• Asian Pacific Journal of Cancer Prevention
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• v.16 no.14
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• pp.5883-5887
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• 2015

Despite recent advances in therapeutic strategies for lung cancer, mortality still is increasing. In the present study, we investigated the anti-cancer effects of KMU-193, 2-(4-Ethoxy-phenyl)-N-{5-[2-fluoro-4-(4-methylpiperazine-1-carbonyl)-phenylamino]-1H-indazol-3-yl}-acetamide in a human non-small cell lung cancer cell line A549. KMU-193 strongly inhibited the proliferation of A549 cells, but it did not have anti-proliferative effect in other types of cancer cell lines. KMU-193 further induced apoptosis in association with activation of caspase-3 and cleavage of PLC-${\gamma}1$. However, KMU-193 had no apoptotic effect in untransformed cells such as TMCK-1 and BEAS-2B. Interestingly, pretreatment with z-VAD-fmk, a pan-caspase inhibitor, strongly abrogated KMU-193-induced apoptosis. KMU-193 treatment enhanced the expression levels of p53 and PUMA. Importantly, p53 siRNA transfection attenuated KMU-193-induced apoptosis. Collectively, these results for the first time demonstrate that KMU-193 has strong apoptotic effects on A549 cells and these are largely mediated through caspase-3- and p53-dependent pathways.

• Journal of Physiology & Pathology in Korean Medicine
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• v.35 no.4
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• pp.117-123
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• 2021

Gefitinib, a first generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI), provides obvious clinical benefit in patients with EGFR-mutant non-small cell lung cancer (NSCLC). However, patients ultimately develop gefitinib resistance which mainly caused by EGFR T790M secondary mutation. In the current study, we investigated whether the root extract of Scutellaria baicalensis (SB) overcomes gefitinib resistance. Gefitinib-resistant H1975 human NSCLC cells (EGFR L858R/T790M double mutant) were treated with gefitinib and/or ethanol extract of SB (ESB) to evaluate the effect of ESB on the gefitinib sensitivity. The cell viability was measured by MTT assay and trypan blue exclusion assay. The colony-forming ability was evaluated by anchorage-dependent colony formation assay. Combined treatment with gefitinib and ESB markedly decreased the cell viability and colony formation than single treatment with gefitinib or ESB in H1975 cells. In addition, cells treated with both gefitinib and ESB exhibited a significant increase of sub-G1 DNA content which indicates apoptotic cells compared with those treated with gefitinib or ESB alone. As a molecular mechanism, combined treatment with gefitinib and ESB strongly downregulated the phosphorylation of ERK and JNK than single treatment with gefitinib or ESB. Taken together, our results demonstrate that ESB sensitizes H1975 cells to gefitinib treatment. We cautiously propose that ESB can be used in combination with gefitinib for the advanced NSCLC patients with acquired resistance to EGFR TKIs.

• Biomolecules & Therapeutics
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• v.31 no.2
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• pp.200-209
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• 2023

Patients with non-small-cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR) amplification or sensitive mutations initially respond to the tyrosine kinase inhibitor gefitinib, however, the treatment becomes less effective over time by resistance mechanism including mesenchymal-epithelial transition (MET) overexpression. A therapeutic strategy targeting MET and EGFR may be a means to overcoming resistance to gefitinib. In the present study, we found that picropodophyllotoxin (PPT), derived from the roots of Podophyllum hexandrum, inhibited both EGFR and MET in NSCLC cells. The antitumor efficacy of PPT in gefitinib-resistant NSCLC cells (HCC827GR), was confirmed by suppression of cell proliferation and anchorage-independent colony growth. In the targeting of EGFR and MET, PPT bound with EGFR and MET, ex vivo, and blocked both kinases activity. The binding sites between PPT and EGFR or MET in the computational docking model were predicted at Gly772/Met769 and Arg1086/Tyr1230 of each ATP-binding pocket, respectively. PPT treatment of HCC827GR cells increased the number of annexin V-positive and subG1 cells. PPT also caused G2/M cell-cycle arrest together with related protein regulation. The inhibition of EGFR and MET by PPT treatment led to decreases in the phosphorylation of the downstream-proteins, AKT and ERK. In addition, PPT induced reactive oxygen species (ROS) production and GRP78, CHOP, DR5, and DR4 expression, mitochondrial dysfunction, and regulated involving signal-proteins. Taken together, PPT alleviated gefitinib-resistant NSCLC cell growth and induced apoptosis by reducing EGFR and MET activity. Therefore, our results suggest that PPT can be a promising therapeutic agent for gefitinib-resistant NSCLC.

• Journal of Physiology & Pathology in Korean Medicine
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• v.21 no.4
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• pp.973-981
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• 2007

In the present study, we investigated the antiproliferative activity of the water extract of Samgibopae-tang (SGBPT) in NCI-H460 and A549 non-small-cell lung cancer cell lines. We found that exposure of A549 cells to SGBPT resulted in the growth inhibition in a dose-dependent manner as measured by MTT assay, however SGBPT did not affect the growth of NCI-H460 cells. The antiproliferative effect by SGBPT treatment in A549 cells was associated with morphological changes such as membrane shrinking and cell rounding up. SGBPT treatment did not induce the cell cycle arrest in both cell lines, however the frequency of sub-G1 population was concentration-dependently increased by SGBPT treatment in A549 cells. SGBPT treatment partially induced the expression of tumor suppressor p53 in A549 cells and the expression of cyclin-dependent kinase inhibitor p21(WAF1/CIP1) was markedly increased in both transcriptional and translational levels in A549 cells. The up-regulation of p21 by SGBPT occurred in a similar a concentration dependent manner to that observed with the inhibition of cell viability and induction of sub-G1 population of the cell cycle. However SGBPT treatment did not affect other growth regulation-related genes such as early growth response-1 (Egr-1), nonsteroidal anti-inflammatory drug-activated gene-1 (NAG-1), inducible nitric oxide synthease (iNOS), cyclooxygenases (COXs), telomere-regulatory factors in A549 as well as NCI-H460 cells. Taken together, these findings suggested that SGBPT-induced inhibition of human lung carcinoma A549 cell growth was aoosciated with the induction of p21 and the results provided important new insights into the possible molecular mechanisms of the anti-cancer activity of SGBPT.

• Proceedings of the Korean Society of Toxicology Conference
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• 2002.11b
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• pp.196-196
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• 2002

p16INK4a tumor suppressor gene transfer in the non-small cell lung cancer cells by transduction of recombinant adenovirus (Ad5CMV-p16) resulted in significant inhibition of cancer cell growth (Anticancer Res., 1998, 18:3257-3261). As a safety concern, we have investigated gene and protein expression after transduction of adenoviral vector (Ad5CMV-p16) in human non-small cell lung cancer (A549) cells by using microarray and 2D gel electrophoresis/ MALDI-TOF.(omitted)

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.4
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• pp.2325-2331
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• 2013

Objective: To detect effects of plumbagin on proliferation and apoptosis in non-small cell lung cancer cell lines, and investigate the underlying mechanisms. Materials and Methods: Human non-small cell lung cancer cell lines A549, H292 and H460 were treated with various concentrations of plumbagin. Cell proliferation rates was determined using both cell counting kit-8 (CCK-8) and clonogenic assays. Apoptosis was detected by annexin V/propidium iodide double-labeled flow cytometry and TUNEL assay. The levels of reactive oxygen species (ROS) were detected by flow cytometry. Activity of NF-${\kappa}B$ was examined by electrophoretic mobility shift assay (EMSA) and luciferase reporter assay. Western blotting was used to assess the expression of both NF-${\kappa}B$ regulated apoptotic-related gene and activation of p65 and $I{\kappa}B{\kappa}$. Results: Plumbagin dose-dependently inhibited proliferation of the lung cancer cells. The IC50 values of plumbagin in A549, H292, and H460 cells were 10.3 ${\mu}mol/L$, 7.3 ${\mu}mol/L$, and 6.1 ${\mu}mol/L$ for 12 hours, respectively. The compound concentration-dependently induced apoptosis of the three cell lines. Treatment with plumbagin increased the intracellular level of ROS, and inhibited the activation of NK-${\kappa}B$. In addition to inhibition of NF-${\kappa}B$/p65 nuclear translocation, the compound also suppressed the degradation of $I{\kappa}B{\kappa}$. ROS scavenger NAC highly reversed the effect of plumbagin on apoptosis and inactivation of NK-${\kappa}B$ in H460 cell line. Treatment with plumbagin also increased the activity of caspase-9 and caspase-3, downregulated the expression of Bcl-2, upregulated the expression of Bax, Bak, and CytC. Conclusions: Plumbagin inhibits cell growth and induces apoptosis in human lung cancer cells through an NF-${\kappa}B$-regulated mitochondrial-mediated pathway, involving activation of ROS.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.7
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• pp.3061-3065
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• 2015

Background: Rac3, a member of the Rac family of small guanosine triphosphatases (GTPases), regulates a variety of cell functions, including the organization of the cytoskeleton, cell migration, and invasion. Overexpression of Rac3 has been reported in several human cancers. However, the role of Rac3 in lung cancer (LC) has not been determined in detail. The purpose of this study was to investigate the effect of silencing of Rac3 expression in human LC cells and the consequences for cell survival. Materials and Methods: Lentivirus small hairpin RNA (shRNA) interference techniques were utilized to knock down the Rac3 gene. Gene and protein expression was quantified by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. LC cell apoptosis was examined by annexin V-APC /propidium iodide staining. Results: Efficient silencing of Rac3 strongly inhibited A549 cell proliferation and colony formation ability, and significantly decreased tumor growth. Moreover, flow cytometry analysis showed that knockdown of Rac3 led to G2/M phase cell cycle arrest as well as an excess accumulation of cells in the G1 and S phase. Conclusions: Thus, functional analysis using shRNAs revealed a critical role for Rac3 in the tumor growth of LC cells. shRNA silencing of Rac3 could provide an effective strategy to treat LC.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.1
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• pp.139-143
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• 2014

Although correlations between platelets and lung cancer has been recognized, effects on non-small cell lung cancer (NSCLC) metastasis remain to be determined in detail. In the present study, wound healing assays revealed a role of platelets in NSCLC cell migration. Thus the mean migration rate of lung adenocarcinoma A549 cells was significantly elevated after co-culture with platelets ($81.7{\pm}0.45%$ vs $41.0{\pm}3.50%$, P<0.01). Expression of GAPDH was examined by reverse transcription-polymerase chain reaction to study the effect of platelets on NSCLC cell proliferation. The result showed that the proliferation of A549 and SPC-A1 cells was not affected. Mouse models were established by transfusing A549 cells and SPC-A1 cells into mice lateral tail veins. We found tumor metastasis nodules in lungs to be increased significantly after co-transfusion with platelets (in A549, $4.33{\pm}0.33$ vs $0.33{\pm}0.33$, P=0.01; in SPC-A1, $2.67{\pm}0.33$ vs $0.00{\pm}0.00$, P=0.01). In addition, consecutive inoperable patients with newly diagnosed NSCLC (TNM stage III or IV) between January 2009 and December 2011 were retrospectively reviewed. Using the Kaplan-Meier method, NSCLC patients with a high platelet counts demonstrated a significantly shorter progression free survival compared with those with a low platelet count (> $200{\times}10^9/L$, 3 months versus ${\leq}200{\times}10^9/L$, 5 months, P=0.001). An elevated platelet count was also identified as an independent prognostic factor by Cox regression analysis for prgression free survival (adjusted hazard ratio: 1.69; 95% CI: 1.16, 2.46; P=0.006). This study suggested that platelets might contribute to the hematogenous metastatic process by promoting cancer cell migration, which eventually affects the prognosis of NSCLC.

• Journal of Life Science
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• v.15 no.5 s.72
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• pp.726-733
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• 2005

Histone deacetylase (HDAC) inhibitors target key steps of tumor development. They inhibit proliferation, induce differentiation and/or apoptotic cell death, and exhibit potent antimetastatic and antiangiogenic properties in cancer cells in vitro and in vivo. Although they are emerging as a promising new treatment strategy in malignancy, how they exert their effect on human non-small cell lung cancer cells is as yet unclear. The present study was undertaken to investiate the underlying mechanism of a HDAC inhibitor trichostatin A (TSA)-induced growth arrest and its effect on the cell cycle control gene products in a human lung carcinoma cell line A549. TSA treaoent induced the growth inhibition and morphological changes in a concentration-dependent manner. Treatment of A549 cells with TSA resulted in a concentration-dependent increased G1 (under 100 ng/ml) and/or G2/M (200 ng/ml) cell population of the cell cycle as determined by flow cytometry Moreover, 200 ng/ml TSA treatment significantly induced the population of sub-G1 cells (23.0 fold of control). This anti-proliferative effect of TSA was accompanied by a marked inhibition of cyclins, positive regulators of cell cycle progression, and cyclin-dependent kinases (Cdks) expression and concomitant induction of tumor suppressor p53 and Cdk inhibitors such as p21 and p27 Although further studies are needed, these findings provide important insights into the possible molecular mechanisms of the anti-cancer activity of TSA in human lung carcinoma cells.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.8
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• pp.3695-3700
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• 2012

Mammalian mediator (MED) is a multi-protein coactivator that has been identified by several research goups. The involvement of the MED complex subunit 19 (MED 19) in the metastasis of lung adenocarcinoma cell line (H1299), which expresses the MED 19 subunit, was here investigated. When MED 19 expression was decreased by RNA interference H1299 cells demonstrated reduced clone formation, arrest in the S phase of the cell cycle, and lowered metastatic capacity. Thus, MED 19 appears to play important roles in the biological behavior of non-small cell lung carcinoma cells. These findings may be important for the development of novel lung carcinoma treatments.