Further, anticancer potential of C4 was evaluated against HT-29 and SW620 colon cancer cell lines considering the factors like cell adhesion and migration, oxidative stress, cell cycle arrest, and apoptosis. Our results showed that C4 significantly inhibited the migration and adhesion of colon cancer cells. C4 significantly increased the intracellular reactive oxygen species (ROS) and induced apoptotic cell death. Cell cycle analysis revealed that C4 interfered in the cell cycle distribution and arrested the cells at the G2/M phase of the cell cycle. Consistent with these results C4 also down-regulated the Bcl-XL and Bcl-2 and up-regulated the caspase-3 expression. These findings introduced C4 as the potential anticancer agent against colon cancer.     

Eupatilin, a dietary flavonoid, induces G2/M cell cycle arrest in human endometrial cancer cells

This study is the first to investigate the antiproliferative effect of eupatilin in human endometrial cancer cells. Eupatilin, a naturally occurring flavonoid isolated from Artemisia princeps, has anti-inflammatory, anti-oxidative, and anti-tumor activities. In the present study, we investigated the potential effect of eupatilin on cell growth and its molecular mechanism of action in human endometrial cancer cells. Eupatilin was more potent than cisplatin in inhibiting cell viability in the human endometrial cancer cell lines Hec1A and KLE. Eupatilin showed relatively low cytotoxicity in normal human endometrial cells HES and HESC cells when compared to cisplatin. Eupatilin induced G2/M phase cell cycle arrest in a time- and dose-dependent manner, as indicated by flow cytometry analysis. In addition, treatment of Hec1A cells with eupatilin resulted in a significant increase in the expression of p21^WAF1/CIP1 and in the phosphorylation of Cdc25C and Cdc2. Knockdown of p21 using specific siRNAs significantly compromised eupatilin-induced cell growth inhibition. Interestingly, levels of mutant p53 in Hec1A cells decreased markedly upon treatment with eupatilin, and p53 siRNA significantly increased p21 expression. Moreover, eupatilin modulated the phosphorylation of protein kinases ERK1/2, Akt, ATM, and Chk2. These results suggest that eupatilin inhibits the growth of human endometrial cancer cells via G2/M phase cell cycle arrest through the up-regulation of p21 by the inhibition of mutant p53 and the activation of the ATM/Chk2/Cdc25C/Cdc2 checkpoint pathway.     

Anthocyanins from roselle extract arrest cell cycle G2/M phase transition via ATM/Chk pathway in p53‐deficient leukemia HL‐60 cells

Cell cycle regulation is an important issue in cancer therapy. Delphinidin and cyanidin are two major anthocyanins of the roselle plant (Hibiscus sabdariffa ). In the present study, we investigated the effect of Hibiscus anthocyanins (HAs) on cell cycle arrest in human leukemia cell line HL‐60 and the analyzed the underlying molecular mechanisms. HAs extracted from roselle calyces (purity 90%) markedly induced G2/M arrest evaluated with flow cytometry analysis. Western blot analyses revealed that HAs (0.1–0.7 mg mL^?1) induced G2/M arrest via increasing Tyr15 phosphorylation of Cdc2, and inducing Cdk inhibitors p27 and p21. HAs also induced phosphorylation of upstream signals related to G2/M arrest such as phosphorylation of Cdc25C tyrosine phosphatase at Ser216, increasing the binding of pCdc25C with 14‐3‐3 protein. HAs‐induced phosphorylation of Cdc25C could be activated by ATM checkpoint kinases, Chk1, and Chk2. We first time confirmed that ATM‐Chk1/2‐Cdc25C pathway as a critical mechanism for G2/M arrest in HAs‐induced leukemia cell cycle arrest, indicating that this compound could be a promising anticancer candidate or chemopreventive agents for further investigation. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1290–1304, 2017.     

Benzylidene derivatives of andrographolide inhibit growth of breast and colon cancer cells in vitro by inducing G(1) arrest and apoptosis

Background and purpose:

Andrographolide, the major phytoconstituent of Andrographis paniculata, was previously shown by us to have activity against breast cancer. This led to synthesis of new andrographolide analogues to find compounds with better activity than the parent compound. Selected benzylidene derivatives were investigated for their mechanisms of action by studying their effects on the cell cycle progression and cell death.

Experimental approach:

Microculture tetrazolium, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and sulphorhodamine B (SRB) assays were utilized in assessing the in vitro growth inhibition and cytotoxicity of compounds. Flow cytometry was used to analyse the cell cycle distribution of control and treated cells. CDK1 and CDK4 levels were determined by western blotting. Apoptotic cell death was assessed by fluorescence microscopy and flow cytometry.

Key results:

Compounds, in nanomolar to micromolar concentrations, exhibited growth inhibition and cytotoxicity in MCF-7 (breast) and HCT-116 (colon) cancer cells. In the NCI screen, 3,19-(2-bromobenzylidene) andrographolide (SRJ09) and 3,19-(3-chloro-4-fluorobenzylidene) andrographolide (SRJ23) showed greater cytotoxic potency and selectivity than andrographolide. SRJ09 and SRJ23 induced G₁ arrest and apoptosis in MCF-7 and HCT-116 cells, respectively. SRJ09 downregulated CDK4 but not CDK1 level in MCF-7 cells. Apoptosis induced by SRJ09 and SRJ23 in HCT-116 cells was confirmed by annexin V-FITC/PI flow cytometry analysis.

Conclusion and implications:

The new benzylidene derivatives of andrographolide are potential anticancer agents. SRJ09 emerged as the lead compound in this study, exhibiting anticancer activity by downregulating CDK4 to promote a G₁ phase cell cycle arrest, coupled with induction of apoptosis.     

Investigation of anti-tumor mechanisms of K2154: characterization of tubulin isotypes, mitotic arrest and apoptotic machinery

Microtubules are crucial targets for cancer chemotherapeutic drugs, and new microtubule-directed agents are of continued interest in drug development. A novel microtubule-directed agent, ethyl-2-[N-ρ-chlorobenzyl-(2′-methoxy)]-anilino-4-oxo -4, 5-dihydro-furan-3-carboxylate, was identified. The compound, designated K2154, inhibited cell proliferation, with IC₅₀ values of 10.3, 15.3, 9.6, 11.2, 12.8 and 12.1 μM in prostate cancer PC-3, hepatocellular carcinoma Hep3B, non-small cell lung cancer A549, colorectal cancer HT29 and HCT116, and P-glycoprotein-rich breast cancer NCI/ADR-RES cells, respectively. Because NCI/ADR-RES cells were susceptible to inhibition by K2154, it indicated that this compound is a poor substrate for P-glycoprotein. In this study, PC-3 cells were used to identify the anticancer mechanisms of K2154. K2154 induced an arrest of the cell cycle at G2/M phase and a subsequent increase of hypodiploid phase in PC-3 cells, whereas it only induced a moderate level of G2/M arrest with little increase of hypodiploid phase in normal prostate cells. K2154 inhibited microtubule assembly in both in vitro turbidity assay and in vivo microtubule spin-down experiment. Immunochemical examination showed that K2154 caused formation of abnormal mitotic characteristics with bipolar spindles, particularly, in β_II- and β_III-tubulin staining. It also induced several pathways, including cyclin B1 up-regulation, dephosphorylation on Tyr¹⁵ and phosphorylation on Thr¹⁶¹ of Cdk1 and Cdc25C phosphorylation, and roscovitine (a Cdk1 inhibitor) significantly inhibited K2154-induced apoptosis, suggesting a pro-apoptotic role of Cdk1. Phosphorylation of Bcl-2 and Bcl-xL and cleavage of Mcl-1, together with activation of caspase-9 and -3, indicated that mitochondrial pathway played a central role in K2154-mediated apoptotic cell death. Additionally, AIF contributed to a late phase of K2154-induced apoptotic pathway. In conclusion, it is suggested that K2154 displays an anticancer activity through a target on microtubules and a subsequent signaling cascade on cell cycle regulation and apoptotic machinery.     

Benzyl isothiocyanate promotes apoptosis of oral cancer cells via an acute redox stress-mediated DNA damage response

Benzyl isothiocyanate (BITC) is a cruciferous vegetable-derived compound with anticancer properties in human cancer cells. However, its anticancer potential and underlying mechanisms remain absent in human oral cancer cells. Results indicate that BITC inhibits growth, promotes G₂/M phase arrest and triggers apoptosis of OC2 cells with a minimal toxicity to normal cells. BITC-induced cell death was completely prevented by pretreatment with thiol-containing redox compounds including N-acetyl-l-cysteine (NAC), glutathione (GSH), dithiothreitol, and 2-mercaptoethanol, but not free radical scavengers mito-TEMPO, catalase, apocynin, l-NAME and mannitol. BITC rapidly produced reactive oxygen species and nitric oxide, triggered oxidative DNA damage. BITC effectively decreased the intracellular GSH and GSH/GSSG ratio and redox balance recovery by thiol-containing redox compounds, but not by free radical scavengers. Accordingly, redox stresses-DNA damage response (DDR) activated ATM, Chk2, p53, and p21 and subsequently resulted in G₂/M phase arrest by inhibiting Cdc2 and cyclin B1. Notably, BITC-induced apoptosis was associated with reduced Mcl-1 and Bcl-2 expression, diminished mitochondrial membrane potential (ΔΨm), and increased PARP cleavage. These BITC-induced redox stress-mediated DDR and apoptosis could be blocked by NAC and GSH. Therefore, BITC can be a rational drug candidate for oral cancer and acted via a redox-dependent pathway.     

Heterocyclic Organobismuth(III) Compound Targets Tubulin to Induce G2/M Arrest in HeLa Cells

Our previous study showed that organobismuth compounds induce apoptosis in human promyelocytic leukemia cells, although solid tumor cell lines were relatively resistant. Herein, we investigated the primary cellular target of these compounds in HeLa cells. One organobismuth compound, bi-chlorodibenzo[c,f][1,5]thiabismocine (compound 3), arrested the cell cycle at G₂/M as assessed by flow cytometry and by upregulating the expression of cyclin B1. At a low concentration (0.5 μM), compound 3 caused cell cycle arrest at the mitotic phase and induced apoptosis. At a higher concentration (>1.0 μM), it induced an arrest in the G₂/M phase, leading to apoptosis. In many cells blocked at the M phase, the organization of microtubules was affected, indicating depolymerization of the microtubule network. Western blotting demonstrated that compound 3 depolymerized microtubules similar to colchicine and nocodazole. Experiments in vitro also showed that compound 3 inhibited the assembly of purified tubulin in a concentration-dependent manner by interacting with the colchicine-binding site of tubulin through its SH groups. Heterocyclic organobismuth compounds are novel tubulin ligands.     

WTC-01, a novel synthetic oxime-flavone compound,destabilizes microtubules in human nasopharyngeal carcinoma cells in vitro and in vivo

Background and Purpose

Dynamic polymerization of microtubules is essential for cancer cell growth and metastasis, and microtubule-disrupting agents have become the most successful anti-cancer agents in clinical use. Besides their antioxidant properties, flavonoids also exhibit strong microtubule-disrupting activity and inhibit tumour growth. We have designed, synthesized and tested a series of oxime/amide-containing flavone derivatives. Here we report the evaluation of one compound, WTC-01 for its anti-proliferative effects in human cancer cells.

Experimental approach

We used a range of cancer cell lines including two human nasopharyngeal carcinoma (NPC) cell lines, measuring proliferation, cell cycle and apoptosis, along with caspase levels and mitochondrial membrane potentials. Assays of tubulin polymerisation in vitro and computer modelling of the colchicine binding site in tubulin were also used. In mice, pharmacokinetics and growth of NPC-derived tumours were studied.

Key Results

WTC-01 was most potent against proliferation of NPC cells (IC₅₀ = 0.45 μM), inducing accumulation of cells in G₂/M and increasing apoptosis, time- and concentration-dependently. The colchicine competition-binding experiments and computer modelling results suggested that WTC-01 causes microtubule disruption via binding to the colchicine-binding site of tubulin resulting in mitochondrial membrane damage and cell apoptosis via activation of caspase-9/-3 without noticeable activation of the caspase-8. Notably, our in vivo studies demonstrated that at doses of 25 and 50 mg·kg⁻¹, WTC-01 exhibited good pharmacokinetic properties and completely inhibited the growth of NPC-TW01 cells in a xenograft nude mouse model.

Conclusions and Implications

WTC-01, a new synthetic oxime-containing flavone, exhibited potent anti-tumour activity against NPC cells and merits further investigation.     

APOPTOSIS INDUCTION AND CELL CYCLE PERTURBATION IN ESTABLISHED CELL LINES BY PEROXYSOMICINE A1 (T-514)†

Peroxysomicine A1, a novel potential anticancer compound induced cell death in established cell lines and in a primary culture of rat neonatal cardiomyocytes. Non-transformed cells are less sensitive to the compound than transformed cell lines. Fluorescent microscopy of dying cells stained with DNA-specific dyes revealed chromatin condensation and nuclear fragmentation as well as membrane blebbing characteristic of apoptosis. Flow cytometry of cells treated with peroxysomicine A1, demonstrated appearance of cells containing less than 2C DNA, that indicated degradation of nuclear DNA, another hallmark of apoptotic cell death. Z-VAD, a nonspecific caspase inhibitor, prevented DNA fragmentation but not cell death registered by permeabilization of cell outer membrane. Peroxysomicine A1 also inhibited proliferation of various cell lines. Flow cytometry analysis showed significant accumulation of dividing cells in G₂/M phases of cell cycle indicating, most likely delay in G₂. These results provide initial insight into the mechanisms of action of peroxysomicine A1 and suggest that peroxysomicine A1 is a potent inhibitor of cell proliferation and inducer of apoptosis and may be a useful antineoplastic chemotherapeutic agent.     

AMP-activated protein kinase-dependent and -independent mechanisms underlying in vitro antiglioma action of compound C

We investigated the effect of compound C, a well-known inhibitor of the intracellular energy sensor AMP-activated protein kinase (AMPK), on proliferation and viability of human U251 and rat C6 glioma cell lines. Compound C caused G₂/M cell cycle block, accompanied by apoptotic glioma cell death characterized by caspase activation, phosphatidylserine exposure and DNA fragmentation. The mechanisms underlying the pro-apoptotic action of compound C involved induction of oxidative stress and downregulation of antiapoptotic molecule Bcl-2, while no alteration of pro-apoptotic Bax was observed. Compound C diminished AMPK phosphorylation and enzymatic activity, resulting in reduced phosphorylation of its target acetyl CoA carboxylase. AMPK activators metformin and AICAR partly prevented the cell cycle block, oxidative stress and apoptosis induced by compound C. The small interfering RNA (siRNA) targeting of human AMPK mimicked compound C-induced G₂/M cell cycle arrest, but failed to induce oxidative stress and apoptosis in U251 glioma cells. In conclusion, our data indicate that AMPK inhibition is required, but not sufficient for compound C-mediated apoptotic death of glioma cells.     

Induction of G2/M phase arrest and apoptosis by a novel indoloquinoline derivative,IQDMA, in K562 cells

The indoloquinoline, IQDMA (N′‐(11H‐indolo[3,2‐c]quinolin‐6‐yl)‐N,N‐dimethylethane‐1,2‐diamine), was identified as a novel antineoplastic agent with broad spectrum of antitumor activities against several human cancer cells. IQDMA‐induced G2/M arrest was accompanied by up‐regulation of the cyclin‐dependent kinase inhibitors (CDKIs), p21 and p27, and down‐regulation of Cdk1and Cdk2. IQDMA had no effect on the levels of cyclin A, cyclin B1, cyclin D3, or Cdc25C. IQDMA also increased apoptosis, as characterized by apoptotic body formation, increase of the sub G₁ population and poly (ADP‐ribose) polymerase (PARP) cleavage. Further mechanistic analysis demonstrated that IQDMA upregulated FasL protein expression, and kinetic studies showed the sequential activation of caspases‐8, ‐3, and ‐9. Both caspase‐8 and caspase‐3 inhibitors, but not a caspase‐9‐specific inhibitor, suppressed IQDMA‐induced cell death. These molecular alterations provide an insight into IQDMA‐caused growth inhibition, G2/M arrest, and apoptotic death of K562 cells. Drug Dev. Res. 67:743–751, 2006. © 2006 Wiley‐Liss, Inc.     

Licochalcone B inhibits growth of bladder cancer cells by arresting cell cycle progression and inducing apoptosis

To examine the mechanisms by which licochalcone B (LCB) inhibits the proliferation of human malignant bladder cancer cell lines (T24 and EJ) in vitro and antitumor activity in vivo in MB49 (murine bladder cancer cell line) tumor model. Exposure of T24 or EJ cells to LCB significantly inhibited cell lines proliferation in a concentration-dependent and time-dependent manner, and resulted in S phase arrest in T24 or EJ cells, respectively. LCB treatment decreased the expression of cyclin A, cyclin-dependent kinase (CDK1 and CDK2) mRNA, cell division cycle 25 (Cdc25A and Cdc25B) protein. In addition, LCB treatment down-regulated Bcl-2 and survivin expression, enhanced Bax expression, activated caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP) protein. Consistently, the tumorigenicity of LCB-treated MB49 cells was limited significantly by using the colony formation assay in vitro and the MB49 tumor model performed in C57BL/6 mice in vivo. These findings provide support for the use of LCB in chemoprevention and bladder cancer therapy.     

Withaferin A modulates the Spindle Assembly Checkpoint by degradation of Mad2–Cdc20 complex in colorectal cancer cell lines

Withania somnifera L. Dunal (Ashwagandha) is used over centuries in the ayurvedic medicines in India. Withaferin A, a withanolide, is the major compound present in leaf extract of the plant which shows anticancer activity against leukemia, breast cancer and colorectal cancer. It arrests the ovarian cancer cells in the G2/M phase in dose dependent manner. In the current study we show the effect of Withaferin A on cell cycle regulation of colorectal cancer cell lines HCT116 and SW480 and its effect on cell fate. Treatment of these cells with this compound leads to apoptosis in a dose dependent manner. It causes the G2/M arrest in both the cell lines. We show that Withaferin A (WA) causes mitotic delay by blocking Spindle assembly checkpoint (SAC) function. Apoptosis induced by Withaferin A is associated with proteasomal degradation of Mad2 and Cdc20, an important constituent of the Spindle Checkpoint Complex. Further overexpression of Mad2 partially rescues the deleterious effect of WA by restoring proper anaphase initiation and keeping more number of cells viable. We hypothesize that Withaferin A kills cancer cells by delaying the mitotic exit followed by inducing chromosome instability.     

Design,Synthesis and Biological Evaluation of Cinnamic Acyl Shikonin Derivatives

Inducing apoptosis is an important and promising therapeutic approach to overcome cancer. Here, we described a series of novel synthesized compounds, cinnamic acyl shikonin derivatives ( 1b – 19b ), which were synthesized starting from shikonin and cinnamic acids, which exhibit anticancer activity via inducing apoptosis in vitro. Our flow cytometry results showed that compound 8b ((E)‐1‐(5,8‐dihydroxy‐1,4‐dioxo‐1,4‐dihydronaphthalen‐2‐yl)‐4‐methylpent ‐3‐enyl‐3‐(3‐(trifluoromethyl) phenyl)acrylate) (IC₅₀ = 0.69, 0.65, 1.62 μm for human SW872‐s, A875 and A549 cell lines, respectively) exhibited conspicuous anticancer activities and has low cell toxicity in vitro. Therefore, we considered that compound 8b is potentially to be a candidate of anticancer agent. The proliferation inhibitory effect of compound 8b was associated with its apoptosis‐inducing effect by activating caspase‐3, caspase‐7, caspase‐9, and PARP. When the level of cleaved caspase‐3, cleaved caspase‐7, cleaved caspase‐9, and cleaved PARP are rise, apoptosis of cancer cells will be induced.     

Design,synthesis, and biological evaluation of tetrahydroisoquinoline stilbene derivatives as potential antitumor candidates

Herein, a novel class of tetrahydroisoquinoline stilbene derivatives were synthesized, and their potential in vitro anticancer activities were evaluated. Most of the compounds displayed inhibitory activity against one or more representative human cancer cell lines (lung cancer A549 cells, breast cancer MCF-7 cells, and human colorectal carcinoma HT-29 cells), especially compound 16 e, which exhibited outstanding cytotoxicity to A549 cells. The tubulin polymerization assay demonstrated that compound 16 e displayed better inhibition than colchicine when tested at the same concentration. It was found that 16 e arrested A549 cells in G2/M phase by downregulating the expression of cell division cycle 2 (Cdc2) and upregulating the expression of proliferating cell nuclear antigen (PCNA) and cyclin B1. Flow cytometry and Western blot analysis indicated that 16 e caused apoptosis via the mitochondrial-dependent apoptotic pathway by reducing mitochondrial membrane potential, inducing ROS accumulation, promoting the release of cytochrome C from the mitochondria into the cytoplasm, and further increasing the protein level of cleaved caspase-3. This work may inspire new ideas for the further improvement of tubulin-related anticancer drugs and treatments.     

Synthesis,molecular docking and biological evaluation of 1‐phenylsulphonyl‐2‐(1‐methylindol‐3‐yl)‐benzimidazole derivatives as novel potential tubulin assembling inhibitors

A series of new 1‐phenylsulphonyl‐2‐(1‐methylindol‐3‐yl)‐benzimidazole derivatives were designed, synthesized and evaluated as potential inhibitors of tubulin polymerization and anthropic cancer cell lines. Among them, compound 33 displayed the most potent tubulin polymerization inhibitory activity in vitro (IC₅₀ = 1.41 μM) and strong antiproliferative activities against A549, Hela, HepG2 and MCF‐7 cell lines in vitro with GI₅₀ value of 1.6, 2.7, 2.9 and 4.3 μM, respectively, comparable with the positive control colchicine (GI₅₀ value of 4.1, 7.2, 9.5 and 14.5 μM, respectively) and CA‐4 (GI₅₀ value of 2.2, 4.3, 6.4 and 11.4 μM, respectively). Simultaneously, we evaluated that compound 33 could effectively induce apoptosis of A549 associated with G2/M phase cell cycle arrest. Immunofluorescence microscopy also clearly indicated compound 33 a potent antimicrotubule agent. Docking simulation showed that compound 33 could bind tightly with the colchicine‐binding site and act as a tubulin inhibitor. Three‐dimensional‐QSAR model was also built to provide more pharmacophore understanding that could be used to design new agents with more potent tubulin assembling inhibitory activity in the future.     

LL-202, a newly synthesized flavonoid,inhibits tumor growth via inducing G2/M phase arrest and cell apoptosis in MCF-7 human breast cancer cells in vitro and in vivo

We recently established that LL-202, a newly synthesized flavonoid, exhibited obvious anticancer effects against human breast cells in vivo and in vitro. The underlying mechanism of its anticancer activity remains to be elucidated. In this study, we demonstrated that LL-202 inhibited the growth and proliferation of human breast cancer MCF-7 cells in a concentration and time-dependent manner. We reported that LL-202 induced both mitochondrial- and death-receptor-mediated apoptosis, which were characterized by the dissipation of mitochondrial membrane potential (ΔΨ_m), cytochrome c (Cyt c) release from mitochondria to cytosol, the activation of several caspases and induction of poly (ADP-ribose) polymerase (PARP) and Bid cleavage. N-acetylcysteine (NAC), a general ROS scavenger, partly blocked the LL-202-induced ROS levels and apoptosis. In addition, LL-202 induced arrest in cell cycle progression at G2/M phase in MCF-7 cells. After the treatment with LL-202, the expression of cell cycle-related proteins, such as cyclin B1, cyclin A, and p-CDK1 (Thr161) were down-regulated, whereas the expression of p21^WAF1/Cip1 and p-CDK1 (Thr14/Tyr15) were up-regulated. Finally, in vivo studies, LL-202 significantly suppressed the growth of MCF-7 breast cancer xenograft tumors in a dose-dependent manner with low systemic toxicity. In conclusion, the results showed that LL-202 had significant anticancer effects against human breast cells via the induction of apoptosis and G2/M phase arrest and it may be a novel anticancer agent for treatment of breast cancer.