G2A as a key modulator of carbonyl stress and apoptosis resistance in glucose-loaded cancer cells.

Cancer cells exhibit high glycolytic activity, metabolizing glucose as their primary energy substrate. Toxic metabolites produced during glycolysis, such as methylglyoxal, induce carbonyl stress (CS), promoting inflammation and oxidative stress. The elevated glucose metabolism in cancer cells creates this toxic environment. However, little research has focused on the molecules mediating these reactions and stresses, and their role in selecting and enriching apoptosis-resistant cells. This study investigated the impact of constitutively suppressing oxidized lipid receptor G2A (GPR132) expression on the relationship between CS and oxidative stress in glucose-loaded cancer cells. G2A has recently attracted attention as a tumor promoter. However, our study shows that G2A suppression under glucose loading significantly reduces CS and associated oxidative stress, thereby enhancing cancer cell survival. This suggests a new mechanism contrary to conventional thinking, involving the acute induction of glyoxalase 1 (Glo1). G2A may thus play a role in selecting and enriching apoptosis-resistant cell populations under high glucose conditions by regulating Glo1 expression. These findings improve our understanding of the adaptive capacity of cancer cells to glucose toxicity.

Neutralization of CX3CL1 Attenuates TGF-ß-Induced Fibroblast Differentiation Through NF-κB Activation and Mitochondrial Dysfunction in Airway Fibrosis.

BACKGROUND: Severe asthma, characterized by inflammation and airway remodeling, involves fibroblast differentiation into myofibroblasts expressing α-SMA. This process leads to the production of fibronectin and connective tissue growth factor (CTGF), driven by factors such as transforming growth factor (TGF)-ß. Furthermore, the persistent presence of myofibroblasts is associated with resistance to apoptosis and mitochondrial dysfunction. The chemokine (C-X3-C motif) ligand 1 (CX3CL1) plays a role in tissue fibrosis. However, it is currently unknown whether neutralization of CX3CL1 decreases TGF-ß-induced fibroblast differentiation and mitochondrial dysfunction in normal human lung fibroblasts (NHLFs). METHODS: CX3CL1/C-X3-C motif chemokine receptor 1 (CX3CR1), CX3CL1 was analyzed by immunofluorescence (IF) or immunohistochemical (IHC) staining of ovalbumin-challenged mice. CX3CL1 release was detected by ELISA. TGF-ß-induced CTGF, fibronectin, and α-SMA expression were evaluated in NHLFs following neutralization of CX3CL1 (TP213) treatment for the indicated times by Western blotting or IF staining. Mitochondrion function was detected by a JC-1 assay and seahorse assay. Cell apoptosis was observed by a terminal uridine nick-end labeling (TUNEL) assay. RESULTS: An increase in CX3CL1 expression was observed in lung tissues from mice with ovalbumin-induced asthma by IF staining. CX3CR1 was increased in the subepithelial layer of the airway by IHC staining. Moreover, CX3CR1 small interfering (si)RNA downregulated TGF-ß-induced CTGF and fibronectin expression in NHLFs. CX3CL1 induced CTGF and fibronectin expression in NHLFs. TGF-ß-induced CX3CL1 secretion from NHLFs. Furthermore, TP213 decreased TGF-ß-induced CTGF, fibronectin, and α-SMA expression in NHLFs. Mitochondrion-related differentially expressed genes (DEGs) were examined after CX3CL1 neutralization in TGF-ß-treated NHLFs. TP213 alleviated TGF-ß-induced mitochondrial dysfunction and apoptosis resistance in NHLFs. CX3CL1 induced p65, IκBα, and IKKα phosphorylation in a time-dependent manner. Furthermore, CX3CL1-induced fibronectin expression and JC-1 monomer were decreased by p65 siRNA. TP213 reduced TGF-ß-induced p65 and α-SMA expression in NHLFs. CONCLUSIONS: These findings suggest that neutralizing CX3CL1 attenuates lung fibroblast activation and mitochondrial dysfunction. Understanding the impacts of CX3CL1 neutralization on fibroblast mitochondrial function could contribute to the development of therapeutic strategies for managing airway remodeling in severe asthma.

Hypo-osmolarity induces apoptosis resistance via TRPV2-mediated AKT-Bcl-2 pathway.

In cirrhosis, several molecular alterations such as resistance to apoptosis could accelerate carcinogenesis. Recently, mechanotransduction has been attracting attention as one of the causes of these disturbances. In patients with cirrhosis, the serum sodium levels progressively decrease in the later stage of cirrhosis, and hyponatremia leads to serum hypo-osmolality. Since serum sodium levels in patients with cirrhosis with liver cancer are inversely related to cancer's number, size, stage, and cumulative survival, we hypothesized that hypo-osmolality-induced mechanotransduction under cirrhotic conditions might contribute to oncogenesis and/or progression of hepatocellular carcinoma (HCC). In this study, we adjusted osmosis of culture medium by changing the sodium chloride concentration and investigated the influence of hypotonic conditions on the apoptosis resistance of an HCC cell line, HepG2, using a serum-deprivation-induced apoptosis model. By culturing the cells in a serum-free medium, the levels of an antiapoptotic protein Bcl-2 were downregulated. In contrast, the hypotonic conditions caused apoptosis resistance by upregulation of Bcl-2. Next, we examined which pathway was involved in the apoptosis resistance. Hypotonic conditions enhanced AKT signaling, and constitutive activation of AKT in HepG2 cells led to upregulation of Bcl-2. Moreover, we revealed that the enhancement of AKT signaling was caused by intracellular calcium influx via a mechanosensor, TRPV2. Our findings suggested that hyponatremia-induced serum hypotonic in patients with cirrhosis promoted the progression of hepatocellular carcinoma.NEW & NOTEWORTHY Our study first revealed that hypo-osmolarity-induced mechanotransduction enhanced calcium-mediated AKT signaling via TRPV2 activation, resulting in contributing to apoptosis resistance. The finding indicates a possible view that liver cirrhosis-induced hyponatremia promotes hepatocellular carcinogenesis.

Venetoclax-Resistant T-ALL Cells Display Distinct Cancer Stem Cell Signatures and Enrichment of Cytokine Signaling.

Therapy resistance remains one of the major challenges for cancer treatment that largely limits treatment benefits and patient survival. The underlying mechanisms that lead to therapy resistance are highly complicated because of the specificity to the cancer subtype and therapy. The expression of the anti-apoptotic protein BCL2 has been shown to be deregulated in T-cell acute lymphoblastic leukemia (T-ALL), where different T-ALL cells display a differential response to the BCL2-specific inhibitor venetoclax. In this study, we observed that the expression of anti-apoptotic BCL2 family genes, such as BCL2, BCL2L1, and MCL1, is highly varied in T-ALL patients, and inhibitors targeting proteins coded by these genes display differential responses in T-ALL cell lines. Three T-ALL cell lines (ALL-SIL, MOLT-16, and LOUCY) were highly sensitive to BCL2 inhibition within a panel of cell lines tested. These cell lines displayed differential BCL2 and BCL2L1 expression. Prolonged exposure to venetoclax led to the development of resistance to it in all three sensitive cell lines. To understand how cells developed venetoclax resistance, we monitored the expression of BCL2, BCL2L1, and MCL1 over the treatment period and compared gene expression between resistant cells and parental sensitive cells. We observed a different trend of regulation in terms of BCL2 family gene expression and global gene expression profile including genes reported to be expressed in cancer stem cells. Gene set enrichment analysis (GSEA) showed enrichment of cytokine signaling in all three cell lines which was supported by the phospho-kinase array where STAT5 phosphorylation was found to be elevated in resistant cells. Collectively, our data suggest that venetoclax resistance can be mediated through the enrichment of distinct gene signatures and cytokine signaling pathways.

MLLT11 Regulates Endometrial Stroma Cell Adhesion, Proliferation and Survival in Ectopic Lesions of Women with Advanced Endometriosis.

MLLT11 is a gene implicated in cell differentiation and the development and progression of human cancers, but whose role in the pathogenesis of endometriosis is still unknown. Using quantitative RT-PCR and immunohistochemistry, we analyzed 37 women with and 33 women without endometriosis for differences in MLLT11 expression. We found that MLLT11 is reduced in the ectopic stroma cells of women with advanced stage endometriosis compared to women without endometriosis. MLLT11 knockdown in control stroma cells resulted in the downregulation of their proliferation accompanied by G1 cell arrest and an increase in the expression of p21 and p27. Furthermore, the knockdown of MLLT11 was associated with increased apoptosis resistance to camptothecin associated with changes in BCL2/BAX signaling. Finally, MLLT11 siRNA knockdown in the control primary stroma cells led to an increase in cell adhesion associated with the transcriptional activation of ACTA2 and TGFB2. We found that the cellular phenotype of MLLT11 knockdown cells resembled the phenotype of the primary endometriosis stroma cells of the lesion, where the levels of MLLT11 are significantly reduced compared to the eutopic stroma cells of women without the disease. Overall, our results indicate that MLLT11 may be a new clinically relevant player in the pathogenesis of endometriosis.

p62 functions as a signal hub in metal carcinogenesis.

A number of metals are toxic and carcinogenic to humans. Reactive oxygen species (ROS) play an important role in metal carcinogenesis. Oxidative stress acts as the converging point among various stressors with ROS being the main intracellular signal transducer. In metal-transformed cells, persistent expression of p62 and erythroid 2-related factor 2 (Nrf2) result in apoptosis resistance, angiogenesis, inflammatory microenvironment, and metabolic reprogramming, contributing to overall mechanism of metal carcinogenesis. Autophagy, a conserved intracellular process, maintains cellular homeostasis by facilitating the turnover of protein aggregates, cellular debris, and damaged organelles. In addition to being a substrate of autophagy, p62 is also a crucial molecule in a myriad of cellular functions and in molecular events, which include oxidative stress, inflammation, apoptosis, cell proliferation, metabolic reprogramming, that modulate cell survival and tumor growth. The multiple functions of p62 are appreciated by its ability to interact with several key components involved in various oncogenic pathways. This review summarizes the current knowledge and progress in studies of p62 and metal carcinogenesis with emphasis on oncogenic pathways related to oxidative stress, inflammation, apoptosis, and metabolic reprogramming.

NOX4 regulates macrophage apoptosis resistance to induce fibrotic progression.

Pulmonary fibrosis is a progressive lung disease often occurring secondary to environmental exposure. Asbestos exposure is an important environmental mediator of lung fibrosis and remains a significant cause of disease despite strict regulations to limit exposure. Lung macrophages play an integral role in the pathogenesis of fibrosis induced by asbestos (asbestosis), in part by generating reactive oxygen species (ROS) and promoting resistance to apoptosis. However, the mechanism by which macrophages acquire apoptosis resistance is not known. Here, we confirm that macrophages isolated from asbestosis subjects are resistant to apoptosis and show they are associated with enhanced mitochondrial content of NADPH oxidase 4 (NOX4), which generates mitochondrial ROS generation. Similar results were seen in chrysotile-exposed WT mice, while macrophages from Nox4-/- mice showed increased apoptosis. NOX4 regulated apoptosis resistance by activating Akt1-mediated Bcl-2-associated death phosphorylation. Demonstrating the importance of NOX4-mediated apoptosis resistance in fibrotic remodeling, mice harboring a conditional deletion of Nox4 in monocyte-derived macrophages exhibited increased apoptosis and were protected from pulmonary fibrosis. Moreover, resolution occurred when Nox4 was deleted in monocyte-derived macrophages in mice with established fibrosis. These observations suggest that NOX4 regulates apoptosis resistance in monocyte-derived macrophages and contributes to the pathogenesis of pulmonary fibrosis. Targeting NOX4-mediated apoptosis resistance in monocyte-derived macrophages may provide a novel therapeutic target to protect against the development and/or progression of pulmonary fibrosis.

Resistance of Human Liver Mesenchymal Stem Cells to FAS-Induced Cell Death.

Mesenchymal stem cells (MSCs) have a pronounced therapeutic potential in various pathological conditions. Though therapeutic effects of MSC transplantation have been studied for a long time, the underlying mechanisms are still not clear. It has been shown that transplanted MSCs are rapidly eliminated, presumably by apoptosis. As the mechanisms of MSC apoptosis are not fully understood, in the present work we analyzed MSC sensitivity to Fas-induced apoptosis using MSCs isolated from the biopsies of liver fibrosis patients (L-MSCs). The level of cell death was analyzed by flow cytometry in the propidium iodide test. The luminescent ATP assay was used to measure cellular ATP levels; and the mitochondrial membrane potential was assessed using the potential-dependent dye JC-1. We found that human L-MSCs were resistant to Fas-induced cell death over a wide range of FasL and anti-Fas mAb concentrations. At the same time, intrinsic death signal inducers CoCl2 and staurosporine caused apoptosis of L-MSCs in a dose-dependent manner. Despite the absence of Fas-induced cell death treatment of L-MSCs with low concentrations of FasL or anti-Fas mAb resulted in a cellular ATP level decrease, while high concentrations of the inducers caused a decline of the mitochondrial membrane potential. Pre-incubation of L-MSCs with the pro-inflammatory cytokine TNF-α did not promote L-MSC cell death. Our data indicate that human L-MSCs have increased resistance to receptor-mediated cell death even under inflammatory conditions.

[Programmed Necrosis Inducers for Cancer Treatment].

Programmed necrosis,a mode of cell death independent of Caspase,is mainly mediated by receptor-interacting protein kinase-1 (RIPK1),receptor-interacting protein kinase-3 (RIPK3),and mixed lineage kinase domain-like protein (MLKL).Studies have demonstrated that programmed necrosis has the dual role of promoting and inhibiting tumor growth and thus we can control the development of tumor by regulating programmed necrosis.The drugs capable of inducing programmed necrosis show potential anti-tumor activity.In addition,inducing programmed necrosis is an effective way to overcome tumor resistance to apoptosis.This paper summarized the mechanisms of programmed necrosis and its relationship with tumors.We focused on the antitumor activity of programmed necrosis inducers including natural products,chemotherapeutic drugs,death receptor ligands,kinase inhibitors,inorganic salts,metal complexes,and metal nanoparticles.These agents will provide new therapeutic candidates for the treatment of tumors,especially the tumors acquiring resistance to apoptosis.

Polygodial and Ophiobolin A Analogues for Covalent Crosslinking of Anticancer Targets.

In a search of small molecules active against apoptosis-resistant cancer cells, including glioma, melanoma, and non-small cell lung cancer, we previously prepared α,ß- and γ,δ-unsaturated ester analogues of polygodial and ophiobolin A, compounds capable of pyrrolylation of primary amines and demonstrating double-digit micromolar antiproliferative potencies in cancer cells. In the current work, we synthesized dimeric and trimeric variants of such compounds in an effort to discover compounds that could crosslink biological primary amine containing targets. We showed that such compounds retain the pyrrolylation ability and possess enhanced single-digit micromolar potencies toward apoptosis-resistant cancer cells. Target identification studies of these interesting compounds are underway.

Impaired Myofibroblast Dedifferentiation Contributes to Nonresolving Fibrosis in Aging.

Idiopathic pulmonary fibrosis (IPF) is a fatal age-associated disease with no cure. Although IPF is widely regarded as a disease of aging, the cellular mechanisms that contribute to this age-associated predilection remain elusive. In this study, we sought to evaluate the consequences of senescence on myofibroblast cell fate and fibrotic responses to lung injury in the context of aging. We demonstrated that nonsenescent lung myofibroblasts maintained the capacity for dedifferentiation, whereas senescent/IPF myofibroblasts exhibited an impaired capacity for dedifferentiation. We previously demonstrated that the transcription factor MyoD acts as a critical switch in the differentiation and dedifferentiation of myofibroblasts. Here, we demonstrate that decreased levels of MyoD preceded myofibroblast dedifferentiation and apoptosis susceptibility in nonsenescent cells, whereas MyoD expression remained elevated in senescent/IPF myofibroblasts, which failed to undergo dedifferentiation and demonstrated resistance to apoptosis. Genetic strategies to silence MyoD restored the susceptibility of IPF myofibroblasts to undergo apoptosis and led to a partial reversal of age-associated persistent fibrosis in vivo. The capacity for myofibroblast dedifferentiation and subsequent apoptosis may be critical for normal physiologic responses to tissue injury, whereas restricted dedifferentiation and apoptosis resistance in senescent cells may underlie the progressive nature of age-associated human fibrotic disorders. These studies support the concept that senescence may promote profibrotic effects via impaired myofibroblast dedifferentiation and apoptosis resistance, which contributes to myofibroblast accumulation and ultimately persistent fibrosis in aging.

Coordinated regulation of cell survival and cell cycle pathways by DDR2-dependent SRF transcription factor in cardiac fibroblasts.

Relative resistance to apoptosis and the ability to proliferate and produce a collagen-rich scar determine the critical role of cardiac fibroblasts in wound healing and tissue remodeling following myocardial injury. Identification of cardiac fibroblast-specific factors and mechanisms underlying these aspects of cardiac fibroblast function is therefore of considerable scientific and clinical interest. In the present study, gene knockdown and overexpression approaches and promoter binding assays showed that discoidin domain receptor 2 (DDR2), a mesenchymal cell-specific collagen receptor tyrosine kinase localized predominantly in fibroblasts in the heart, acts via ERK1/2 MAPK-activated serum response factor (SRF) transcription factor to enhance the expression of antiapoptotic cIAP2 in cardiac fibroblasts, conferring resistance against oxidative injury. Furthermore, DDR2 was found to act via ERK1/2 MAPK-activated SRF to transcriptionally upregulate Skp2 that in turn facilitated post-translational degradation of p27, the cyclin-dependent kinase inhibitor that causes cell cycle arrest, to promote G1-S transition, as evidenced by Rb phosphorylation, increased proliferating cell nuclear antigen (PCNA) levels, and flow cytometry. DDR2-dependent ERK1/2 MAPK activation also suppressed forkhead box O 3a (FoxO3a)-mediated transcriptional induction of p27. Inhibition of the binding of collagen type I to DDR2 using WRG-28 indicated the obligate role of collagen type I in the activation of DDR2 and its regulatory role in cell survival and cell cycle protein expression. Notably, DDR2 levels positively correlated with SRF, cIAP2, and PCNA levels in cardiac fibroblasts from spontaneously hypertensive rats. To conclude, DDR2-mediated ERK1/2 MAPK activation facilitates coordinated regulation of cell survival and cell cycle progression in cardiac fibroblasts via SRF.NEW & NOTEWORTHY Relative resistance to apoptosis and the ability to proliferate and produce a collagen-rich scar enable cardiac fibroblasts to play a central role in myocardial response to injury. This study reports novel findings that mitogen-stimulated cardiac fibroblasts exploit a common regulatory mechanism involving collagen receptor (DDR2)-dependent activation of ERK1/2 MAPK and serum response factor to achieve coordinated regulation of apoptosis resistance and cell cycle progression, which could facilitate their survival and function in the injured myocardium.

Mechanisms of Apoptosis Resistance to NK Cell-Mediated Cytotoxicity in Cancer.

Natural killer (NK) cells are major contributors to immunosurveillance and control of tumor development by inducing apoptosis of malignant cells. Among the main mechanisms involved in NK cell-mediated cytotoxicity, the death receptor pathway and the release of granules containing perforin/granzymes stand out due to their efficacy in eliminating tumor cells. However, accumulated evidence suggest a profound immune suppression in the context of tumor progression affecting effector cells, such as NK cells, leading to decreased cytotoxicity. This diminished capability, together with the development of resistance to apoptosis by cancer cells, favor the loss of immunogenicity and promote immunosuppression, thus partially inducing NK cell-mediated killing resistance. Altered expression patterns of pro- and anti-apoptotic proteins along with genetic background comprise the main mechanisms of resistance to NK cell-related apoptosis. Herein, we summarize the main effector cytotoxic mechanisms against tumor cells, as well as the major resistance strategies acquired by tumor cells that hamper the extrinsic and intrinsic apoptotic pathways related to NK cell-mediated killing.

Chemistry and biology of ophiobolin A and its congeners.

Ophiobolin A is a fungal secondary metabolite that was found to have significant activity against apoptosis-resistant glioblastoma cells through the induction of a non-apoptotic cell death, offering an innovative strategy to combat this aggressive cancer. The current article aims to make the bridge between the anti-cancer effects of ophiobolin A and its unique reaction with primary amines and suggests that pyrrolylation of lysine residues on its intracellular target protein(s) and/or phosphatidylethanolamine lipid is responsible for its biological effects. The article also discusses chemical derivatization of ophiobolin A to establish first synthetically generated structure-activity relationship. Finally, the reported total synthesis efforts toward the ophiobolin class of sesterterpenes are discussed and identified as a fertile area for improvement in pursuit of these molecules as anticancer agents.

Protein Tyrosine Phosphatase-N13 Promotes Myofibroblast Resistance to Apoptosis in Idiopathic Pulmonary Fibrosis.

RATIONALE: Idiopathic pulmonary fibrosis (IPF) is a progressive, fibrotic interstitial lung disease characterized by (myo)fibroblast accumulation and collagen deposition. Resistance to Fas-induced apoptosis is thought to facilitate (myo)fibroblast persistence in fibrotic lung tissues by poorly understood mechanisms. OBJECTIVES: To test the hypothesis that PTPN13 (protein tyrosine phosphatase-N13) is expressed by IPF lung (myo)fibroblasts, promotes their resistance to Fas-induced apoptosis, and contributes to the development of pulmonary fibrosis. METHODS: PTPN13 was localized in lung tissues from patients with IPF and control subjects by immunohistochemical staining. Inhibition of PTPN13 function in primary IPF and normal lung (myo)fibroblasts was accomplished by: 1) downregulation with TNF-α (tumor necrosis factor-α)/IFN-γ, 2) siRNA knockdown, or 3) a cell-permeable Fas/PTPN13 interaction inhibitory peptide. The role of PTPN13 in the development of pulmonary fibrosis was assessed in mice with genetic deficiency of PTP-BL, the murine ortholog of PTPN13. MEASUREMENTS AND MAIN RESULTS: PTPN13 was constitutively expressed by (myo)fibroblasts in the fibroblastic foci of patients with IPF. Human lung (myo)fibroblasts, which are resistant to Fas-induced apoptosis, basally expressed PTPN13 in vitro. TNF-α/IFN-γ or siRNA-mediated PTPN13 downregulation and peptide-mediated inhibition of the Fas/PTPN13 interaction in human lung (myo)fibroblasts promoted Fas-induced apoptosis. Bleomycin-challenged PTP-BL-/- mice, while developing inflammatory lung injury, exhibited reduced pulmonary fibrosis compared with wild-type mice. CONCLUSIONS: These findings suggest that PTPN13 mediates the resistance of human lung (myo)fibroblasts to Fas-induced apoptosis and promotes pulmonary fibrosis in mice. Our results suggest that strategies aimed at interfering with PTPN13 expression or function may represent a novel strategy to reduce fibrosis in IPF.

Caffeic Acid Phenethyl Ester Rescues Pulmonary Arterial Hypertension through the Inhibition of AKT/ERK-Dependent PDGF/HIF-1α In Vitro and In Vivo.

Pulmonary arterial hypertension (PAH) is characterized by pulmonary arterial proliferation and remodeling, resulting in a specific increase in right ventricle systolic pressure (RVSP) and, ultimately right ventricular failure. Recent studies have demonstrated that caffeic acid phenethyl ester (CAPE) exerts a protective role in NF-κB-mediated inflammatory diseases. However, the effect of CAPE on PAH remains to be elucidated. In this study, monocrotaline (MCT) was used to establish PAH in rats. Two weeks after the induction of PAH by MCT, CAPE was administrated by intraperitoneal injection once a day for two weeks. Pulmonary hemodynamic measurements and pulmonary artery morphological assessments were examined. Our results showed that administration of CAPE significantly suppressed MCT-induced vascular remodeling by decreasing the HIF-1α expression and PDGF-BB production, and improved in vivo RV systolic performance in rats. Furthermore, CAPE inhibits hypoxia- and PDGF-BB-induced HIF-1α expression by decreasing the activation of the AKT/ERK pathway, which results in the inhibition of human pulmonary artery smooth muscle cells (hPASMCs) proliferation and prevention of cells resistant to apoptosis. Overall, our data suggest that HIF-1α is regarded as an alternative target for CAPE in addition to NF-κB, and may represent a promising therapeutic agent for the treatment of PAH diseases.

Role of CCL20 mediated immune cell recruitment in NF-κB mediated TRAIL resistance of pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) represents one of the deadliest cancers. From a clinical view, the transcription factor NF-κB is of particular importance, since this pathway confers apoptosis resistance and limits drug efficacy. Whereas the role of the most abundant NF-κB subunit p65/RelA in therapeutic resistance is well documented, only little knowledge of the RelA downstream targets and their functional relevance in TRAIL mediated apoptosis in PDAC is available. In the present study TRAIL resistant and sensitive PDAC cell lines were analyzed for differentially expressed RelA target genes, to define RelA downstream targets mediating TRAIL resistance. The most upregulated target gene was then further functionally characterized. Unbiased genome-wide expression analysis demonstrated that the chemokine CCL20 represents the strongest TRAIL inducible direct RelA target gene in resistant PDAC cells. Unexpectedly, targeting CCL20 by siRNA, blocking antibodies or by downregulation of the sole CCL20 receptor CCR6 had no effect on PDAC cell death or cancer cell migration, arguing against an autocrine role of CCL20 in PDAC. However, by using an ex vivo indirect co-culture system we were able to show that CCL20 acts paracrine to recruit immune cells. Importantly, CCL20-recruited immune cells further increase TRAIL resistance of CCL20-producing PDAC cells. In conclusion, our data show a functional role of a RelA-CCL20 pathway in PDAC TRAIL resistance. We demonstrate how the therapy-induced cross-talk of cancer cells with immune cells affects treatment responses, knowledge needed to tailor novel bi-specific treatments, which target tumor cell as well as immune cells.

Synthetic analogues of the montanine-type alkaloids with activity against apoptosis-resistant cancer cells.

In a search of small molecules active against apoptosis-resistant cancer cells, a skeletal rearrangement of alkaloid haemanthamine was utilized to generate a series of compounds possessing the alkaloid montanine ring system. The synthesized compounds were found to inhibit proliferation of cancer cells resistant to apoptosis at micromolar concentrations. Selected compounds were also active against patient-derived glioblastoma cells expressing stem-cell markers. This is the first report describing the preparation of synthetic analogues of the montanine-type alkaloids with antiproliferative activity. The compounds prepared in the current investigation appear to be a useful starting point for the development of agents to fight cancers with apoptosis resistance, and thus, associated with poor prognoses.

Nrf2 mediates redox adaptation in NOX4-overexpressed non-small cell lung cancer cells.

The redox adaptation mechanisms in cancer cells are very complex and remain largely unclear. Our previous studies have confirmed that NADPH oxidase 4 (NOX4) is abundantly expressed in non-small cell lung cancer (NSCLC) and confers apoptosis resistance on NSCLC cells. However, the comprehensive mechanisms for NOX4-mediated oxidative resistance of cancer cells remain still undentified. The present study found that NOX4-derived H2O2 enhanced the nuclear factor erythroid 2-related factor 2 (Nrf2) stability via disruption of redox-dependent proteasomal degradation and stimulated its activity through activation of PI3K signaling. Specifically, the results showed that ectopic NOX4 expression did not induce apoptosis of A549 cells; however, inhibition of Nrf2 resulted in obvious apoptotic death of NOX4-overexpressed A549 cells, accompanied by a significant increase in H2O2 level and decrease in GSH content. Besides, inhibition of Nrf2 could suppress cell growth and efficiently reverse the enhancement effect of NOX4 on cell growth. The in vivo data confirmed that inhibition of Nrf2 could interfere apoptosis resistance in NOX4-overexpressed A549 tumors and led to cell growth inhibition. In conclusion, these results reveal that Nrf2 is critically involved in redox adaptation regulation in NOX4-overexpressed NSCLC cells. Therefore, NOX4 and Nrf2 may be promising combination targets against malignant progression of NSCLC.

TRAIL/NF-κB/CX3CL1 Mediated Onco-Immuno Crosstalk Leading to TRAIL Resistance of Pancreatic Cancer Cell Lines.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignant neoplasms and registers rising death rates in western countries. Due to its late detection in advanced stages, its extremely aggressive nature and the minimal effectiveness of currently available therapies, PDAC is a challenging problem in the clinical field. One characteristic of PDAC is a distinct desmoplasia consisting of fibroblasts, endothelial and immune cells as well as non-cellular components, contributing to therapy resistance. It is well established that the NF-κB signaling pathway controls inflammation, cancer progression and apoptosis resistance in PDAC. This study attempts to identify NF-κB target genes mediating therapy resistance of humane PDAC cell lines towards death ligand induced apoptosis. By using a genome wide unbiased approach the chemokine CX3CL1 was established as a central NF-κB target gene mediating therapy resistance. While no direct impact of CX3CL1 expression on cancer cell apoptosis was identified in co-culture assays it became apparent that CX3CL1 is acting in a paracrine fashion, leading to an increased recruitment of inflammatory cells. These inflammatory cells in turn mediate apoptosis resistance of PDAC cells. Therefore, our data dissect a bifunctional cross-signaling pathway in PDAC between tumor and immune cells giving rise to therapy resistance.