An Animal Model That Mimics Human Herpesvirus 6B Pathogenesis.

Human herpesvirus 6B (HHV-6B), a T-lymphotropic virus, infects almost exclusively humans. An animal model of HHV-6B has not been available. Here, we report the first animal model to mimic HHV-6B pathogenesis; the model is based on humanized mice in which human immune cells were engrafted and maintained. For HHV-6B replication, adequate human T-cell activation (which becomes susceptible to HHV-6B) is necessary in this murine model. Here, we found that an additional transfer of human mononuclear cells to humanized mice resulted in an explosive proliferation of human activated T cells, which could be representative of graft-versus-host disease (GVHD) because the primary transfer of human cells was not sufficient to increase the number and ratio of human T cells. Mice infected with HHV-6B became weak and/or died approximately 7 to 14 days later. Quantitative PCR analysis revealed that the spleen and lungs were the major sites of HHV-6B replication in this model, and this was corroborated by the detection of viral proteins in these organs. Histological analysis also revealed the presence of megakaryocytes, indicating HHV-6B infection. Multiplex analysis of cytokines/chemokines in sera from the infected mice showed secretions of human cytokines/chemokines as reported for both in vitro infection and clinical samples, indicating that the secreted cytokines could affect pathogenesis. This is the first animal model showing HHV-6B pathogenesis, and it will be useful for elucidating the pathogenicity of HHV-6B, which is related to GVHD and idiopathic pneumonia syndrome.IMPORTANCE Human herpesvirus 6B (HHV-6B) is a ubiquitous virus that establishes lifelong latent infection only in humans, and the infection can reactivate, with severe complications that cause major problems. A small-animal model of HHV-6B infection has thus been desired for research regarding the pathogenicity of HHV-6B and the development of antiviral agents. We generated humanized mice by transplantation with human hematopoietic stem cells, and here, we modified the model by providing an additional transfer of human mononuclear cells, providing the proper conditions for efficient HHV-6B infection. This is the first humanized mouse model to mimic HHV-6B pathogenesis, and it has great potential for research into the in vivo pathogenesis of HHV-6B.

Role of tyrosine kinase-independent phosphorylation of EGFR with activating mutation in cisplatin-treated lung cancer cells.

Epidermal growth factor receptor (EGFR) mutation is one of the hallmarks of cancer progression and resistance to anticancer therapies, particularly non-small cell lung carcinomas (NSCLCs). In contrast to the canonical EGFR activation in which tyrosine residues are engaged, we have demonstrated that the non-canonical pathway is triggered by phosphorylation of serine and threonine residues through p38 and ERK MAPKs, respectively. The purpose of this study is to investigate the role of non-canonical EGFR pathway in resistance mechanism against cisplatin treatment. Wild type and mutated (exon 19 deletion) EGFR-expressing cells responded similarly to cisplatin by showing MAPK-mediated EGFR phosphorylation. It is interesting that internalization mechanism of EGFR was switched from tyrosine kinase-dependent to p38-dependent fashions, which is involved in a survival pathway that counteracts cisplatin treatment. We therefore introduce a potential combinatorial therapy composed of p38 inhibition and cisplatin to block the activation of EGFR, therefore inducing cancer cell death and apoptosis.

Helicobacter pylori Induces Serine Phosphorylation of EGFR via Novel TAK1-p38 Activation Pathway in an HB-EGF-Independent Manner.

BACKGROUND: The interaction of Helicobacter pylori with gastric epithelial cells can result in the activation of transcription factor NF-κB via TGF-ß-activated kinase 1 (TAK1). In this study, we have demonstrated the role of H. pylori in the activation of EGFR via TAK1-mediated phosphorylation of p38. MATERIALS AND METHODS: Gastric epithelial AGS or MKN-45 cells were co-cultured with wild-type or cagA(-) H. pylori strains. H. pylori was added to the cells, and the activation of EGFR, p65 (NF-κB) subunit, p38, ERK, and TAK1 was examined by Western blotting. Infected cells were pretreated with or without ligands, chemical inhibitors, anti-HB-EGF antibody, and siRNAs to evaluate the effects on phosphorylation of various EGFR residues. Fluorescence microscopy and flow cytometry were performed to detect the internalization of EGFR. RESULTS: Incubating cells with wild-type and CagA(-) H. pylori strains resulted in the rapid and transient phosphorylation of serine residues of EGFR. RNAi experiments using siRNA against TAK1 and p38 pathways blocked the phosphorylation of serine residue. Immunofluorescence and flow cytometry revealed that EGFR was internalized in H. pylori-infected cells after EGFR phosphorylation in a p38-dependent manner. In contrast, pretreatment with gefitinib and anti-HB-EGF antibody did not block both the phosphorylation and internalization of EGFR. CONCLUSION: Helicobacter pylori induces internalization of EGFR via novel TAK1-p38-serine activation pathway which is independent of HB-EGF. The interaction between TAK1 and EGFR in H. pylori-infected cells might open new dimensions in understanding H. pylori-associated gastric carcinogenesis.

Anti-inflammatory effect of cinnamaldehyde in Helicobacter pylori induced gastric inflammation.

Cinnamomum cassia is widely employed for gastrointestinal complaints such as dyspepsia, flatulence, diarrhea, and vomiting. Studies report cinnamaldehyde (CM) as a major active constituent of cinnamon. The aim of this study was to evaluate the anti-inflammatory mechanism of CM on Helicobacter (H.) pylori-infected gastric epithelial cells in order to validate cinnamon traditional use in gastrointestinal (GI)-related disorders. AGS/MKN-45 cells and H. pylori (193C) were employed for co-culture experiments. Anti-H. pylori cytotoxic and anti-adhesion activity of CM were determined. Enzyme linked immunosorbent assay, real time polymerase chain reaction analysis and immunoblotting were used to measure the effect on interleukin-8 (IL-8) secretion/expression. The effect on activation of nuclear factor kappa B (NF-κB) was determined by immunoblot analysis. The non-cytotoxic CM (≤125 µM) was also non-bactericidal at the given time, suggesting the effect in H. pylori/cell co-culture system was not due to alteration in H. pylori viability or the toxicity to the cells. Also, CM did not show any anti-adhesion effect against H. pylori/cell co-culture. However, pre-incubation of the cells with CM significantly inhibited the IL-8 secretion/expression from H. pylori-infected cells (p<0.01). In addition, CM suppressed H. pylori-induced NF-κB activation and prevented degradation of inhibitor (I)-κB This study provides evidence that the anti-inflammatory effect of C. cassia on H. pylori-infected gastric cells is due to blockage of the NF-κB pathway by cinnamaldehyde. This agent can be considered as a potential candidate for in vivo and clinical studies against various H. pylori related gastric pathogenic processes.

Preclinical evaluation of the efficacy of an antibody to human SIRPα for cancer immunotherapy in humanized mouse models.

Tumor-associated macrophages (TAMs) are abundant in the tumor microenvironment and are considered potential targets for cancer immunotherapy. To examine the antitumor effects of agents targeting human TAMs in vivo, we here established preclinical tumor xenograft models based on immunodeficient mice that express multiple human cytokines and have been reconstituted with a human immune system by transplantation of human CD34+ hematopoietic stem and progenitor cells (HIS-MITRG mice). HIS-MITRG mice supported the growth of both human cell line (Raji)- and patient-derived B cell lymphoma as well as the infiltration of human macrophages into their tumors. We examined the potential antitumor action of an antibody to human SIRPα (SE12C3) that inhibits the interaction of CD47 on tumor cells with SIRPα on human macrophages and thereby promotes Fcγ receptor-mediated phagocytosis of the former cells by the latter. Treatment with the combination of rituximab (antibody to human CD20) and SE12C3 inhibited Raji tumor growth in HIS-MITRG mice to a markedly greater extent than did rituximab monotherapy. This enhanced antitumor effect was dependent on human macrophages and attributable to enhanced rituximab-dependent phagocytosis of lymphoma cells by human macrophages. Treatment with rituximab and SE12C3 also induced reprogramming of human TAMs toward a proinflammatory phenotype. Furthermore, the combination treatment essentially prevented the growth of patient-derived diffuse large B cell lymphoma in HIS-MITRG mice. Our findings thus support the study of HIS-MITRG mice as a model for the preclinical evaluation in vivo of potential therapeutics, such as antibodies to human SIRPα, that target human TAMs.

Distinct roles of transforming growth factor-beta-activated kinase 1 (TAK1)-c-Rel and interferon regulatory factor 4 (IRF4) pathways in human T cell lymphotropic virus 1-transformed T helper 17 cells producing interleukin-9.

Investigation of helper T cell markers in HTLV-1-transformed cell lines demonstrated that HuT-102 has an IL-9-producing Th17 phenotype. We confirmed the vital role of retinoic acid-related orphan receptor C, a Th17 transcription factor, in the expression of IL-17. Interferon regulatory factor 4 (IRF4), a transcription factor overexpressed in all HTLV-1-infected cells, regulated IL-17 and IL-9 concomitantly. We further demonstrated a novel pathway for the regulation of Tax-induced cytokines, IL-9 and IL-6, through TAK1-mediated nuclear accumulation of c-Rel. A microarray analysis for IRF4 knocked down HuT-102 cells showed a significant up-regulation in the set of genes related to Th1, mainly IFN-γ and several transcription factors. T-bet and IRF1, but not STAT1 and IRF9, participated in counteracting the inhibitory effect of IRF4 on the production of IFN-γ. Finally, suppression of both IRF4 and c-Rel resulted in the reduced proliferation. Collectively, these findings indicate that TAK1-c-Rel and IRF4 pathways play distinct roles in the maintenance of IL-9-producing Th17 phenotype of HTLV-1-transformed cells.

Human T cell lymphotropic virus 1 manipulates interferon regulatory signals by controlling the TAK1-IRF3 and IRF4 pathways.

We previously reported that human T cell lymphotropic virus 1 (HTLV-1) Tax oncoprotein constitutively activates transforming growth factor-beta-activated kinase 1 (TAK1). Here, we established Tax-positive HuT-102 cells stably transfected with a short hairpin RNA vector (HuT-shTAK1 cells) and investigated the physiological function of TAK1. Microarray analysis demonstrated that several interferon (IFN)-inducible genes, including chemokines such as CXCL10 and CCL5, were significantly down-regulated in HuT-shTAK1 cells. In contrast, Tax-mediated constitutive activation of nuclear factor-kappaB (NF-kappaB) was intact in HuT-shTAK1 cells. IFN-regulatory factor 3 (IRF3), a critical transcription factor in innate immunity to viral infection, was constitutively activated in a Tax-dependent manner. Activation of IRF3 and IRF3-dependent gene expressions was dependent on TAK1 and TANK-binding kinase 1 (TBK1). On the other hand, IRF4, another member in the IRF family of transcription factors overexpressed in a Tax-independent manner, negatively regulated TAK1-dependent IRF3 transcriptional activity. Together, HTLV-1 manipulates IFN signaling by regulating both positive and negative IRFs.

Immune Checkpoint Regulators: A New Era Toward Promising Cancer Therapy.

During the last century, our battle against cancer has been inaugurated upon three main approaches; surgery, radiation and chemotherapy. The latest findings on the effectiveness of immunotherapy in cancer management offer a ray of hope after decades of research and studies on the best treatment methods. Immunotherapy has proven effective in the surveillance and destruction of cancer- causing cells, demonstrating its ability to suppress cancer through controlling the wellestablished immune-editing process. Immuno-editing is a process that comprises three principal elements; elimination, equilibrium, and escape, and is paramount to the comprehension of checkpoint inhibition. Cancer cells employ various approaches to evade the elimination step leading to its immune- escape. The escape mechanism encompasses the up-regulation of negative co-signals that block successful activation of cancer-eradicating immune cells, developing cytokine background that favors the immunosuppressive tumor microenvironment (TME), or dropping the expression of tumor- specific proteins known as neo-antigens, therefore reducing the immunogenic activity against cancer cells. Today, checkpoint inhibitors are considered as a primary approach in our fight against cancer. Strategies targeting the inhibitory roles of checkpoint inhibitors have been shown effective against different cancer types and stages, and some already gained the FDA's approval. This review seeks to comprehensively cover the historical background as well as the most recent updates for the role of immune checkpoint regulators in the maintenance of immune homeostatic balance as well as keeping the tumorigenic cells in check.

Modulation of activation-induced cytidine deaminase by curcumin in Helicobacter pylori-infected gastric epithelial cells.

BACKGROUND: Anomalous expression of activation-induced cytidine deaminase (AID) in Helicobacter pylori-infected gastric epithelial cells has been postulated as one of the key mechanisms in the development of gastric cancer. AID is overexpressed in the cells through nuclear factor (NF)-kappaB activation by H. pylori and hence, inhibition of NF-kappaB pathway can downregulate the expression of AID. Curcumin, a spice-derived polyphenol, is known for its anti-inflammatory activity via NF-kappaB inhibition. Therefore, it was hypothesized that curcumin might suppress AID overexpression via NF-kappaB inhibitory activity in H. pylori-infected gastric epithelial cells. MATERIALS AND METHODS: MKN-28 or MKN-45 cells and H. pylori strain 193C isolated from gastric cancer patient were used for co-culture experiments. Cells were pretreated with or without nonbactericidal concentrations of curcumin. Apoptosis was determined by DNA fragmentation assay. Enzyme-linked immunosorbent assay was performed to evaluate the anti-adhesion activity of curcumin. Real-time polymerase chain reaction was employed to evaluate the expression of AID mRNA. Immunoblot assay was performed for the analysis of AID, NF-kappaB, inhibitors of NF-kappaB (IkappaB), and IkappaB kinase (IKK) complex regulation with or without curcumin. RESULTS: The adhesion of H. pylori to gastric epithelial cells was not inhibited by curcumin pretreatment at nonbactericidal concentrations (< or =10 micromol/L). Pretreatment with nonbactericidal concentration of curcumin downregulated the expression of AID induced by H. pylori. Similarly, NF-kappaB activation inhibitor (SN-50) and proteasome inhibitor (MG-132) also downregulated the mRNA expression of AID. Moreover, curcumin (< or =10 micromol/L) has suppressed H. pylori-induced NF-kappaB activation via inhibition of IKK activation and IkappaB degradation. CONCLUSION: Nonbactericidal concentrations of curcumin downregulated H. pylori-induced AID expression in gastric epithelial cells, probably via the inhibition of NF-kappaB pathway. Hence, curcumin can be considered as a potential chemopreventive candidate against H. pylori-related gastric carcinogenesis.

Role of mitochondria in rescuing glycolytically inhibited subpopulation of triple negative but not hormone-responsive breast cancer cells.

Triple-negative breast cancer (TNBC) subtype is among the most aggressive cancers with the worst prognosis and least therapeutic targetability while being more likely to spread and recur. Cancer transformations profoundly alter cellular metabolism by increasing glucose consumption via glycolysis to support tumorigenesis. Here we confirm that relative to ER-positive cells (MCF7), TNBC cells (MBA-MD-231) rely more on glycolysis thus providing a rationale to target these cells with glycolytic inhibitors. Indeed, iodoacetate (IA), an effective GAPDH inhibitor, caused about 70% drop in MDA-MB-231 cell viability at 20 µM while 40 µM IA was needed to decrease MCF7 cell viability only by 30% within 4 hours of treatment. However, the triple negative cells showed strong ability to recover after 24 h whereas MCF7 cells were completely eliminated at concentrations <10 µM. To understand the mechanism of MDA-MB-231 cell survival, we studied metabolic modulations associated with acute and extended treatment with IA. The resilient TNBC cell population showed a significantly greater count of cells with active mitochondria, lower apoptotic markers, normal cell cycle regulations, moderately lowered ROS, but increased mRNA levels of p27 and PARP1; all compatible with enhanced cell survival. Our results highlight an interplay between PARP and mitochondrial oxidative phosphorylation in TNBC that comes into play in response to glycolytic disruption. In the light of these findings, we suggest that combined treatment with PARP and mitochondrial inhibitors may provide novel therapeutic strategy against TNBC.

Prospects for repurposing CNS drugs for cancer treatment.

Drug repurposing is the idea of using an already approved drug for another disease or disorder away from its initial use. This new approach ensures the reduction in high cost required for developing a new drug in addition to the time consumed, especially in the tumor disorders that show an unceasing rising rate with an unmet success rate of new anticancer drugs. In our review, we will review the anti-cancer effect of some CNS drugs, including both therapeutic and preventive, by searching the literature for preclinical or clinical evidence for anticancer potential of central nervous system drugs over the last 8 years period (2010-2018) and including only evidence from Q1 journals as indicated by Scimago website (www.scimagojr.com). We concluded that Some Central Nervous system drugs show a great potential as anti-cancer in vitro, in vivo and clinical trials through different mechanisms and pathways in different types of cancer that reveal a promising evidence for the repurposing of CNS drugs for new indications.

Mechanisms for DNA-damaging agent-induced inactivation of ErbB2 and ErbB3 via the ERK and p38 signaling pathways.

Cisplatin (CDDP) and doxorubicin (DOX) are chemotherapeutic drugs that trigger apoptosis by inducing DNA-damage. A previous study using breast cancer cells demonstrated the negative feedback modulation of the epidermal growth factor receptor (EGFR) and receptor tyrosine-protein kinase erbB-2 (ErbB2) via extracellular signal-regulated kinase (ERK)-mediated phosphorylation of conserved Thr-669 and Thr-677 residues, respectively, in the juxtamembrane domain. In addition, CDDP has been identified to cause negative feedback inhibition of activated EGFR in lung cancer cells. In the present study, the role of phosphorylation in the feedback control of the ErbB2/ErbB3 heterodimer in human breast and gastric cancer cells was investigated. Phosphorylation of ErbB2 at Thr-677 was induced by CDDP and DOX, which in turn reduced tyrosine autophosphorylation of ErbB2 and ErbB3. Treatment with trametinib, a mitogen-activated protein kinase inhibitor that blocks ERK-mediated Thr-677 phosphorylation, and substitution of Thr-677 to alanine, blocked the feedback inhibition of ErbB2 and ErbB3. In addition, these agents caused the degradation of ErbB proteins through the activation of p38 mitogen-activated protein kinase (p38) and ERK. These results demonstrate that chemotherapeutic agents trigger ERK- and p38-mediated post-translational downregulation of ErbB receptors.

Retrospective screening of microarray data to identify candidate IFN-inducible genes in a HTLV-1 transformed model.

HuT-102 cells are considered one of the most representable human T-lymphotropic virus 1 (HTLV-1)-infected cell lines for studying adult T-cell lymphoma (ATL). In our previous studies, genome-wide screening was performed using the GeneChip system with Human Genome Array U133 Plus 2.0 for transforming growth factor-ß-activated kinase 1 (TAK1)-, interferon regulatory factor 3 (IRF3)- and IRF4-regulated genes to demonstrate the effects of interferon-inducible genes in HuT-102 cells. Our previous findings demonstrated that TAK1 induced interferon inducible genes via an IRF3-dependent pathway and that IRF4 has a counteracting effect. As our previous data was performed by manual selection of common interferon-related genes mentioned in the literature, there has been some obscure genes that have not been considered. In an attempt to maximize the outcome of those microarrays, the present study reanalyzed the data collected in previous studies through a set of computational rules implemented using 'R' software, to identify important candidate genes that have been missed in the previous two studies. The final list obtained consisted of ten genes that are highly recommend as potential candidate for therapies targeting the HTLV-1 infected cancer cells. Those genes are ATM, CFTR, MUC4, PARP14, QK1, UBR2, CLEC7A (Dectin-1), L3MBTL, SEC24D and TMEM140. Notably, PARP14 has gained increased attention as a promising target in cancer cells.

The Unfolded Protein Response: A Novel Therapeutic Target for Poor Prognostic BRAF Mutant Colorectal Cancer.

BRAFV600E mutations occur in ∼10% of colorectal cancer cases, are associated with poor survival, and have limited responses to BRAF/MEK inhibition with or without EGFR inhibition. There is an unmet need to understand the biology of poor prognostic BRAFMT colorectal cancer. We have used differential gene expression and pathway analyses of untreated stage II and stage III BRAFMT (discovery set: n = 31; validation set: n = 26) colorectal cancer, and an siRNA screen to characterize the biology underpinning the BRAFMT subgroup with poorest outcome. These analyses identified the unfolded protein response (UPR) as a novel and druggable pathway associated with the BRAFMT colorectal cancer subgroup with poorest outcome. We also found that oncogenic BRAF drives endoplasmic reticulum (ER) stress and UPR pathway activation through MEK/ERK. Furthermore, inhibition of GRP78, the master regulator of the UPR, using siRNA or small molecule inhibition, resulted in acute ER stress and apoptosis, in particular in BRAFMT colorectal cancer cells. In addition, dual targeting of protein degradation using combined Carfilzomib (proteasome inhibitor) and ACY-1215 (HDAC6-selective inhibitor) treatment resulted in marked accumulation of protein aggregates, acute ER stress, apoptosis, and therapeutic efficacy in BRAFMT in vitro and xenograft models. Mechanistically, we found that the apoptosis following combined Carfilzomib/ACY-1215 treatment is mediated through increased CHOP expression. Taken together, our findings indicate that oncogenic BRAF induces chronic ER stress and that inducers of acute ER stress could be a novel treatment strategy for poor prognostic BRAFMT colorectal cancer. Mol Cancer Ther; 17(6); 1280-90. ©2018 AACR.

Hyperthermia and radiation reduce the toxic side-effects of bufadienolides for cancer therapy.

Bufadienolides are constituents of the traditional Chinese medicine Chan Su and are found in toad venom. Cardiovascular side-effects are one of the limiting factors towards developing bufadienolides as chemotherapeutic agents. Thus, in the present study, low doses of bufalin and cinobufotalin, prominent members of the bufadienolides, were investigated for their cytotoxic activity in combination with hyperthermia (HT) or radiation (Rad) therapy. In addition, the underlying mechanism involved was investigated. A DNA fragmentation assay, viability assay and microscopic observation were primarily used to assess the effect of low doses of the two drugs in human lymphoma U937 cells. Furthermore, the effects of these drugs on the mitochondrial membrane potential (MMP) and apoptotic-associated protein activation were investigated. HT/bufadienolide- and RT/bufadienolide-treated samples significantly increased the DNA fragmentation percentile and decreased the MMP, as well as increasing the apoptotic features observed microscopically within a relatively short time (6 h) after treatment. The two combinations affected the expression of important apoptotic markers, including caspase-3 and BH3 interacting domain death agonist. The findings of the current study confirm the additive effect of HT with this group of drugs, directing a novel therapeutic avenue for the clinical use of bufadienolides at lower doses with more restrained cardio toxic side-effects.

Rapid versus gradual bladder decompression in acute urinary retention.

OBJECTIVE: To demonstrate a benefit in diminished adverse events such as hypotension and hematuria with gradual drainage of the bladder when compared to rapid decompression in patients with acute urinary retention (AUR) due to benign prostatic hyperplasia in a case-control study. METHODS: Sixty-two patients matched our selection criteria presenting with AUR. They were divided into two groups - the first was managed by rapid drainage of the bladder, the second was managed by gradual drainage through a urethral catheter (The first 100 mL immediately evacuated, then the rest evacuated gradually over 2 h). RESULTS: The mean age was 64.4 and 63.2 years in the first and second group, respectively. Diagnosed cause was benign hyperplasia of the prostate. Hematuria occurred in two patients in the first group and none in the second group. The two cases of hematuria were mild and treated conservatively. After the relief of the obstruction, the mean blood pressure was noticed to decrease by 15 mmHg and 10 mmHg in the first and second group, respectively, however, no one developed significant hypotension. Pain relief was achieved after complete drainage in the first group and after the evacuation of 100 mL in the second group. CONCLUSIONS: We conclude that there is no significant difference between rapid and gradual decompression of the bladder in patients with AUR. Hematuria and hypotension may occur after rapid decompression of the obstructed urinary bladder, but these complications are rarely clinically significant.

Bardoxolone-methyl inhibits migration and metabolism in MCF7 cells.

Bardoxolone-methyl (BAR) is reported to have anti-inflammatory, anti-proliferative and anti-fibrotic effects. BAR activates Nrf2 and may ameliorate oxidative stress through induction of antioxidant genes. However, off-target effects, probably concentration and NFkB-dependent, have limited the clinical use of BAR. Nrf2 regulates expression of antioxidant and mitochondrial genes and has been proposed as a target for both obesity and breast cancer. Therefore, we explored whether BAR can alter migration and proliferation in the MCF7 cell line and whether metabolic function is affected by BAR. Incubation with BAR caused a time-dependent migratory inhibition and an associated decrease in mitochondrial respiration. Both migratory and mitochondrial inhibition by BAR were further enhanced in the presence of fatty acids. In addition to the activation of Nrf2, BAR altered the expression of target mRNA GCLC and UCP1. After 24 h, BAR inhibited both glycolytic capacity, reserve (p < 0.05) and oxidative phosphorylation (p < 0.001) with an associated increase in mitochondrial ROS and loss of intracellular glutathione in MCF7 cells; however, impairment of mitochondrial activity was prevented by N-acetyl cysteine. The fatty acid, palmitate, increased mitochondrial ROS, impaired migration and oxidative phosphorylation but palmitate toxicity towards MCF7 could not be inhibited by N-acetyl cysteine suggesting that they exert effects through different pathways. BAR-activated AKT, induced DNA damage and inhibited cell proliferation. When the proteasome was inhibited, there was loss of BAR-mediated changes in p65 phosphorylation and SOD2 expression suggesting non-canonical NFkB signaling effects. These data suggest that BAR-induced ROS are important in inhibiting MCF7 migration and metabolism by negatively affecting glycolytic capacity and mitochondrial function.

Inhibition of p38 mitogen-activated protein kinase potentiates the apoptotic effect of berberine/tumor necrosis factor-related apoptosis-inducing ligand combination therapy.

It was previously reported that berberine (BBR) and tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) exhibited a synergistic apoptotic effect on triple negative breast cancer (TNBC) cells. In addition, the BBR/TRAIL combination treatment sensitized TRAIL-resistant TNBC cells to TRAIL. The aim of the present study was to investigate a novel pathway for enhancing the apoptotic effect of BBR/TRAIL through mitogen-activated protein kinases (MAPKs). Selective inhibitors and small interfering RNAs were utilized to understand the role of p38 MAPK in this pathway. The results demonstrated that p38 MAPK was activated in response to the combination therapy in TRAIL-resistant TNBC cells. In addition, it was revealed that the inhibition of p38 enhanced apoptosis in epidermal growth factor receptor (EGFR)-overexpressing MDA-MB-468 TNBC cells and EGFR-mutant PC-9 non-small-cell lung carcinoma cells, which was associated with the downregulation of EGFR serine phosphorylation. Viability assays for these two cell lines also confirmed the significant reduction of cell viability following p38 inhibition in BBR/TRAIL-treated cells. In conclusion, the present study provided novel evidence for the role of p38 in suppressing BBR/TRAIL-mediated apoptosis and its association with EGFR, which may explain the mechanism of treatment resistance in certain types of cancer.

TRAIL combinations: The new 'trail' for cancer therapy (Review).

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) therapy is anticipated to be one of the most effective cancer treatments. However, resistance to TRAIL therapy remains a challenge facing the development of anticancer strategies. To circumvent this problem, TRAIL combinations have been experimented with for over ten years to induce synergism or sensitize resistant cancer cells. By analyzing the signaling pathways triggered by these combinations, this review has defined a set of core targets for novel combinatorial treatments. The review suggests specific pathways to be targeted together with TRAIL for more efficient treatment, including cellular FLICE inhibitory protein and its downstream survival factors, the Bcl-2 family and other prominent targets. The suggested pathways provide new avenues for more effective TRAIL-based cancer therapy.

Piperine enhances the efficacy of TRAIL-based therapy for triple-negative breast cancer cells.

BACKGROUND/AIM: Triple-negative breast cancer (TNBC) is most the aggressive type of breast cancer and is poorly responsive to endocrine therapeutics; however, one of the most attractive treatments is tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-based therapies. To identify compounds that enhance the efficacy of TRAIL-based therapies, we screened 55 compounds from natural products in combination with TRAIL in TNBC cells. MATERIALS AND METHODS: Human TNBC cells, MDA-MB-468 and MDA-MB-231, and murine TNBC cells, 4T1, were used. Cell viability, apoptotic cells, and cell cycle were quantified by the WST-1 assay, annexin-V/7-amino-actinomycinD (7-AAD) staining and Propidium iodide (PI) staining, respectively. In vivo effects of piperine were evaluated in the orthotopic-inoculated 4T1-luc mouse model. RESULTS: After screening, we identified piperine as the most potent adjuvant at enhancing the efficacy of TRAIL-based therapies in TNBC cells in vitro and in vivo, which might be mediated through inhibition of survivin and p65 phosphorylation. CONCLUSION: Piperine may enhance TRAIL-based therapeutics for TNBC.