Gut microbiota composition and metabolite profiling in smokers: a comparative study between emphysema and asymptomatic individuals with therapeutic implications.

BACKGROUND: Diet has a crucial role in the gut microbiota, and dysbiosis in the gut and lungs has been suggested to be associated with chronic obstructive pulmonary disease. We compared the diet, microbiome and metabolome between asymptomatic smokers and those with emphysema. METHODS: We enrolled 10 asymptomatic smokers with preserved lung function and 16 smokers with emphysema with severe airflow limitation. Dietary intake information was gathered by a self-reported questionnaire. Sputum and faecal samples were collected for microbial and metabolomics analysis. A murine model of emphysema was used to determine the effect of metabolite supplementation. RESULTS: Despite having a similar smoking history with emphysema patients, asymptomatic smokers had higher values of body mass index, fibre intake and faecal acetate level. Linear discriminant analysis identified 17 microbial taxonomic members that were relatively enriched in the faeces of asymptomatic smokers. Analysis of similarity results showed dissimilarity between the two groups (r=0.287, p=0.003). Higher acetate level was positively associated with forced expiratory volume in one second in the emphysema group (r=0.628, p=0.012). Asymptomatic smokers had a greater number of species associated with acetate and propionate (r>0.6) than did those with emphysema (30 vs 19). In an emphysema mouse model, supplementation of acetate and propionate reduced alveolar destruction and the production of proinflammatory cytokines, and propionate decreased the CD3+CD4+IL-17+ T-cell population in the lung and spleen. CONCLUSION: Smokers with emphysema showed differences in diet, microbiome and short-chain fatty acids compared with asymptomatic smokers. Acetate and propionate showed therapeutic effects in a smoking-induced murine model of emphysema.

GLUT3 Promotes Epithelial-Mesenchymal Transition via TGF-ß/JNK/ATF2 Signaling Pathway in Colorectal Cancer Cells.

Glucose transporter (GLUT) 3, a member of the GLUTs family, is involved in cellular glucose utilization and the first step in glycolysis. GLUT3 is highly expressed in colorectal cancer (CRC) and it leads to poor prognosis to CRC patient outcome. However, the molecular mechanisms of GLUT3 on the epithelial-mesenchymal transition (EMT) process in metastatic CRC is not yet clear. Here, we identified that activation of the c-Jun N-terminal kinase (JNK)/activating transcription factor-2 (ATF2) signaling pathway by transforming growth factor-ß (TGF-ß) promotes GLUT3-induced EMT in CRC cells. The regulation of GLUT3 expression was significantly associated with EMT-related markers such as E-cadherin, α- smooth muscle actin (α-SMA), plasminogen activator inhibitor-1 (PAI-1), vimentin and zinc finger E-box binding homeobox 1 (ZEB1). We also found that GLUT3 accelerated the invasive ability of CRC cells. Mechanistically, TGF-ß induced the expression of GLUT3 through the phosphorylation of JNK/ATF2, one of the SMAD-independent pathways. TGF-ß induced the expression of GLUT3 by increasing the phosphorylation of JNK, the nuclear translocation of the ATF2 transcription factor, and the binding of ATF2 to the promoter region of GLUT3, which increased EMT in CRC cells. Collectively, our results provide a new comprehensive mechanism that GLUT3 promotes EMT process through the TGF-ß/JNK/ATF2 signaling pathway, which could be a potential target for the treatment of metastatic CRC.

Xanthohumol Interferes with the Activation of TGF-ß Signaling in the Process Leading to Intestinal Fibrosis.

Fibrosis has various biological processes and affects almost every organ, especially in patients with inflammatory bowel disease, including Crohn's disease, who experience discomfort caused by intestinal fibrosis, which is a problem that needs to be resolved. TGF-ß signaling is known to act as a key regulator of intestinal fibrosis, and its modulation could be an excellent candidate for fibrosis therapy. Xanthohumol (XN) has various effects, including anti-inflammation and anti-cancer; however, the detailed mechanism of TGF-ß signaling has not yet been studied. The purpose of this study was to investigate the mechanism underlying the anti-fibrotic effect of XN on TGF-ß1-induced intestinal fibrosis using primary human intestinal fibroblasts (HIFs). In this study, to check the anti-fibrotic effects of XN on intestinal fibrosis, we assessed the expression of fibrosis-related genes in TGF-ß1-stimulated HIFs by qPCR, immunoblotting, and immunofluorescence staining. As a result, XN showed the ability to reduce the expression of fibrosis-associated genes increased by TGF-ß1 treatment in HIFs and restored the cell shape altered by TGF-ß1. In particular, XN repressed both NF-κB- and Smad-binding regions in the α-SMA promoter, which is important in fibrosis. In addition, XN inhibited NF-κB signaling, including phosphorylated-IkBα and cyclooxygenase-2 expression, and TNF-α-stimulated transcriptional activity of NF-κB. XN attenuated TGF-ß1-induced phosphorylation of Smad2 and Smad3, and the transcriptional activity of CAGA. Particularly, XN interfered with the binding of TGF-Receptor I (TßRI) and Smad3 by binding to the kinase domain of the L45 loop of TßRI, thereby confirming that the fibrosis mechanism did not proceed further. In conclusion, XN has an inhibitory effect on TGF-ß1-induced intestinal fibrosis in HIFs, significantly affecting TGF-ß/Smad signaling.

High-fiber diets attenuate emphysema development via modulation of gut microbiota and metabolism.

Dietary fiber functions as a prebiotic to determine the gut microbe composition. The gut microbiota influences the metabolic functions and immune responses in human health. The gut microbiota and metabolites produced by various dietary components not only modulate immunity but also impact various organs. Although recent findings have suggested that microbial dysbiosis is associated with several respiratory diseases, including asthma, cystic fibrosis, and allergy, the role of microbiota and metabolites produced by dietary nutrients with respect to pulmonary disease remains unclear. Therefore, we explored whether the gut microbiota and metabolites produced by dietary fiber components could influence a cigarette smoking (CS)-exposed emphysema model. In this study, it was demonstrated that a high-fiber diet including non-fermentable cellulose and fermentable pectin attenuated the pathological changes associated with emphysema progression and the inflammatory response in CS-exposed emphysema mice. Moreover, we observed that different types of dietary fiber could modulate the diversity of gut microbiota and differentially impacted anabolism including the generation of short-chain fatty acids, bile acids, and sphingolipids. Overall, the results of this study indicate that high-fiber diets play a beneficial role in the gut microbiota-metabolite modulation and substantially affect CS-exposed emphysema mice. Furthermore, this study suggests the therapeutic potential of gut microbiota and metabolites from a high-fiber diet in emphysema via local and systemic inflammation inhibition, which may be useful in the development of a new COPD treatment plan.

Cyanidin Chloride Induces Apoptosis by Inhibiting NF-κB Signaling through Activation of Nrf2 in Colorectal Cancer Cells.

Colorectal cancer (CRC) is the third most common cancer worldwide and a leading cause of cancer-related deaths in developed countries. Anthocyanins are a class of flavonoids, widely distributed in food, exhibiting important biological effects. Cyanidin chloride (CyCl) is the common type of anthocyanin with antioxidative and anti-inflammatory potential. The present study aimed to investigate the molecular mechanisms underlying the chemotherapeutic effects of CyCl in colorectal cancer cells. We found that CyCl treatment induced apoptosis as well as a significant inhibition of cellular proliferation and colony formation in three colon cancer HCT116, HT29, and SW620 cells. In addition, CyCl suppressed nuclear factor-kappa B (NF-κB) signaling and induced the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in tumor necrosis factor-alpha (TNF-α)-stimulated colon cancer cells. Nrf2 and NF-κB are two key transcription factors regulating antioxidative responses and cellular proliferation, respectively. In this study, knockdown of Nrf2 by small interfering RNA (siRNA) transfection inhibited the effect of CyCl on NF-κB signaling and apoptosis, suggesting that there is functional crosstalk between Nrf2 and NF-κB. Our findings demonstrate the important role of Nrf2 in inducing apoptosis through the involvement of NF-κB signaling in colorectal cancer cells, suggesting that CyCl may be used as a potential therapeutic agent for CRC.

Hepatoprotective Effects of Steamed and Freeze-Dried Mature Silkworm Larval Powder against Ethanol-Induced Fatty Liver Disease in Rats.

Silkworm, Bombyx mori, contains high amounts of beneficial nutrients, including amino acids, proteins, essential minerals, and omega-3 fatty acids. We have previously reported a technique for producing steamed and freeze-dried mature silkworm larval powder (SMSP), which makes it easier to digest mature silkworm. In this study, we investigated the preventive effects of SMSP on alcoholic fatty liver disease and elucidated its mechanism of action. Male Sprague-Dawley rats treated with SMSP (50 mg/kg) or normal diet (AIN-76A) were administered 25% ethanol (3 g/kg body weight) by oral gavage for 4 weeks. SMSP administration for 4 weeks significantly decreased hepatic fat accumulation in ethanol-treated rats by modulating lipogenesis and fatty acid oxidation-related molecules such as sirtuin 1, AMP-activated protein kinase, and acetyl-CoA carboxylase 1. Moreover, SMSP administration significantly diminished the levels of triglyceride in liver tissues by as much as 35%, as well as lowering the serum levels of triglyceride, gamma glutamyl transpeptidase, alanine transaminase, and aspartate aminotransferase in ethanol-treated rats. SMSP supplementation also decreased the pro-inflammatory tumor necrosis factor-alpha and interleukin 1 beta levels and cytochrome P450 2E1 generating oxidative stress. These results suggest that SMSP administration may be possible for the prevention of alcoholic liver disease.

Administration of Steamed and Freeze-Dried Mature Silkworm Larval Powder Prevents Hepatic Fibrosis and Hepatocellular Carcinogenesis by Blocking TGF-ß/STAT3 Signaling Cascades in Rats.

Hepatocellular carcinoma (HCC) is the leading cause of cancer-related deaths worldwide and the majority of HCC patients occur with a background of hepatic fibrosis and cirrhosis. We have previously reported the hepatoprotective effects of steamed and freeze-dried mature silkworm larval powder (SMSP) in a chronic ethanol-treated rat model. Here, we assessed the anti-fibrotic and anti-carcinogenic effects of SMSP on diethylnitrosamine (DEN)-treated rats. Wistar rats were intraperitoneally injected with DEN once a week for 12 or 16 weeks with or without SMSP administration (0.1 and 1 g/kg). SMSP administration significantly attenuated tumor foci formation and proliferation in the livers of the rats treated with DEN for 16 weeks. SMSP administration also inhibited hepatic fibrosis by decreasing the levels of collagen fiber and the expression of pro-collagen I and alpha-smooth muscle actin (α-SMA). Moreover, SMSP supplementation improved the major parameters of fibrosis such as transforming growth factor-ß (TGF-ß), connective tissue growth factor (CTGF), tumor necrosis factor-alpha (TNF-α), plasminogen activator inhibitor-1 (PAI-1), and collagen type I (Col1A1) in the livers from the rats treated with DEN for 16 weeks. As s possible mechanisms, we investigated the effects of SMSP on the TGF-ß and signal transducer and activator of transcription 3 (STAT3)-mediated signaling cascades, which are known to promote hepatic fibrosis. We found that SMSP treatment inhibited the activation of TGF-ß and the phosphorylation of STAT3 pathway in DEN-treated rats. Moreover, SMSP administration suppressed the expressions of the target genes of TGF-ß and STAT3 induced by DEN treatment. Our findings provide experimental evidences that SMSP administration has inhibitory effects of hepatic fibrosis and HCC induced by DEN in vivo and could be a promising strategy for the prevention or treatment of hepatic fibrosis and hepatocellular carcinogenesis.

Ampelopsin Induces DR5-Mediated Apoptotic Cell Death in EBV-Infected Cells through the p38 Pathway.

Ampelopsin (AMP) is a well-known flavonoid that exerts a number of biological and pharmacological effects including anticancer effects against several cancer cell lines. In this study, we investigated the anticancer activity of AMP against Epstein-Barr virus (EBV)-positive cells and its mechanism of action. Our results showed that AMP dose-dependently inhibited cell viability and induced apoptotic cell death in EBV-positive cells without cytotoxicity in EBV-negative cells. In particular, AMP induced caspase-8 dependent apoptosis via upregulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and death receptor (DR5). Knockdown of DR5 by RNA interference blocked AMP-induced apoptosis. Furthermore, AMP dose-dependently activated p38 mitogen-activated protein kinases (MAPKs) in EBV-positive cells. Additionally, SB203580 (a p38-MAPK inhibitor) effectively inhibited apoptotic cell death. These results demonstrate that treatment with AMP induces the apoptosis of EBV-positive cells through upregulation of TRAIL/DR5 and activation of p38 signaling. Therefore, these results provide experimental information for developing AMP as a new therapeutic drug against EBV-positive cancer.

LMP1 and 2A Induce the Expression of Nrf2 Through Akt Signaling Pathway in Epstein-Barr Virus-Transformed B Cells.

The transcription factor Nrf2, which regulates the expression of antioxidant and cytoprotective enzymes, contributes to cell proliferation and resistance to chemotherapy. Nrf2 is also dysregulated in many cancers such as lung, head and neck, and breast cancers, but its role in Epstein-Barr virus (EBV)-transformed B cells is still not understood. Here, we investigated EBV infection-induced Nrf2 activation in B cells by analyzing translocation of Nrf2 from the cytosol to the nucleus. In addition, we confirmed expression of the target genes in response to increased Nrf2 activation in EBV-transformed B cells. We demonstrated that knockdown of LMP1 and 2A blocks the translocation of Nrf2 to the nucleus and reduces ROS production in EBV-transformed B cells. Further, we showed that inhibition of Akt prevents Nrf2 activation. Moreover, knockdown of Nrf2 induces apoptotic cell death in EBV-transformed B cells. In conclusion, our study demonstrates that Nrf2 promotes proliferation of EBV-transformed B cells through the EBV-related proteins LMP1 and 2A and Akt signaling, implicating Nrf2 as a potential molecular target for EBV-associated disease.

Active Hexose Correlated Compound (AHCC) Inhibits the Proliferation of Ovarian Cancer Cells by Suppressing Signal Transducer and Activator of Transcription 3 (STAT3) Activation.

The purpose of this study was to investigate the antiproliferative effect of active hexose correlated compound (AHCC), derived from basidiomycete mushroom culture, on ovarian cancer cell lines. An in vitro growth inhibition assay was performed using AHCC in ovarian cancer cell lines. Western blotting was performed to investigate the mechanism of the observed antiproliferative effect of AHCC. We identified that ovarian cancer cell viability was significantly reduced through treatment with AHCC compared to that in the control. AHCC inhibited constitutive signal transducer and activator of transcription 3 (STAT3) phosphorylation in ovarian cancer cell lines. In contrast, treatment with pervanadate, a protein tyrosine phosphatase inhibitor, reversed AHCC-induced STAT3 suppression. AHCC treatment induced the expression of SHP-1, a protein tyrosine phosphatase, and suppressed the expression of cyclin D1, Bcl-2, Mcl-1, survivin, and VEGF, which are STAT3-regulated gene products that are associated with cell proliferation or apoptosis. These results suggest that AHCC has an antiproliferative effect on ovarian cancer cell lines, via STAT3 phosphorylation; thus, this compound has the potential to be a complementary and alternative anticancer therapy for the treatment of ovarian cancer.

Ginsenoside Rh2 induces apoptosis and inhibits epithelial-mesenchymal transition in HEC1A and Ishikawa endometrial cancer cells.

BACKGROUND: Anticancer effect of ginsenoside Rh2 has been found in various cancer cells. However, the anticancer effect of Rh2 in endometrial cancer cells is still unclear. We aimed to determine the anticancer effect of Rh2 and elucidate its mechanism in endometrial cancer cells, using HEC1A and Ishikawa cell lines, in this study. METHODS: Cell proliferation was assessed by MTT assay, and cell apoptosis was visualized by TdT mediated-dUTP Nick-End Labeling (TUNEL) method. Western blot were performed to detect the expression of apoptosis and epithelial-mesenchymal transition (EMT)-related proteins. Further, cell invasion and migration assays were conducted to estimate cell migration and invasion abilities. RESULTS: Rh2 treatment significantly suppressed cell proliferation in HEC1A and Ishikawa cells, in dose-dependent manner. Levels of cleaved poly adenosine diphosphate-ribose polymerase (PARP) and cleaved caspase-3 increased in the both cell lines with Rh2 compared with control. In Western blotting analysis after Rh2 treatment, the expression of E-cadherin increased, while the expression of EMT-related proteins including vimentin, TGF-ß, and Snail markedly decreased in both cell lines. The cell invasion and migration assays results indicated that Rh2 inhibited the cell invasion and migration in HEC1A cells. CONCLUSIONS: Our findings suggested that Rh2 exerts the anticancer effect in endometrial cancer cells through the apoptosis induction and EMT inhibition.

The microbiome of the lung and its extracellular vesicles in nonsmokers, healthy smokers and COPD patients.

Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory disease, and bacterial infection plays a role in its pathogenesis. Bacteria secrete nanometer-sized extracellular vesicles (EVs), which may induce more immune dysfunction and inflammation than the bacteria themselves. We hypothesized that the microbiome of lung EVs might have distinct characteristics depending on the presence of COPD and smoking status. We analyzed and compared the microbiomes of 13 nonsmokers with normal spirometry, 13 smokers with normal spirometry (healthy smokers) and 13 patients with COPD by using 16S ribosomal RNA gene sequencing of surgical lung tissue and lung EVs. Subjects were matched for age and sex in all groups and for smoking levels in the COPD and healthy smoker groups. Each group included 12 men and 1 woman with the same mean age of 65.5 years. In all groups, EVs consistently showed more operational taxonomic units (OTUs) than lung tissue. In the healthy smoker and COPD groups, EVs had a higher Shannon index and a lower Simpson index than lung tissue and this trend was more prominent in the COPD group. Principal component analysis (PCA) showed clusters based on sample type rather than participants' clinical characteristics. Stenotrophomonas, Propionibacterium and Alicyclobacillus were the most commonly found genera. Firmicutes were highly present in the EVs of the COPD group compared with other samples or groups. Our analysis of the lung microbiome revealed that the bacterial communities present in the EVs and in the COPD group possessed distinct characteristics with differences in the OTUs, diversity indexes and PCA clustering.

Melatonin enhances arsenic trioxide-induced cell death via sustained upregulation of Redd1 expression in breast cancer cells.

Melatonin is implicated in various physiological functions, including anticancer activity. However, the mechanism(s) of its anticancer activity is not well understood. In the present study, we investigated the combined effects of melatonin and arsenic trioxide (ATO) on cell death in human breast cancer cells. Melatonin enhanced the ATO-induced apoptotic cell death via changes in the protein levels of Survivin, Bcl-2, and Bax, thus affecting cytochrome c release from the mitochondria to the cytosol. Interestingly, we found that the cell death induced by co-treatment with melatonin and ATO was mediated by sustained upregulation of Redd1, which was associated with increased production of reactive oxygen species (ROS). Combined treatment with melatonin and ATO induced the phosphorylation of JNK and p38 MAP kinase downstream from Redd1 expression. Rapamycin and S6K1 siRNA enhanced, while activation of mTORC1 by transfection with TSC2 siRNA suppressed the cell death induced by melatonin and ATO treatment. Taken together, our findings suggest that melatonin enhances ATO-induced apoptotic cell death via sustained upregulation of Redd1 expression and inhibition of mTORC1 upstream of the activation of the p38/JNK pathways in human breast cancer cells.

Inhibition of S6K1 enhances dichloroacetate-induced cell death.

PURPOSE: The unique metabolic profile of cancer (aerobic glycolysis) is an attractive therapeutic target for cancer. Dichloroacetate (DCA), an inhibitor of pyruvate dehydrogenase kinase, has been shown to reverse glycolytic phenotype and induce mitochondrion-dependent apoptosis. In the present study, we investigated the effects of S6 kinase 1 (S6K1) inhibition on DCA-induced cell death and the underlying mechanisms in breast cancer cells. METHODS: Cell death was evaluated by annexin V and PI staining. The synergistic effects of DCA and PF4708671 were assessed by isobologram analysis. Small interfering RNA (siRNA) was used for suppressing gene expression. The mRNA and protein levels were measured by RT-PCR and Western blot analysis, respectively. RESULTS: PF4708671, a selective inhibitor of S6K1, and knockdown of S6K1 with specific siRNA enhanced DCA-induced cell death. Interestingly, a combination of DCA/PF4708671 markedly reduced protein expression of a glycolytic enzyme, hexokinase 2 (HK2). Suppression of HK2 activity using specific siRNA and 2-deoxyglucose (2-DG) further enhanced cell sensitivity to DCA/PF4708671. Overexpression of Myc-tagged HK2 rescued cell death induced by DCA/PF4708671. CONCLUSIONS: Based on these findings, we propose that inhibition of S6K1, in combination with the glycolytic inhibitor, DCA, provides effective cancer therapy.

HS-133, a novel fluorescent phosphatidylinositol 3-kinase inhibitor as a potential imaging and anticancer agent for targeted therapy.

As PI3K/Akt signaling is frequently deregulated in a wide variety of human tumors, PI3K inhibitors are an emerging class of drugs for cancer treatment. The monitoring of the drug behavior and distribution in the biological system can play an important role for targeted therapy and provide information regarding the response or resistance to available therapies. In this study, therefore, we have developed a family of xanthine derivatives, serving as a dual function exhibiting fluorescence, as well as inhibiting PI3K. Among them, HS-133 showed anti-proliferative effects and was monitored for its subcellular localization by a fluorescence microscopy. HS-133 suppressed the PI3K/Akt pathway and induced cell cycle arrest at the G0/G1 phase. The induction of apoptosis by HS-133 was confirmed by the increases of the cleaved PARP, caspase-3, and caspase-8. Furthermore, HS-133 decreased the protein expression of HIF-1α and VEGF, as well inhibited the tube formation and migration of the human umbilical vein endothelial cells. In vivo imaging also showed that tumors were visualized fluorescent with HS-133, and its oral administration significantly inhibited the growth of tumor in SkBr3 mouse xenograft models. Thus, we suggest that HS-133 may be used as a fluorescent anticancer agent against human breast cancer.

HS-438, a new inhibitor of imatinib-resistant BCR-ABL T315I mutation in chronic myeloid leukemia.

Imatinib is a selective breakpoint cluster region-Abelson (BCR-ABL) tyrosine kinase inhibitor (TKI) that has significantly improved the prognosis of patients with chronic myeloid leukemia (CML). However, T315I gene mutations of the BCR-ABL kinase domain have been shown to confer resistance to imatinib. In the present study, we synthesized a novel BCR-ABL inhibitor, HS-438, and identified its anti-leukemic effects in vitro and in vivo. We found that HS-438 strongly inhibited the expression of BCR-ABL signaling pathways in wild-type BCR-ABL (BaF3/WT) cells as well as T315I-mutated BCR-ABL (BaF3/T315I) cells with resistance to imatinib. HS-438 induced cell cycle arrest, particularly during the G0/G1 cell cycle phase, and induced apoptosis. In BaF3/T315I xenograft models, HS-438 significantly delayed tumor growth, unlike imatinib. In summary, we suggest that HS-438 may be a novel drug candidate with the therapeutic potential to target BCR-ABL and overcome imatinib resistance in patients with CML.

Tanshinone IIA induces autophagic cell death via activation of AMPK and ERK and inhibition of mTOR and p70 S6K in KBM-5 leukemia cells.

Although tanshinone IIA (Tan IIA) from Salviae miltiorrhizae was known to induce apoptosis in various cancers, its underlying mechanism of autophagic cell death was not reported yet. Thus, in the present study, the molecular mechanism of autophagic cell death by Tan IIA was investigated in KBM-5 leukemia cells. Tan IIA significantly increased the expression of microtubule-associated protein light chain 3 (LC3) II as a hallmark of autophagy in western blotting and immunofluorescence staining. Tan IIA augmented the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) and attenuated the phosphorylation of mammalian target of rapamycin (mTOR) and p70 S6K in a dose-dependent manner. Conversely, autophagy inhibitor 3-methyladenine partly reversed the cytotoxicity and the phosphorylation of AMPK, mTOR and p70 S6K induced by Tan IIA in KBM-5 leukemia cells. In addition, Tan IIA dramatically activated the extracellular signal regulated kinase (ERK) signaling pathway including Raf, ERK and p90 RSK in a dose-dependent and time-dependent manner. Consistently, ERK inhibitor PD184352 suppressed LC3-II activation induced by Tan IIA, whereas PD184352 and PD98059 did not affect poly (ADP-ribose) polymerase cleavage and sub-G1 accumulation induced by Tan IIA in KBM-5 leukemia cells. Furthermore, Tan IIA could induce autophagy via LC3-II activation in various cancer cells such as prostate (PC-3), multiple myeloma (U266), lung (NCI-H460), and breast (MDA-MB-231) cells. Overall, these findings suggest that Tan IIA induces autophagic cell death via activation of AMPK and ERK and inhibition of mTOR and p70 S6K in KBM-5 cells as a potent natural compound for leukemia treatment.

Induction of apoptosis and suppression of angiogenesis of hepatocellular carcinoma by HS-159, a novel phosphatidylinositol 3-kinase inhibitor.

The phosphatidylinositol 3-kinase (PI3K) pathway plays a central role in cell proliferation and survival in human cancer and is emerging as an attractive therapeutic target. In this study, we synthesized a novel PI3Kα inhibitor, HS-159 [N-(5-(3-(3-cyanophenyl)imidazo[1,2-a]pyridin-6-yl)pyridin-3-yl)benzenesulfonamide] and evaluated its anticancer effects on Huh-7 human hepatocellular carcinoma (HCC) cells. HS-159 effectively inhibited the phosphorylation of downstream PI3K effectors such as Akt, mTOR and P70S6 kinases in a dose-dependent manner. This compound also induced apoptosis and increased the fraction of apoptotic cells in the sub-G1 phase as well as the levels of cleaved PARP, caspase-3 and -9. Furthermore, HS-159 decreased the expression of hypoxia inducible factor-1α and vascular endothelial growth factor which play important roles in angiogenesis. The anti-angiogenic effect of HS-159 was confirmed by the suppression of tube formation and migration of human umbilical vein endothelial cells in vitro. Collectively, our results demonstrate that HS-159 exhibited anticancer activities including the induction of apoptosis and inhibition of angiogenesis by blocking the PI3K/Akt pathway in Huh-7 cells. Therefore, we suggest that this drug may be potentially used for targeted HCC therapy.

Discovery of new benzothiazole-based inhibitors of breakpoint cluster region-Abelson kinase including the T315I mutant.

The existence of drug resistance caused by mutations in the break-point cluster region-Abelson tyrosine kinase (Bcr-Abl) kinase domain remains a clinical challenge due to limited effective treatment options for chronic myeloid leukemia (CML). Herein we report a novel series of benzothiazole-based inhibitors that are effective against wild-type and T315I mutant Bcr-Abl kinases. The original hit compound, nocodazole, was extensively modified through a structure-based drug design strategy, especially by varying the groups at the C2 and C6 positions of the scaffold. In addition, the introduction of water-solubilizing groups at the terminal ethyl group resulted in enhanced physicochemical properties and potency in cellular inhibition. Several compounds inhibited the kinase activity of both wild-type Bcr-Abl and the T315I mutant with IC50 values in the picomolar range and exhibited good antiproliferative effects on Ba/F3 cell lines transformed with either wild-type or T315I mutant Bcr-Abl.

Activation of c-Jun N-terminal kinase mediates tanshinone IIA-induced apoptosis in KBM-5 chronic myeloid leukemia cells.

Aim of this study was to identify the molecular mechanisms of tanshinone IIA-induced apoptosis in chronic myelogenous leukemia (CML) cells. Cytotoxicity of tanshinone IIA was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Our data demonstrate that tanshinone IIA induced apoptosis by increasing the sub-G1 DNA contents and DNA fragmentation in KBM-5 CML cell line. In addition, tanshinone IIA significantly reduced mitochondrial membrane potential (MMP), mediated cytochrome c release from mitochondria and activated caspase-3 and 9, indicating mitochondria-dependent apoptosis by tanshinone IIA. Tanshinone IIA attenuated expression of several apoptosis-related proteins such as c-inhibitor of apoptosis protein (IAP) 2, Mcl-1(L) and Bcl-2. Interestingly, although tanshinone IIA notably enhanced the phosphorylation of both c-Jun N-terminal protein kinase (JNK) and p38, JNK inhibitor, but not p38 inhibitor, reversed tanshinone IIA-induced apoptosis. Our findings suggest that tanshinone IIA induces mitochondria-dependent apoptosis via activation of JNK in KBM 5 cells as a potent anti-cancer agent for CML therapy.