Jolkinolide B induces cell cycle arrest and apoptosis in MKN45 gastric cancer cells and inhibits xenograft tumor growth in vivo

Gastric cancer is one of the most common digestive carcinomas throughout the world and represents high mortality. There is an urgent quest for seeking a novel and efficient antigastric cancer drug. Euphorbia fischeriana Steud had long been used as a traditional Chinese medicine for the treatment of cancer. According to the basic theory of traditional Chinese medicine, its antitumor mechanism is ‘to combat poison with poison’. However, its effective material foundation of it is still ambiguous. In our previous work, we studied the chemical constituents of E. fischeriana Steud. Jolkinolide B (JB) is an ent-abietane-type diterpenoid we isolated from it. The purpose of the present study was to investigate the antigastric effect and mechanism of JB. Results revealed that JB could suppress the proliferation of MKN45 cells in vitro and inhibit MKN45 xenograft tumor growth in nude mice in vivo. We further investigated its anticancer mechanism. On the one hand, JB caused DNA damage in gastric cancer MKN45 cells and induced the S cycle arrest by activating the ATR-CHK1-CDC25A-Cdk2 signaling pathway, On the other hand, JB induced MKN45 cells apoptosis through the mitochondrial pathway, and ultimately effectively inhibited the growth of gastric cancer cells. These results suggest that JB appears to be a promising candidate drug with antigastric cancer activity and warrants further research.     

A novel pro-apoptosis gene PNAS4 that induces apoptosis in A549 human lung adenocarcinoma cells and inhibits tumor growth in mice

The gene PNAS4 is a high conservative gene that shares high homology of sequence in various organisms from plants to animals. We found overexpression of human PNAS4 induced apoptosis and arrested cell cycle in S phase in A549 human lung adenocarcinoma cells. In C57BL/6 mice model of Lewis lung carcinoma, overexpression of mouse PNAS4 significantly suppressed tumor growth and prolonged survival time through induction of tumor cell apoptosis, exhibiting effective antitumor. Our original investigations in vitro and vivo indicated PNAS4 is a novel pro-apoptosis gene, which could be used as a potential target of cancer biotherapy in future.     

Inhibition of cellular proliferation and induction of apoptosis in human lung adenocarcinoma A549 cells by T-type calcium channel antagonist

The anti-proliferative and apoptotic activities of new T-type calcium channel antagonist, 6e (BK10040) on human lung adenocarcinoma A549 cells were investigated. The MTT assay results indicated that BK10040 was cytotoxic against human lung adenocarcinoma (A549) and pancreatic cancer (MiaPaCa2) cells in a dose-dependent manner with IC₅₀ of 2.25 and 0.93 μM, respectively, which is ca. 2-fold more potent than lead compound KYS05090 despite of its decreased T-type calcium channel blockade. As a mode of action for cytotoxic effect of BK10040 on lung cancer (A549) cells, this cancer cell death was found to have the typical features of apoptosis, as evidenced by the accumulation of positive cells for annexin V. In addition, BK10040 triggered the activations of caspases 3 and 9, and the cleavages of poly (ADP-ribose) polymerase (PARP). Moreover, the treatment with z-VAD-fmk (a broad spectrum caspase inhibitor) significantly prevented BK10040-induced apoptosis. Based on these results, BK10040 may be used as a potential therapeutic agent for human lung cancer via the potent apoptotic activity.     

p38 inhibitor SB203580 sensitizes the resveratrol-induced apoptosis in human lung adenocarcinoma (A549) cells

Based on our recent findings that resveratrol, a natural plant polyphenol found in red grape skins as well as other food products, induces apoptosis via a caspase-independent intrinsic pathway in human lung adenocarcinoma cells, this study is designed to explore whether SB203580, a p38 inhibitor, potentiates the resveratrol-induced apoptosis of human lung adenocarcinoma (A549) cells. We found that pretreatment with SB203580 enhanced the resveratrol-induced apoptosis by accelerating the intrinsic apoptotic pathway including Bax activation, loss of mitochondrial membrane potential, and activation of both caspase-9 and -3. Although treatment with resveratrol alone did not induce caspase-8 activation, cotreatment with both SB203580 and resveratrol not only enhanced FasL cleavage but also activated caspase-8, indicating that the extrinsic apoptotic pathway may be involved in the synergistic effect. Collectively, we for the first time demonstrate that SB203580 synergistically enhances the resveratrol-induced apoptosis by accelerating Bax-mediated intrinsic pathway and initiating extrinsic pathway, suggesting a possible alternative therapeutic strategy for human lung cancer.     

Transforming growth factor beta-1 enhances cytotoxic effect of doxorubicin in human lung adenocarcinoma cells of A549 line

Transforming growth factor beta-1 (TGFbeta-1) is a regulator of cell proliferation, differentiation and apoptosis. Doxorubicin (adriamycin), an anthracycline drug causing double-strand DNA breaks, is widely used in anticancer chemotherapy. Here we demonstrated that TGFbeta-1 enhanced cytotoxic (proapoptotic) action of doxorubicin towards cultured human lung carcinoma A549 cells. Western-blot analysis and immunocytochemistry were used to show that doxorubicin induced PARP degradation in A549 cells, and TGFbeta-1 enhanced that action of the drug. The obtained results suggest a possibility of biomodulating effect of TGFbeta-1 on tumor cell treatment with doxorubicin.     

Furano-1,2-naphthoquinone inhibits EGFR signaling associated with G2/M cell cycle arrest and apoptosis in A549 cells

Furano-1,2-naphthoquinone (FNQ), prepared from 2-hydroxy-1,4-naphthoquinone and chloroacetaldehyde in an efficient one-pot reaction, exhibits an anti-carcinogenic effect. FNQ exerted anti-proliferative activity with the G(2)/M cell cycle arrest and apoptosis in A549 cells. FNQ-induced G(2)/M arrest was correlated with a marked decrease in the expression levels of cyclin A and cyclin B, and their activating partner cyclin-dependent kinases (Cdk) 1 and 2 with concomitant induction of p53, p21, and p27. FNQ-induced apoptosis was accompanied with Bax up-regulation and the down-regulation of Bcl-2, X-linked inhibitor of apoptosis (XIAP), and survivin, resulting in cytochrome c release and sequential activation of caspase-9 and caspase-3. Western blot analysis revealed that FNQ suppressed EGFR phosphorylation and JAK2, STAT3, and STAT5 activation, but increased in activation of p38 MAPK and c-Jun NH2-terminal kinase (JNK) stress signal. The combined treatment of FNQ with AG1478 (a specific EGFR inhibitor) significantly enhanced the G(2)/M arrest and apoptosis, and also led to up-regulation in Bax, p53, p21, p27, release of mitochondrial cytochrome c, and down-regulation of Bcl-2, XIAP, survivin, cyclin A, cyclin B, Cdk1, and Cdk2 in A549 cells. These findings suggest that FNQ-mediated cytotoxicity of A549 cell related with the G(2)/M cell cycle arrest and apoptosis via inactivation of EGFR-mediated signaling pathway.     

The role of reactive oxygen species (ROS) production on diallyl disulfide (DADS) induced apoptosis and cell cycle arrest in human A549 lung carcinoma cells

Diallyl disulfide (DADS), an oil soluble constituent of garlic (Allium sativum), has been reported to cause antimutagentic and anticarcinogenic effects in vitro and in vivo by modulating phases I and II enzyme activities. In recent years, several studies suggested that the chemopreventive effects of DADS can also be attributed to induction of cell cycle arrest and apoptosis in cancer cells. In the present study, we reported that DADS-induced cell cycle arrest at G2/M and apoptosis in human A549 lung cancer cells in a time- and dose-dependent manner. Additionally, a significant increase of intracellular reactive oxygen species (ROS) was induced in A549 cells less than 0.5 h after DADS treatment, indicating that ROS may be an early event in DADS-modulated apoptosis. Treatment of A549 cells with N-acetyl cysteine (NAC) completely abrogated DADS-induced cell cycle arrest and apoptosis. The result indicated that oxidative stress modulates cell proliferation and cell death induced by DADS.     

Effects of karanjin on cell cycle arrest and apoptosis in human A549, HepG2 and HL-60 cancer cells

Background

We have investigated the potential anticancer effects of karanjin, a principal furanoflavonol constituent of the Chinese medicine Fordia cauliflora, using cytotoxic assay, cell cycle arrest, and induction of apoptosis in three human cancer cell lines (A549, HepG2 and HL-60 cells).

Results

MTT cytotoxic assay showed that karanjin could inhibit the proliferation and viability of all three cancer cells. The induction of cell cycle arrest was observed via a PI (propidium iodide)/RNase Staining Buffer detection kit and analyzed by flow cytometry: karanjin could dose-dependently induce cell cycle arrest at G2/M phase in the three cell lines. Cell apoptosis was assessed by Annexin V-FITC/PI staining: all three cancer cells treated with karanjin exhibited significantly increased apoptotic rates, especially in the percentage of late apoptosis cells.

Conclusion

Karanjin can induce cancer cell death through cell cycle arrest and enhance apoptosis. This compound may be effective clinically for cancer pharmacotherapy.     

Arsenic trioxide inhibits the growth of A498 renal cell carcinoma cells via cell cycle arrest or apoptosis

Previously, we showed that arsenic trioxide potently inhibited the growth of myeloma cells and head and neck cancer cells. Here, we demonstrate that arsenic trioxide inhibited the proliferation of all the renal cell carcinoma cell lines (ACHN, A498, Caki-2, Cos-7, and Renca) except only one cell line (Caki-1) with IC(50) of about 2.5-10 microM. Arsenic trioxide induced a G(1) or a G(2)-M phase arrest in these cells. When we examined the effects of this drug on A498 cells, arsenic trioxide (2.5 microM) decreased the levels of CDK2, CDK6, cyclin D1, cyclin E, and cyclin A proteins. Although p21 protein was not increased by arsenic trioxide, this drug markedly enhanced the binding of p21 with CDK2. In addition, the activities of CDK2- and CDK6-associated kinase were reduced in association with hypophosphorylation of Rb protein. Arsenic trioxide (10 microM) also induced apoptosis in A498 cells. Apoptotic process of A498 cells was associated with the changes of Bcl-(XL), caspase-9, caspase-3, and caspase-7 proteins as well as mitochondria transmembrane potential (Deltapsi(m)) loss. Taken together, these results demonstrate that arsenic trioxide inhibits the growth of renal cell carcinoma cells via cell cycle arrest or apoptosis.     

Taiwanin A induced cell cycle arrest and p53-dependent apoptosis in human hepatocellular carcinoma HepG2 cells

Taiwanin A, a lignan isolated from Taiwania cryptomerioides Hayata, has previously been reported to have cytotoxicity against human tumor cells, but the mechanisms are unclear. In this study, we examined the molecular mechanism of cell death of human hepatocellular carcinoma HepG2 cells induced by Taiwanin A. Taiwanin A has been found to induce cell cycle arrest at G2/M phase as well as caspase-3-dependent apoptosis within 24 h. We performed both in vitro turbidity assay and immunofluorescence staining of tubulin to show that Taiwanin A can inhibit microtubule assembly. Moreover, the tumor suppressor protein p53 in HepG2 cells was activated by Taiwanin A within 12 h. Inhibition of p53 by either pifithrin-alpha or by short hairpin RNA which blocks p53 expression attenuates Taiwanin A cytotoxicity. Our results demonstrate that Taiwanin A can act as a new class of microtubule damaging agent, arresting cell cycle progression at mitotic phase and inducing apoptosis through the activation of p53.     

Escin reduces cell proliferation and induces apoptosis on glioma and lung adenocarcinoma cell lines

Aesculus hippocastanum (the horse chestnut) seed extract has a wide variety of biochemical and pharmacological effects including anti-inflammatory, antianalgesic, and antipyretic activities. The main active compound of this plant is escin. It is known that several medicinal herbs with anti-inflammatory properties have been found to have a role in the prevention and treatment of cancer. In the present study, the cytotoxic effects of escin in the C6 glioma and A549 cell lines were analyzed by MTT. Apoptotic effects of escin on both cell lines were evaluated by Annexin V binding capacity with flow cytometric analysis. Structural and ultrastructural changes were also evaluated using transmission electron microscopy. The results indicated that escin has potent antiproliferative effects against C6 glioma and A549 cells. These effects are both dose and time dependent. Taken together, escin possesses cell cycle arrest on G0/G1 phase and selective apoptotic activity on A549 cells as indicated by increased Annexin V-binding capacity, bax protein expression, caspase-3 activity and morphological changes obtained from micrographs by transmission electron microscopy.     

Silencing of the type 1 insulin-like growth factor receptor increases the sensitivity to apoptosis and inhibits invasion in human lung adenocarcinoma A549 cells

The type 1 insulin-like growth factor receptor (IGF-1R), which is over-expressed or activated in many human cancers, including lung cancer, mediates cancer cell proliferation and metastasis. Several studies indicate that blocking IGF-1R expression can inhibit tumor cell proliferation and metastasis. In this study, inhibition of the endogenous IGF-1R by recombinant adenoviruses encoding short hairpin RNAs against IGF-1R was found to significantly suppress IGF-1R expression, arrest the cell cycle, enhance the apoptotic response, and inhibit proliferation, adhesion, invasion and migration in A549 cells. Moreover, silencing IGF-1R decreases the expression of invasive-related genes including matrix metalloproteinase-2 (MMP-2), MMP-9, and urokinase-plasminogen activator (u-PA), and the phosphorylation of Akt and ERK1/2. These results suggest that the silencing of IGF-1R has the potential to be an effective cancer gene therapy strategy for human lung cancer.     

Swertia chirayita suppresses the growth of non-small cell lung cancer A549 cells and concomitantly induces apoptosis via downregulation of JAK1/STAT3 pathway

Lung carcinoma is the leading cause of cancer-related mortalities worldwide, and present therapeutical interventions are not successful enough to treat this disease in many cases. Recent years have witnessed a surge in exploring natural compounds for their antiproliferative efficacy to expedite the characterization of novel anticancer chemotherapeutics. Swertia chirayita is a valued medicinal herb and possess intrinsic pharmaceutical potential. However, elucidation of its anticancer effects at molecular levels remains unclear and needs to be investigated. We assessed the anticancer and apoptotic efficacy of S. chirayita ethanolic extract (Sw-EtOH) on non-small cell lung cancer (NSCLC) A549 cells during this exploratory study. The results elucidated that S. chirayita extract induced toxic effects within lung cancer cells by ~1 fold during cytotoxicity and LDH release assay at a 400 μg/ml concentration. Sw-EtOH extract elevates the level of ROS, resulting in the disruption of Δψm and release of cytosolic cytochrome c by 3.15 fold. Activation of caspases-3, -8 & -9 also escalated by ~1 fold, which further catalyze the augmentation of PARP cleavage (~3 folds), resulting in a four-fold increase in Sw-EtOH induced apoptosis. The gene expression analysis further demonstrated that Sw-EtOH extracts inhibited JAK1/STAT3 signaling pathway by down-regulating the levels of JAK1 and STAT3 to nearly half a fold. Treatment of Sw-EtOH modulates the expression level of various STAT3 associated proteins, including Bcl-XL, Bcl-2, Mcl-1, Bax, p53, Fas, Fas-L, cyclinD1, c-myc, IL-6, p21 and p27 in NSCLC cells. Thus, our study provided a strong impetus that Sw-EtOH holds the translational potential of being further evaluated as efficient cancer therapeutics and a preventive agent for the management of NSCLC.     

Synthesis of novel pyrazole carboxamide derivatives and discovery of modulators for apoptosis or autophagy in A549 lung cancer cells

A series of novel 3-aryl-1-arylmethyl-1H-pyrazole-5-carboxamide derivatives 3a–l, were synthesized by the reaction of 3-aryl-1-arylmethyl-1H-pyrazole-5-carbonyl chloride with substituted amine in excellent yields. The compounds 3e–h could suppress A549 lung cancer cell growth. More interestingly, compounds 3e and 3f might inhibit the A549 cell growth by inducing apoptosis; whereas compounds 3g and 3h with fluorine group might inhibit the A549 cell growth by inducing autophagy.     

Induction of nucleolin translocation by acharan sulfate in A549 human lung adenocarcinoma

Acharan sulfate (AS), isolated from the giant African snail Achatina fulica, is a novel glycosaminoglycan, consisting primarily of the repeating disaccharide structure α‐D ‐N‐acetylglucosaminyl (1 → 4) 2‐sulfoiduronic acid. AS shows anti‐tumor activity in vitro and in vivo. Despite this activity, AS is only weakly cytotoxic towards cancer cells. We examine the interactions between AS and cell‐surface proteins in an effort to explain this anti‐tumor activity. Using flow cytometry and affinity column chromatography, we confirm that AS has strong affinity to specific cell‐surface proteins including nucleolin (NL) in A549 human lung adenocarcinomas. Surprisingly, we found the translocation of NL from nucleus to cytoplasm under the stimulation of AS (100 µg/ml) in vitro. Also, as NL exits the nucleus, the levels of growth factors such as bFGF and signaling cascade proteins, such as p38, p53, and pERK, are altered. These results suggest that the communication between AS and NL plays a critical role on signal transduction in tumor inhibition. J. Cell. Biochem. 110: 1272–1278, 2010. Published 2010 Wiley‐Liss, Inc.