Ergosta-7,9(11),22-trien-3ß-ol Alleviates Intracerebral Hemorrhage-Induced Brain Injury and BV-2 Microglial Activation.

Intracerebral hemorrhage (ICH) is a devastating neurological disorder characterized by an exacerbation of neuroinflammation and neuronal injury, for which few effective therapies are available at present. Inhibition of excessive neuroglial activation has been reported to alleviate ICH-related brain injuries. In the present study, the anti-ICH activity and microglial mechanism of ergosta-7,9(11),22-trien-3ß-ol (EK100), a bioactive ingredient from Asian medicinal herb Antrodia camphorate, were evaluated. Post-treatment of EK100 significantly attenuated neurobehavioral deficit and MRI-related brain lesion in the mice model of collagenase-induced ICH. Additionally, EK100 alleviated the inducible expression of cyclooxygenase (COX)-2 and the activity of matrix metalloproteinase (MMP)-9 in the ipsilateral brain regions. Consistently, it was shown that EK100 concentration-dependently inhibited the expression of COX-2 protein in Toll-like receptor (TLR)-4 activator lipopolysaccharide (LPS)-activated microglial BV-2 and primary microglial cells. Furthermore, the production of microglial prostaglandin E2 and reactive oxygen species were attenuated by EK100. EK100 also attenuated the induction of astrocytic MMP-9 activation. Among several signaling pathways, EK100 significantly and concentration-dependently inhibited activation of c-Jun N-terminal kinase (JNK) MAPK in LPS-activated microglial BV-2 cells. Consistently, ipsilateral JNK activation was markedly inhibited by post-ICH-treated EK100 in vivo. In conclusion, EK100 exerted the inhibitory actions on microglial JNK activation, and attenuated brain COX-2 expression, MMP-9 activation, and brain injuries in the mice ICH model. Thus, EK100 may be proposed and employed as a potential therapeutic agent for ICH.

Haloperidol Abrogates Matrix Metalloproteinase-9 Expression by Inhibition of NF-κB Activation in Stimulated Human Monocytic Cells.

Much evidence has indicated that matrix metalloproteinases (MMPs) participate in the progression of neuroinflammatory disorders. The present study was undertaken to investigate the inhibitory effect and mechanism of the antipsychotic haloperidol on MMP activation in the stimulated THP-1 monocytic cells. Haloperidol exerted a strong inhibition on tumor necrosis factor- (TNF-) α-induced MMP-9 gelatinolysis of THP-1 cells. A concentration-dependent inhibitory effect of haloperidol was observed in TNF-α-induced protein and mRNA expression of MMP-9. On the other hand, haloperidol slightly affected cell viability and tissue inhibition of metalloproteinase-1 levels. It significantly inhibited the degradation of inhibitor-κB-α (IκBα) in activated cells. Moreover, it suppressed activated nuclear factor-κB (NF-κB) detected by a mobility shift assay, NF-κB reporter gene, and chromatin immunoprecipitation analyses. Consistent with NF-κB inhibition, haloperidol exerted a strong inhibition of lipopolysaccharide- (LPS-) induced MMP-9 gelatinolysis but not of transforming growth factor-ß1-induced MMP-2. In in vivo studies, administration of haloperidol significantly attenuated LPS-induced intracerebral MMP-9 activation of the brain homogenate and the in situ in C57BL/6 mice. In conclusion, the selective anti-MMP-9 activation of haloperidol could possibly involve the inhibition of the NF-κB signal pathway. Hence, it was found that haloperidol treatment may represent a bystander of anti-MMP actions for its conventional psychotherapy.

3-Aroylindoles display antitumor activity in vitro and in vivo: Effects of N1-substituents on biological activity.

A series of 3-aroylindole hydroxamic acids (10-17) were developed based on the concept of a structural combination of tubulin and histone deacetylase (HDAC) inhibitors. This was accomplished by introducing hydroxamic acid-containing moieties at the N1 position of the tubulin assembly inhibitor, compound 9 (SCB01A, BPR0L075, phase II trial). Most of synthetic compounds produced in this way displayed comparable HDAC inhibitory activity, and four (10, 12-14) of them also inhibit tubulin assembly. Notably, compound 12 possesses not only tubulin and HDAC inhibitory activity but also shows HDAC6 selectivity over other HDAC isoforms. In addition, it exhibits remarkable inhibitory activity against the growth cancer cells in vitro and in vivo (PC3 and RPMI-8226 cells). Notably, it suppresses the growth of multiple myeloma xenografts without leading to the death of teated animals like reference compound. In sum, this study provided potential compounds with safer profiles for cancer treatment.

Mycobacterium tuberculosis Upregulates TNF-α Expression via TLR2/ERK Signaling and Induces MMP-1 and MMP-9 Production in Human Pleural Mesothelial Cells.

BACKGROUND: Tumor necrosis factor (TNF)-α and matrix metalloproteinases (MMPs) are elevated in pleural fluids of tuberculous pleuritis (TBP) where pleural mesothelial cells (PMCs) conduct the first-line defense against Mycobacterium tuberculosis (MTB). However, the clinical implication of TNF-α and MMPs in TBP and the response of PMCs to MTB infection remain unclear. METHODS: We measured pleural fluid levels of TNF-α and MMPs in patients with TBP (n = 18) or heart failure (n = 18) as controls. Radiological scores for initial effusion amount and residual pleural fibrosis at 6-month follow-up were assessed. In vitro human PMC experiments were performed to assess the effect of heat-killed M. tuberculosis H37Ra (MTBRa) on the expression of TNF-α and MMPs. RESULTS: As compared with controls, the effusion levels of TNF-α, MMP-1 and MMP-9 were significantly higher and correlated positively with initial effusion amount in patients with TBP, while TNF-α and MMP-1, but not MMP-9, were positively associated with residual pleural fibrosis of TBP. Moreover, effusion levels of TNF-α had positive correlation with those of MMP-1 and MMP-9 in TBP. In cultured PMCs, MTBRa enhanced TLR2 and TLR4 expression, activated ERK signaling, and upregulated TNF-α mRNA and protein expression. Furthermore, knockdown of TLR2, but not TLR4, significantly inhibited ERK phosphorylation and TNF-α expression. Additionally, both MTBRa and TNF-α markedly induced MMP-1 and MMP-9 synthesis in human PMCs, and TNF-α neutralization substantially reduced the production of MMP-1, but not MMP-9, in response to MTBRa stimulation. CONCLUSION: MTBRa activates TLR2/ERK signalings to induce TNF-α and elicit MMP-1 and MMP-9 in human PMCs, which are associated with effusion volume and pleural fibrosis and may contribute to pathogenesis of TBP. Further investigation of manipulation of TNF-α and MMP expression in pleural mesothelium may provide new insights into the mechanisms and rational treatment strategies for TBP.

Carbamazepine attenuates inducible nitric oxide synthase expression through Akt inhibition in activated microglial cells.

UNLABELLED: Abstract Background: Carbamazepine, which was developed primarily for the treatment of epilepsy, is now also useful for the treatment of non-epileptic disorders and inflammatory hyperalgesia. However, the mechanism of its anti-neuroinflammatory action remains poorly understood. OBJECTIVE: This study elucidates the anti-neuroinflammatory capacity of carbamazepine on microglial activation and the relative mechanisms involved. MATERIALS AND METHODS: The microglial BV-2 cells were pretreated with carbamazepine for 15 min before activation by lipopolysaccharide (LPS). After LPS stimulation, the expression of inducible nitric oxide synthase (iNOS) was analyzed by Western blotting (WB) and reverse transcription-polymerase chain reaction. Signaling proteins and cyclooxygenase (COX)-2 were also evaluated by WB. The levels of nitrate and tumor necrosis factor (TNF)-α were analyzed by the Griess method and enzyme-linked immunosorbant assay, respectively. The formation of intracellular reactive oxygen species (ROS) was examined by fluorescent analysis. RESULTS: Carbamazepine strongly attenuated LPS-induced production of NO and iNOS protein at concentrations of 5, 10, and 20 µM. Consistently, it could markedly suppress iNOS mRNA expression stimulated by LPS. Among the signaling pathways, LPS-mediated IκBα degradation or JNK MAPK phosphorylation was not affected by carbamazepine. Interestingly, it was found that carbamazepine could concentration-dependently inhibit LPS-activated phospho-Akt expression. Nevertheless, LPS-induced ROS production was not affected by carbamazepine. Carbamazepine (20 µM) affected either COX-2 expression or TNF-α production induced by LPS with approximately 70% and 51% inhibition, respectively. DISCUSSION AND CONCLUSION: Our findings showed that carbamazepine exerted selective inhibition on LPS-induced microglial iNOS expression through the down-regulation of Akt activation, and thus may play a pivotal role of anti-neuroinflammation in its therapeutic efficacy.

Histone deacetylase inhibitor impairs plasminogen activator inhibitor-1 expression via inhibiting TNF-α-activated MAPK/AP-1 signaling cascade.

Tumor necrosis factor-(TNF-)-α upregulates plasminogen activator inhibitor-(PAI-) 1 expression in pleural mesothelial cells (PMCs), contributing to fibrin deposition and pleural fibrosis. Histone deacetylases (HDACs) have been found implicated in fibrogenesis. However, the roles of TNF-α or HDAC in the regulation of PAI-1 expression have not been well investigated. We aimed to examine the effects and mechanisms of HDAC inhibition on TNF-α-induced PAI-1 expression in human PMCs. MeT-5A human PMCs were treated with TNF-α in the presence or absence of the m-carboxycinnamic acid bishydroxamide (CBHA), an HDAC class II inhibitor, and the HDAC activity, PAI-1 protein expression, mRNA, and activated signalings were analyzed. CBHA abrogated TNF-α-induced HDAC activity, PAI-1 protein and, mRNA expression in MeT-5A cells. Moreover, CBHA significantly enhanced mitogen-activated protein kinase phosphatase-(MKP-) 5/MKP-1 expression and inhibited p38/JNK activations, ATF2/c-Jun translocation, and PAI-1 promoter activity. Altogether, our data suggest that HDAC inhibition may abrogate TNF-α-activated MAPK/AP-1 signaling and PAI-1 expression in human PMCs. Given the antifibrotic effect through PAI-1 abrogation, CBHA may be utilized as a novel agent in the treatment of fibrotic diseases.

Cephalochromin induces G0/G1 cell cycle arrest and apoptosis in A549 human non-small-cell lung cancer cells by inflicting mitochondrial disruption.

The fungus-derived compound cephalochromin, isolated from the fermented broth of Cosmospora vilior YMJ89051501, shows growth-inhibitory and apoptotic activity against human lung cancer A549 cells in a concentration-dependent manner with an IC50 value of 2.8 µM at 48 h. Cephalochromin induced cell cycle arrest at the G0/G1 phase through down-regulation of cyclin D1, cyclin E, Cdk 2, and Cdk 4 expressions. Cephalochromin markedly increased the hypodiploid sub-G1 phase (apoptosis) of the cell cycle at 48 h as measured by flow cytometric analysis. Reactive oxygen species generation and loss of the mitochondrial membrane potential (MMP) were also markedly induced by cephalochromin. Moreover, the immunoblotting assays showed that cephalochromin reduced survivin and Bcl-xL expression and induced the activation of caspase-8, -9, and -3 and the cleavage of poly(ADP-ribose) polymerase, indicating the involvement of a caspase signaling cascade. The caspase inhibitor Z-VAD-fmk significantly suppressed cephalochromin-induced apoptosis. Cephalochromin also triggered LC3 II, autophagic marker, expression. Taken together, this is the first report that cephalochromin induced an antiproliferative effect on human lung cancer cells through mitochondrial disruption and down-regulation of survivin, leading to cell cycle arrest at the G0/G1 phase, loss of MMP, and subsequently apoptotic cell death.

Polyketides from the littoral plant associated fungus Pseudallescheria boydii.

Four previously unreported chemical entities, boydone A (1), boydone B (2), botryorhodine F (3), and botryorhodine G (4), along with five known compounds, fusidilactone A (5), (R)-(-)-mevalonolactone (6), (R)-(-)-lactic acid (7), ovalicin (8), and botryorhodine C (9), were isolated from the ethyl acetate extracts of the fermented broths of the fungal strain Pseudallescheria boydii NTOU2362. The structures of 1-9 were characterized on the basis of their spectroscopic data analyses. The absolute configurations of 1 and 2 were established by comparison with the literature and the modified Mosher's method. The growth inhibitory activities of 1-9 against the A549 non-small-cell lung cancer cell line were evaluated, and 2 and 8 exhibited moderate to potent bioactivities with GI50 values of 41.3 and 4.1 µM, respectively, in comparison with fluorouracil (GI50 = 3.6 µM).

Butein Inhibits Angiogenesis of Human Endothelial Progenitor Cells via the Translation Dependent Signaling Pathway.

Compelling evidence indicates that bone marrow-derived endothelial progenitor cells (EPCs) can contribute to postnatal neovascularization and tumor angiogenesis. EPCs have been shown to play a "catalytic" role in metastatic progression by mediating the angiogenic switch. Understanding the pharmacological functions and molecular targets of natural products is critical for drug development. Butein, a natural chalcone derivative, has been reported to exert potent anticancer activity. However, the antiangiogenic activity of butein has not been addressed. In this study, we found that butein inhibited serum- and vascular endothelial growth factor- (VEGF-) induced cell proliferation, migration, and tube formation of human EPCs in a concentration dependent manner without cytotoxic effect. Furthermore, butein markedly abrogated VEGF-induced vessels sprouting from aortic rings and suppressed microvessel formation in the Matrigel implant assay in vivo. In addition, butein concentration-dependently repressed the phosphorylation of Akt, mTOR, and the major downstream effectors, p70S6K, 4E-BP1, and eIF4E in EPCs. Taken together, our results demonstrate for the first time that butein exhibits the antiangiogenic effect both in vitro and in vivo by targeting the translational machinery. Butein is a promising angiogenesis inhibitor with the potential for treatment of cancer and other angiogenesis-related diseases.

Trichodermin induces cell apoptosis through mitochondrial dysfunction and endoplasmic reticulum stress in human chondrosarcoma cells.

Chondrosarcoma is the second most common primary bone tumor, and it responds poorly to both chemotherapy and radiation treatment. Nalanthamala psidii was described originally as Myxosporium in 1926. This is the first study to investigate the anti-tumor activity of trichodermin (trichothec-9-en-4-ol, 12,13-epoxy-, acetate), an endophytic fungal metabolite from N. psidii against human chondrosarcoma cells. We demonstrated that trichodermin induced cell apoptosis in human chondrosarcoma cell lines (JJ012 and SW1353 cells) instead of primary chondrocytes. In addition, trichodermin triggered endoplasmic reticulum (ER) stress protein levels of IRE1, p-PERK, GRP78, and GRP94, which were characterized by changes in cytosolic calcium levels. Furthermore, trichodermin induced the upregulation of Bax and Bid, the downregulation of Bcl-2, and the dysfunction of mitochondria, which released cytochrome c and activated caspase-3 in human chondrosarcoma. In addition, animal experiments illustrated reduced tumor volume, which led to an increased number of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive cells and an increased level of cleaved PARP protein following trichodermin treatment. Together, this study demonstrates that trichodermin is a novel anti-tumor agent against human chondrosarcoma cells both in vitro and in vivo via mitochondrial dysfunction and ER stress.

Pycnidione, a fungus-derived agent, induces cell cycle arrest and apoptosis in A549 human lung cancer cells.

Pycnidione, a small tropolone first isolated from the fermented broth of Theissenia rogersii 92031201, exhibits antitumor activities through an undefined mechanism. The present study evaluated the effects and mechanisms of pycnidione on the growth and death of A549 human lung cancer cells. Pycnidione significantly inhibited the proliferation of A549 cells in a concentration-dependent manner, with a 50% growth inhibition (GI(50)) value of approximately 9.3nM at 48h. Pycnidione significantly decreased the expression of cyclins D1 and E and induced G(1)-phase cell cycle arrest and a subsequent increase in the sub-G(1) phase population. Pycnidione also markedly reduced the expression of survivin and activated caspase-8 and -3, increased reactive oxygen species (ROS) generation, caused the collapse of the mitochondrial membrane potential (MMP), and enhanced PAI-1 production, thus triggering apoptosis in the A549 cells. Taken together, pycnidione exerts anti-proliferative effects on human lung cancer cells through the induction of cell cycle arrest and apoptosis. Therefore, testing of its effects in vivo is warranted to evaluate its potential as a therapeutic agent against lung cancer.

Suppression of matrix metalloproteinase-9 expression by andrographolide in human monocytic THP-1 cells via inhibition of NF-κB activation.

There is much evidence indicating that human leukemic cells and monocytes/macrophages synthesize, and secrete, several matrix metalloproteinases (MMPs), and participate in the degradation of extracellular matrix components in tissue lesions. In this study, we investigated the effects and mechanisms of andrographolide, extracted from the herb Andrographis paniculata, on human monocytic MMPs expression and activation. Andrographolide (1-50 µM) exhibited concentration-dependent inhibition of MMP-9 activation, induced by either tumor necrosis factor-α (TNF-α), or lipopolysaccharide (LPS), in THP-1cells. In addition, andrographolide did not present an inhibitory effect on MMP-9 enzymatic activity at a concentration of 50 µM. By contrast, enzyme-linked immunosorbent assay (ELISA) showed that andrographolide partially affect TIMP-1 levels. Western blot analysis showed that both TNF-α, and LPS stimulators attenuated MMP-9 protein expression in a concentration-dependent manner. Using reverse transcription polymerase chain reaction (RT-PCR), we found that andrographolide suppressed expression of MMP-9 messenger RNA. Furthermore, we also found that andrographolide could significantly inhibit the degradation of inhibitor-κB-α (IκB-α) induced by TNF-α. We used electrophoretic mobility shift assay and reporter gene detection to show that andrographolide also markedly inhibited NF-κB signaling, anti-translocation and anti-activation. In conclusion, we demonstrate that andrographolide attenuates MMP-9 expression, and its main mechanism might involve the NF-κB signal pathway. These results provide new opportunities for the development of new anti-inflammatory and leukemic therapies.

Chemical modification and anticancer effect of prenylated flavanones from Taiwanese propolis.

Our previous studies demonstrated that eight prenylated flavanones (1-8), isolated from Taiwanese propolis, were capable of a broad spectrum of biological activities. Among them, nymphaeol A (3), nymphaeol B (4) and nymphaeol C (7), abundant in Taiwanese propolis, exhibited cytotoxicity against cancer cell lines. It therefore seemed interesting to improve their activity via a semi-synthetic strategy. In this study, 12 novel prenylated flavanones were synthesised in our laboratory and their activities were assessed for two human prostate cancer cell lines, PC-3 and DU-145, and a human hepatoma cell line, Hep-3B. Of these compounds, 10c, 11 and 12 showed more potent cytotoxicity against the PC-3 cell line than 5-Fu. Using cytometric analysis followed by double staining with annexin V-FITC and propidium iodide, it was observed that these compounds induced apoptosis as well. This suggests that prenylated flavanones 10c, 11 and 12 may have anticancer potential for further development.

Histone deacetylase inhibitor m-carboxycinnamic acid bis-hydroxamide attenuates plasminogen activator inhibitor-1 expression in human pleural mesothelial cells.

Plasminogen activator inhibitor-1 (PAI-1), primarily up-regulated by transforming growth factor (TGF)-ß, is essential in the development of fibrosis. Histone deacetylase (HDAC) was shown to modulate gene expression and fibrogenesis in various tissues. However, the implications of HDAC in terms of PAI-1 expression and pleural fibrosis remain unclear. In this study, we examined the effects of m-carboxycinnamic acid bis-hydroxamide (CBHA), a hybrid-polar HDAC inhibitor, on the TGF-ß1-induced expression of PAI-1 in a human pleural mesothelial cell line (MeT-5A). MeT-5A cells were treated with TGF-ß1 in the presence or absence of CBHA. We assayed the expression and stability of PAI-1 mRNA and protein, PAI-1 promoter activity, the activation of Smad signaling, the protein-protein interactions of Smads with transcriptional cofactors Sp1 and coactivator p300, and the expression of the mRNA-stabilizing protein nucleolin. The results indicate that CBHA significantly inhibited TGF-ß1-induced PAI-1 mRNA and protein expression, and attenuated PAI-1 promoter activity in MeT-5A cells. CBHA abrogated TGF-ß1-induced Smad4 nuclear translocation, but not Smad2/3 activation. Furthermore, the association of Smad4 with p300, but not with Sp1, was disrupted by CBHA. Alternatively, CBHA suppressed TGF-ß1-induced nucleolin expression, and thereby destabilized PAI-1 mRNA and decreased PAI-1 protein concentrations. These findings suggest that the inhibition of HDAC activity by CBHA may attenuate PAI-1 expression through the modulation of cellular signaling at multiple levels. Given the down-regulating effect of CBHA on PAI-1 expression, HDAC inhibitors should be tested further in animal models as potential therapeutic agents for pleural fibrosis.

Chemical constituents of the fermented broth of the ascomycete Theissenia cinerea 89091602.

One new betaenone, theissenoic acid (1), together with three new acetogenins, theissenolactones A-C (2-4, resp.), were isolated from the fermented broth of Theissenia cinerea 89091602 isolated in Taiwan. The structures of 1-4 were elucidated by spectroscopic methods. Biological tests revealed that 3 and 4 exhibited moderate growth-inhibitory activities against A549 lung cancer cell line with GI(50) values of 14.9 and 47.9 µM, respectively.

Aristolochic acid downregulates monocytic matrix metalloproteinase-9 by inhibiting nuclear factor-κB activation.

Aristolochic acid (AA)-associated nephropathy was described as being characterized by a rapid progressive enhancement of interstitial renal fibrosis. Renal tissue fibrosis occurs because of an imbalance of extracellular matrix (ECM) accumulation and matrix metalloproteinase (MMP) activation. Much evidence indicates that inflammatory renal disease including monocyte and mesangial interactions is linked to the development and progression of renal remodeling. In this study, we found that AA showed concentration-dependent inhibition of tumor necrosis factor (TNF)-α-induced MMP-9 activation with an IC(50) value of 6.4±0.5µM in human monocytic THP-1 cells. A similar effect was also noted with different ratios of AAs (types I and II). However, AA had no inhibitory effect on the intact enzymatic activity of MMP-9 at a concentration of 20µM. On the other hand, the level of tissue inhibitor of metalloproteinase (TIMP)-1 was not induced by AA, but it suppressed TNF-α-induced MMP-9 protein and messenger RNA expressions. AA also significantly inhibited TNF-α-induced IκBα degradation. Furthermore, an electrophoretic mobility shift assay and a reported gene study, respectively, revealed that AA inhibited TNF-α-induced NF-κB translocation and activation. In addition, compared to other NF-κB inhibitors, AA exerted significant inhibition of MMP-9 activation and monocyte chemotactic protein-1-directed invasion. From these results, we concluded that AA, a natural compound, inhibits TNF-α-induced MMP-9 in human monocytic cells possibly through the NF-κB signal pathway. These results also imply that AA may be involved in alteration of matrix homeostasis during renal fibrosis in vivo.

Hypertonicity-enhanced TNF-α release from activated human monocytic THP-1 cells requires ERK activation.

BACKGROUND: Hypertonic stress enhances tumor necrosis factor (TNF)-α expression in activated monocytes. However, the underlying mechanism is unknown. The produced TNF-α is primarily cleaved and released by TNF-α-converting enzyme (TACE), and the surface expression of TACE is down-regulated by endocytosis. As hypertonicity inhibits endocytosis, we evaluated the mechanism of hypertonicity-induced TNF-α release from activated human monocytic THP-1 cells. METHODS: THP-1 cells were stimulated with lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA) in the presence or absence of hypertonic agents (150 mM sucrose or 150-300 mM NaCl). The amount of TNF-α mRNA and protein, surface expression of TACE and activation of signaling pathways (mitogen-activated protein kinase, Akt and NF-κB) were assayed. RESULTS: Hypertonic sucrose and NaCl significantly enhanced TNF-α release from THP-1 cells upon LPS or PMA stimulation. Hypertonic sucrose and other endocytosis inhibitors increased surface expression of TACE, but their effects on TNF-α release were inconsistent. This enhancement effect by hypertonicity was not attenuated by inhibition of TACE or IκB kinase, but it was blocked by cycloheximide and a MAP/ERK kinase inhibitor. The LPS- or PMA-induced TNF-α mRNA expression was not increased; rather, it was inhibited by hypertonicity. ERK1/2 was re-activated after sucrose treatment in LPS-stimulated THP-1 cells. CONCLUSIONS: Hypertonicity-enhanced TNF-α protein synthesis from LPS- or PMA-activated THP-1 cells requires ERK activation and may proceed without TACE. GENERAL SIGNIFICANCE: A vast amount of TNF-α production was regulated by a crucial post-transcriptional manner in activated human monocytic leukemia cells, and it may possibly be contributed to the cachexia condition.

Methoxychalcone induces cell-cycle arrest and apoptosis in human hormone-resistant prostate cancer cells through PI 3-kinase-independent inhibition of mTOR pathways.

BACKGROUND: Chalcones are contained in fruits and vegetables, and have been suggested to display anticancer activities. In this study, the anticancer mechanism of WJ9708011 (a methoxychalcone derivative) was delineated in human prostate cancer cells. METHOD: Cell proliferation was examined by sulforhodamine B and clonogenic assays. Cell-cycle progression and mitochondrial membrane potential (DeltaPsi(m)) were detected by flow cytometric analysis. Expressions of protein and mRNA were detected by Western blot and RT-PCR technique, respectively. The protein synthesis was examined by [(3)H]leucine incorporation assay. The overexpression or knockdown techniques for specific target protein were also used in this study. RESULTS: WJ9708011 induced time- and concentration-dependent G1 arrest of the cell cycle and subsequent apoptosis in human prostate cancer cells. The G1-arrest effect was confirmed by down-regulated expressions of several G1-phase regulators, including cyclin D1, cyclin E, cyclin-dependent kinase (Cdk)-4, Cdk2, phospho-RB, E2F-1, and Cdc25A. The mRNA expressions of cyclin D1 and cyclin E were also inhibited through the suppression of NF-kappaB. WJ9708011 blocked the protein synthesis and inhibited mammalian target of rapamycin (mTOR) signaling pathways. The suppression of mTOR pathways were irrespective of Akt- and AMPK-activated protein kinase (AMPK), but were attributed to mitochondrial stress, in which the down-regulation of survivin protein level may play a crucial role. CONCLUSIONS: The data suggest that WJ9708011 induces transcriptional and translational suppression of cell-cycle regulators that might be through Akt- and AMPK-independent loss of DeltaPsi(m) and inhibition of mTOR signaling pathway, leading to G1 arrest of the cell cycle and subsequent apoptotic cell death.

Mana-Hox displays anticancer activity against prostate cancer cells through tubulin depolymerization and DNA damage stress.

Tubulin and deoxyribonucleic acid (DNA) are two potential targets for the development of cancer chemotherapeutic agents. Mana-Hox is a synthetic derivative of beta-carboline, a structure relevant to marine sponge component, manzamine. In this study, Mana-Hox induced an inhibition of cell proliferation in several types of human cancer cell lines, including androgen-independent prostate cancer PC-3 and DU-145, hepatocellular carcinoma Hep3B and HepG2, and colorectal cancer HT-29 cells. The p53-null PC-3 cells were used for to anticancer mechanisms. Mana-Hox stimulated an increase of ataxia telangiectasia mutated (ATM) phosphorylation on Ser-1981, indicating the induction of DNA double-strand breaks. It also displayed an inhibitory effect on tubulin polymerization using tubulin turbidity assay and immunofluorescence identification. However, it only showed a minor inhibition on the activity of Aurora kinase and histone deacetylase. Mana-Hox induced mitotic arrest of the cell cycle identified by downregulation of cyclin E, cyclin A, and cyclin-dependent kinase 2 (Cdk2) and an increase of MPM-2 expression. Next, it caused Bcl-2 phosphorylation on Ser-70, downregulation of Mcl-1 expression, and activation of caspase-3, leading to apoptotic cell death. Notably, Mana-Hox was not a P-glycoprotein (P-gp) substrate and showed equipotent activity against P-gp-rich cancer cells. We conclude that Mana-Hox induces dual effects on DNA damage and tubulin depolymerization, leading to mitotic arrest and activation of mitochondria-mediated apoptotic pathways. Data provide evidence that the anticancer strategy of dual-action targets could be a potential anticancer approach.

WRC-213, an l-methionine-conjugated mitoxantrone derivative, displays anticancer activity with reduced cardiotoxicity and drug resistance: identification of topoisomerase II inhibition and apoptotic machinery in prostate cancers.

Anthracyclines and anthracenediones are well-known cancer chemotherapeutic agents but their uses are limited with cardiotoxicity and drug resistance. Several l- and d-form amino acids were introduced into the anthraquinone skeleton and numerous derivatives were synthesized for the evaluation of anticancer activity. The screening tests showed that WRC-213, an l-methionine conjugation, was the most effective derivative to inhibit proliferative effect of human androgen-independent prostate cancer PC-3 cells (IC50=50 nM). In an extension evaluation, WRC-213 displayed a potent anti-proliferative activity in various cancer cell lines, including non-small cell lung cancer A549, androgen-independent prostate cancer DU145, colorectal cancer HT-29, breast cancer MCF-7 and hepatocellular carcinoma Hep3B and HepG2. It induced cell-cycle arrest at S and G2, but not mitotic phase, in PC-3 cells. The comet assay revealed that induction of DNA damage and inhibition of topoisomerase II were the primary insults. After the checkpoint arrest of the cell-cycle, WRC-213 induced the mitochondria-mediated intrinsic apoptotic pathway, including Mcl-1 cleavage, Bcl-2 down-regulation and activation of caspase-9/caspase-3 cascades. Survivin degradation and caspase-2 activation also contributed to WRC-213-induced apoptosis. Moreover, the assessment of cytotoxicity in H9c2 cardiomyocytes and drug resistance in NCI/ADR-RES cells demonstrated that WRC-213 showed much lower cardiotoxicity and P-glycoprotein-related resistance than those of mitoxantrone, etoposide and doxorubicin. In conclusion, it is suggested that WRC-213 is a potential topoisomerase II inhibitor with reduced cardiotoxicity and drug resistance. It inhibits topoisomerase II activity and induces chromosomal DNA strand breaks, leading to S and G2 arrest of the cell-cycle and activation of mitochondria-mediated apoptotic pathways.