Serine synthesis and catabolism in starved lung cancer and primary bronchial epithelial cells.

Serine and glycine give rise to important building blocks in proliferating cells. Both amino acids are either synthesized de novo or taken up from the extracellular space. In lung cancer, serine synthesis gene expression is variable, yet, expression of the initial enzyme, phosphoglycerate dehydrogenase (PHGDH), was found to be associated with poor prognosis. While the contribution of de novo synthesis to serine pools has been shown to be enhanced by serine starvation, the impact of glucose deprivation, a commonly found condition in solid cancers is poorly understood. Here, we utilized a stable isotopic tracing approach to assess serine and glycine de novo synthesis and uptake in different lung cancer cell lines and normal bronchial epithelial cells in variable serine, glycine, and glucose conditions. Under low glucose supplementation (0.2 mM, 3-5% of normal plasma levels), serine de novo synthesis was maintained or even activated. As previously reported, also gluconeogenesis supplied carbons from glutamine to serine and glycine under these conditions. Unexpectedly, low glucose treatment consistently enhanced serine to glycine conversion, along with an up-regulation of the mitochondrial one-carbon metabolism enzymes, serine hydroxymethyltransferase (SHMT2) and methylenetetrahydrofolate dehydrogenase (MTHFD2). The relative contribution of de novo synthesis greatly increased in low serine/glycine conditions. In bronchial epithelial cells, adaptations occurred in a similar fashion as in cancer cells, but serine synthesis and serine to glycine conversion, as assessed by label enrichments and gene expression levels, were generally lower than in (PHGDH positive) cancer cells. In summary, we found a variable contribution of glucose or non-glucose carbon sources to serine and glycine and a high adaptability of the downstream one-carbon metabolism pathway to variable glucose supply.

Metformin Impedes Oxidation of LDL In Vitro.

Metformin is the most commonly prescribed glucose-lowering drug for the treatment of type 2 diabetes. The aim of this study was to investigate whether metformin is capable of impeding the oxidation of LDL, a crucial step in the development of endothelial dysfunction and atherosclerosis. LDL was oxidized by addition of CuCl2 in the presence of increasing concentrations of metformin. The extent of LDL oxidation was assessed by measuring lipid hydroperoxide and malondialdehyde concentrations, relative electrophoretic mobilities, and oxidation-specific immune epitopes. Cytotoxicity of oxLDL in the vascular endothelial cell line EA.hy926 was assessed using the alamarBlue viability test. Quantum chemical calculations were performed to determine free energies of reactions between metformin and radicals typical for lipid oxidation. Metformin concentration-dependently impeded the formation of lipid hydroperoxides, malondialdehyde, and oxidation-specific immune epitopes when oxidation of LDL was initiated by addition of Cu2+. The cytotoxicity of oxLDL was reduced when it was obtained under increasing concentrations of metformin. The quantum chemical calculations revealed that only the reaction of metformin with hydroxyl radicals is exergonic, whereas the reactions with hydroperoxyl radicals or superoxide radical anions are endergonic. Metformin, beside its glucose-lowering effect, might be a suitable agent to impede the development of atherosclerosis and associated CVD. This is due to its capability to impede LDL oxidation, most likely by scavenging hydroxyl radicals.

Circulating microRNAs as molecular biomarkers for lung adenocarcinoma.

BACKGROUND: The potential of microRNAs (miRNAs) as molecular tumor biomarkers for early diagnosis and prognosis in lung cancer is still unclear. OBJECTIVE: To analyze expression of miRNAs in A549 lung adenocarcinoma (LUAD) cells and in primary, non-malignant bronchial epithelial (BE) cells from healthy donors. To analyze the most prominently deregulated miRNAs in plasma samples of LUAD patients and healthy donors. MATERIALS AND METHODS: The expression of 752 miRNAs in LUAD and BE cells was assessed by RT-qPCR with mean-centering restricted normalization. The relative plasma levels of 18 miRNAs in LUAD patients and healthy donors were analyzed using RT-qPCR and normalized to miR-191-5p and miR-16-3p. Putative interactions between miRNAs and their target genes were investigated in silico. RESULTS: Out of 752 miRNAs, 37 miRNAs were significantly deregulated in A549 cells compared to BE cells. MiR-15b-3p, miR-148a-3p, miR-193b-3p, and miR-195-5p were significantly deregulated in plasma samples of LUAD patients compared to donors. The target genes of those four miRNAs are involved in essential mechanisms in cancer development and progression. CONCLUSIONS: There are substantial differences between cancer and control miRNA expression in vitro and in plasma samples of LUAD patients compared to healthy donors. Four deregulated miRNAs are promising as a diagnostic biomarker for adenocarcinoma of the lung.

Bisdemethoxycurcumin sensitizes the response of cisplatin resistant non-small cell lung carcinoma cell lines by activating apoptosis and autophagy.

Lung cancer belongs to the most frequent and deadliest cancer types worldwide, non-small cell lung carcinoma (NSCLC) being the most frequent type. Development of chemoresistance in NSCLC patients is common and responsible for bad outcome. Curcuminoids are naturally occurring substances with prominent cytotoxic effects in different cancer cells. Here we analyzed influence of bisdemethoxycurcumin (BDMC) on phenotype and molecular mechanisms in cisplatin-sensitive NSCLC cell lines (A549 and H460) and their cisplatin-resistant counterparts. NSCLC cell lines were exposed to BDMC and analyzed by cell viability, proliferation, and motility assays, as well as fluorescence-activated cell sorting. Immunoblotting was assessed to detect apoptosis and autophagy. Colony-formation assay and multicellular tumor spheroid model were used to investigate the effects of BDMC. Expression levels of different Hedgehog-pathway genes were determined by RT-qPCR analysis. We identified substantial cytotoxic effects of BDMC on NSCLC cells in general and on cisplatin-resistant NSCLC cells in special. BDMC markedly decreased the cell viability by inducing apoptosis and autophagy in a cell-type specific manner. BDMC emphasized cisplatin-induced cell death and inhibited cell cycle progression of cisplatin-resistant NSCLC cells. Scratch-closure, colony formation, and multicellular spheroid growth in cisplatin-resistant NSCLC cell lines were inhibited by BDMC. Expression profile analyses of different Hedgehog-pathway regulatory genes showed that Gli1, the mean transcriptional regulator of this pathway, was markedly decreased upon the BDMC treatment, this decrement being most prominent in cisplatin-resistant cells. Our data identified BDMC as a potent substance that may be suitable for combined cisplatin-based therapy in cisplatin-resistant subpopulation of NSCLC patients.

Only Subclinical Alterations in the Haemostatic System of People with Diabetes after COVID-19 Vaccination.

People with diabetes have an increased risk of experiencing adverse COVID-19 outcomes. COVID-19 vaccination is, therefore, highly recommended. However, people with diabetes have an inherently elevated risk of thrombotic events and the impact of the vaccination on the coagulation system in this patient population remains to be elucidated. The aim of this study was to investigate the impact of COVID-19 vaccination on the haemostatic system in people with type 1 or type 2 diabetes. We evaluated the effects of COVID-19 vaccination (BioNTech Pfizer, Moderna, AstraZeneca) on standard coagulation parameters, whole blood coagulation (Thrombelastometry), platelet function (impedance aggregation), and thrombin generation (calibrated automated thrombography) in people with type 1 diabetes mellitus (n = 41) and type 2 diabetes mellitus (n = 37). Blood sampling points were prior to vaccination and two weeks after the respective vaccination. Thrombelastometry measurements indicated moderately increased clot formation post-vaccination in people with type 1, as well as with type 2, diabetes: "Clot formation times" were significantly shorter, and both "maximum clot firmness" and "alpha angles" were significantly higher, as compared to the respective pre-vaccination values. Therefore, TEM parameters were not altered after vaccination in patients receiving ASA. Moreover, platelet aggregation was enhanced in people with type 1 diabetes, and plasma levels of D-Dimer were increased in people with type 2 diabetes, following COVID-19 vaccination. All other standard coagulation parameters, as well as thrombin generation, were not affected by the vaccination. The coagulation responses of people with diabetes to COVID-19 vaccination were only subclinical and comparable to those observed in healthy individuals. Our findings suggest that people with diabetes do not face an increased activation of the coagulation post-vaccination.

15d-PGJ2 Promotes ROS-Dependent Activation of MAPK-Induced Early Apoptosis in Osteosarcoma Cell In Vitro and in an Ex Ovo CAM Assay.

Osteosarcoma (OS) is the most common type of bone tumor, and has limited therapy options. 15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) has striking anti-tumor effects in various tumors. Here, we investigated molecular mechanisms that mediate anti-tumor effects of 15d-PGJ2 in different OS cell lines. Human U2-OS and Saos-2 cells were treated with 15d-PGJ2 and cell survival was measured by MTT assay. Cell proliferation and motility were investigated by scratch assay, the tumorigenic capacity by colony forming assay. Intracellular ROS was estimated by H2DCFDA. Activation of MAPKs and cytoprotective proteins was detected by immunoblotting. Apoptosis was detected by immunoblotting and Annexin V/PI staining. The ex ovo CAM model was used to study growth capability of grafted 15d-PGJ2-treated OS cells, followed by immunohistochemistry with hematoxylin/eosin and Ki-67. 15d-PGJ2 substantially decreased cell viability, colony formation and wound closure capability of OS cells. Non-malignant human osteoblast was less affected by 15d-PGJ2. 15d-PGJ2 induced rapid intracellular ROS production and time-dependent activation of MAPKs (pERK1/2, pJNK and pp38). Tempol efficiently inhibited 15d-PGJ2-induced ERK1/2 activation, while N-acetylcystein and pyrrolidine dithiocarbamate were less effective. Early but weak activation of cytoprotective proteins was overrun by induction of apoptosis. A structural analogue, 9,10-dihydro-15d-PGJ2, did not show toxic effects in OS cells. In the CAM model, we grafted OS tumors with U2-OS, Saos-2 and MG-63 cells. 15d-PGJ2 treatment resulted in significant growth inhibition, diminished tumor tissue density, and reduced tumor cell proliferation for all cell lines. Our in vitro and CAM data suggest 15d-PGJ2 as a promising natural compound to interfere with OS tumor growth.

PCK2 opposes mitochondrial respiration and maintains the redox balance in starved lung cancer cells.

Cancer cells frequently lack nutrients like glucose, due to insufficient vascular networks. Mitochondrial phosphoenolpyruvate carboxykinase, PCK2, has recently been found to mediate partial gluconeogenesis and hence anabolic metabolism in glucose starved cancer cells. Here we show that PCK2 acts as a regulator of mitochondrial respiration and maintains the redox balance in nutrient-deprived human lung cancer cells. PCK2 silencing increased the abundance and interconversion of tricarboxylic acid (TCA) cycle intermediates, augmented mitochondrial respiration and enhanced glutathione oxidation under glucose and serum starvation, in a PCK2 re-expression reversible manner. Moreover, enhancing the TCA cycle by PCK2 inhibition severely reduced colony formation of lung cancer cells under starvation. As a conclusion, PCK2 contributes to maintaining a reduced glutathione pool in starved cancer cells besides mediating the biosynthesis of gluconeogenic/glycolytic intermediates. The study sheds light on adaptive responses in cancer cells to nutrient deprivation and shows that PCK2 confers protection against respiration-induced oxidative stress.

Endothelial Lipase Modulates Paraoxonase 1 Content and Arylesterase Activity of HDL.

Endothelial lipase (EL) is a strong modulator of the high-density lipoprotein (HDL) structure, composition, and function. Here, we examined the impact of EL on HDL paraoxonase 1 (PON1) content and arylesterase (AE) activity in vitro and in vivo. The incubation of HDL with EL-overexpressing HepG2 cells decreased HDL size, PON1 content, and AE activity. The EL modification of HDL did not diminish the capacity of HDL to associate with PON1 when EL-modified HDL was incubated with PON1-overexpressing cells. The overexpression of EL in mice significantly decreased HDL serum levels but unexpectedly increased HDL PON1 content and HDL AE activity. Enzymatically inactive EL had no effect on the PON1 content of HDL in mice. In healthy subjects, EL serum levels were not significantly correlated with HDL levels. However, HDL PON1 content was positively associated with EL serum levels. The EL-induced changes in the HDL-lipid composition were not linked to the HDL PON1 content. We conclude that primarily, the interaction of enzymatically active EL with HDL, rather than EL-induced alterations in HDL size and composition, causes PON1 displacement from HDL in vitro. In vivo, the EL-mediated reduction of HDL serum levels and the consequently increased PON1-to-HDL ratio in serum increase HDL PON1 content and AE activity in mice. In humans, additional mechanisms appear to underlie the association of EL serum levels and HDL PON1 content.

Phospholipid dynamics in ex vivo lung cancer and normal lung explants.

In cancer cells, metabolic pathways are reprogrammed to promote cell proliferation and growth. While the rewiring of central biosynthetic pathways is being extensively studied, the dynamics of phospholipids in cancer cells are still poorly understood. In our study, we sought to evaluate de novo biosynthesis of glycerophospholipids (GPLs) in ex vivo lung cancer explants and corresponding normal lung tissue from six patients by utilizing a stable isotopic labeling approach. Incorporation of fully 13C-labeled glucose into the backbone of phosphatidylethanolamine (PE), phosphatidylcholine (PC), and phosphatidylinositol (PI) was analyzed by liquid chromatography/mass spectrometry. Lung cancer tissue showed significantly elevated isotopic enrichment within the glycerol backbone of PE, normalized to its incorporation into PI, compared to that in normal lung tissue; however, the size of the PE pool normalized to the size of the PI pool was smaller in tumor tissue. These findings indicate enhanced PE turnover in lung cancer tissue. Elevated biosynthesis of PE in lung cancer tissue was supported by enhanced expression of the PE biosynthesis genes ETNK2 and EPT1 and decreased expression of the PC and PI biosynthesis genes CHPT1 and CDS2, respectively, in different subtypes of lung cancer in publicly available datasets. Our study demonstrates that incorporation of glucose-derived carbons into the glycerol backbone of GPLs can be monitored to study phospholipid dynamics in tumor explants and shows that PE turnover is elevated in lung cancer tissue compared to normal lung tissue.

Distribution and prognostic significance of gluconeogenesis and glycolysis in lung cancer.

Inhibition of glycolysis has been considered as a therapeutic approach in aggressive cancers including lung cancer. Abbreviated gluconeogenesis, mediated by phosphoenolpyruvate carboxykinase (PEPCK), was recently discovered to partially circumvent the need for glycolysis in lung cancer cells. However, the interplay of glycolysis and gluconeogenesis in lung cancer is still poorly understood. Here, we analyzed the expression of GLUT1, the prime glucose transporter, and of PCK1 and PCK2, the cytoplasmic and mitochondrial isoforms of PEPCK, in 450 samples of non-small cell lung cancer (NSCLC) and in 54 NSCLC metastases using tissue microarrays and whole tumor sections. Spatial distribution was assessed by automated image analysis. Additionally, glycolytic and gluconeogenic gene expression was inferred from The Cancer Genome Atlas (TCGA) datasets. We found that PCK2 was preferentially expressed in the lung adenocarcinoma subtype, while GLUT1 expression was higher in squamous cell carcinoma. GLUT1 and PCK2 were inversely correlated, GLUT1 showing elevated expression in larger tumors while PCK2 was highest in smaller tumors. However, a mixed phenotype showing the presence of both, glycolytic and gluconeogenic cancer cells was frequent. In lung adenocarcinoma, PCK2 expression was associated with significantly improved overall survival, while the opposite was found for GLUT1. The metabolic tumor microenvironment and the 3-dimensional context play an important role in modulating both pathways, since PCK2 expression preferentially occurred at the tumor margin and hypoxia regulated both, glycolysis and gluconeogenesis, in NSCLC cells in vitro, albeit in opposite directions. PCK1/2 expression was enhanced in metastases compared to primary tumors, possibly related to the different environment. The results of this study show that glycolysis and gluconeogenesis are activated in NSCLC in a tumor size and oxygenation modulated manner and differentially correlate with outcome. The frequent co-activation of gluconeogenesis and glycolysis in NSCLC should be considered in potential future therapeutic strategies targeting cancer cell metabolism.

MicroRNA-182-5p regulates hedgehog signaling pathway and chemosensitivity of cisplatin-resistant lung adenocarcinoma cells via targeting GLI2.

Lung cancer is one of the deadliest cancers worldwide. Late diagnosis at an advanced, inoperable stage makes chemotherapy a treatment of choice, yet, with low response rates. The hedgehog signaling pathway (HHSP) is often reactivated in cancer. We identified miR-182-5p as a regulator of GLI2, a transcriptional regulator of the HHSP, and explored the role of the miR-182-5p/GLI2 axis in carcinogenesis and cisplatin resistance of lung adenocarcinoma (LADC). Expression of miRNAs and target genes was analyzed by RT-qPCR, expression of the GLI-protein family in LADC and adjacent lung tissue (n = 27 pairs) by immunohistochemistry. MiR-182-5p was manipulated, and data were generated by immunoblotting, immunofluorescence, apoptosis, proliferation/viability, dual-luciferase-, and colony forming assays. MiR-182-5p was down-regulated in cisplatin-resistant LADC cells and directly targeted GLI2. Interference with miR-182-5p or GLI2 silencing resulted in modulation of cell proliferation, clonogenic potential, and cisplatin-sensitivity. HHSP was markedly reactivated in LADC tissue compared to adjacent non-malignant lung tissue. Our results indicate that the miR-182-5p/GLI2 axis modulates tumorigenesis and cisplatin-resistance in LADC cells, by influencing the HHSP. Therefore, this axis might be considered as a potential biomarker and future therapeutic target in LADC patients.

Targeting TMEM16A to reverse vasoconstriction and remodelling in idiopathic pulmonary arterial hypertension.

Our systematic analysis of anion channels and transporters in idiopathic pulmonary arterial hypertension (IPAH) showed marked upregulation of the Cl- channel TMEM16A gene. We hypothesised that TMEM16A overexpression might represent a novel vicious circle in the molecular pathways causing pulmonary arterial hypertension (PAH).We investigated healthy donor lungs (n=40) and recipient lungs with IPAH (n=38) for the expression of anion channel and transporter genes in small pulmonary arteries and pulmonary artery smooth muscle cells (PASMCs).In IPAH, TMEM16A was strongly upregulated and patch-clamp recordings confirmed an increased Cl- current in PASMCs (n=9-10). These cells were depolarised and could be repolarised by TMEM16A inhibitors or knock-down experiments (n=6-10). Inhibition/knock-down of TMEM16A reduced the proliferation of IPAH-PASMCs (n=6). Conversely, overexpression of TMEM16A in healthy donor PASMCs produced an IPAH-like phenotype. Chronic application of benzbromarone in two independent animal models significantly decreased right ventricular pressure and reversed remodelling of established pulmonary hypertension.Our findings suggest that increased TMEM16A expression and activity comprise an important pathologic mechanism underlying the vasoconstriction and remodelling of pulmonary arteries in PAH. Inhibition of TMEM16A represents a novel therapeutic approach to reverse remodelling in PAH.

The glycerol backbone of phospholipids derives from noncarbohydrate precursors in starved lung cancer cells.

Cancer cells are reprogrammed to consume large amounts of glucose to support anabolic biosynthetic pathways. However, blood perfusion and consequently the supply with glucose are frequently inadequate in solid cancers. PEPCK-M (PCK2), the mitochondrial isoform of phosphoenolpyruvate carboxykinase (PEPCK), has been shown by us and others to be functionally expressed and to mediate gluconeogenesis, the reverse pathway of glycolysis, in different cancer cells. Serine and ribose synthesis have been identified as downstream pathways fed by PEPCK in cancer cells. Here, we report that PEPCK-M-dependent glycerol phosphate formation from noncarbohydrate precursors (glyceroneogenesis) occurs in starved lung cancer cells and supports de novo glycerophospholipid synthesis. Using stable isotope-labeled glutamine and lactate, we show that PEPCK-M generates phosphoenolpyruvate and 3-phosphoglycerate, which are at least partially converted to glycerol phosphate and incorporated into glycerophospholipids (GPL) under glucose and serum starvation. This pathway is required to maintain levels of GPL, especially phosphatidylethanolamine (PE), as shown by stable shRNA-mediated silencing of PEPCK-M in H23 lung cancer cells. PEPCK-M shRNA led to reduced colony formation after starvation, and the effect was partially reversed by the addition of dioleyl-PE. Furthermore, PEPCK-M silencing abrogated cancer growth in a lung cancer cell xenograft model. In conclusion, glycerol phosphate formation for de novo GPL synthesis via glyceroneogenesis is a newly characterized anabolic pathway in cancer cells mediated by PEPCK-M under conditions of severe nutrient deprivation.

No erythropoietin-induced growth is observed in non-small cell lung cancer cells.

Lung cancer patients have the highest incidence of anemia among patients with solid tumors. The use of recombinant human erythropoietin (Epo) has consistently been shown to reduce the need for blood transfusions and to increase hemoglobin levels in lung cancer patients with chemotherapy-induced anemia. However, clinical and preclinical studies have prompted concerns that Epo and the presence of its receptor, EpoR, in tumor cells may be responsible for adverse effects and, eventually, death. The question has been raised whether Epo promotes tumor growth and inhibits the death of cancer cells. In this study, we investigated the presence and functionality of EpoR, as well as the implications of Epo upon the proliferation and survival of lung cancer cells. Since the protein expression of both Epo and EpoR is induced by hypoxia, which is frequently present in lung cancer, the cells were treated with Epo under both normoxic and hypoxic conditions (1% O2). By using quantitative (real-time) PCR, western blot analysis, and immunocytochemical staining, three non-small cell lung cancer (NSCLC) cell lines (A427, A549 and NCI-H358) were analyzed for the expression of EpoR and its specific downstream signaling pathways [Janus kinase 2 (Jak2)-signal transducer and activator of transcription 5 (STAT5), phosphatidylinositol-3-kinase (PI3K)-Akt, mitogen-activated protein (MAP) kinase]. The effects of 100 U/ml Epo on cell proliferation and cisplatin-induced apoptosis were assessed. All NSCLC cell lines expressed EpoR mRNA and protein, while these levels differed considerably between the cell lines. We found the constitutive phosphorylation of EpoR and most of its downstream signaling pathways (STAT5, Akt and ERK1/2) independently of Epo administration. While Epo markedly enhanced the proliferation and reduced apoptosis of Epo-dependent UT-7/Epo leukemia cells, it did not affect tumor cell proliferation or the cisplatin-induced apoptosis of NSCLC cells. Thus, this in vitro study suggests that there are no tumor-promoting effects of Epo in the NSCLC cell lines studied, neither under normoxic nor under hypoxic conditions.

Impact of Endothelial Lipase on Cholesterol Efflux Capacity of Serum and High-density Lipoprotein.

Endothelial lipase (EL) is a potent modulator of the structural and functional properties of HDL. Impact of EL on cholesterol efflux capacity (CEC) of serum and isolated HDL is not well understood and apparently contradictory data were published. Here, we systematically examined the impact of EL on composition and CEC of serum and isolated HDL, in vitro and in vivo, using EL-overexpressing cells and EL-overexpressing mice. CEC was examined in a validated assay using 3H-cholesterol labelled J774 macrophages. In vitro EL-modification of serum resulted in complex alterations, including enrichment of serum with lipid-free/-poor apoA-I, decreased size of human (but not mouse) HDL and altered HDL lipid composition. EL-modification of serum increased CEC, in line with increased lipid-free/-poor apoA-I formation. In contrast, CEC of isolated HDL was decreased likely through altered lipid composition. In contrast to in vitro results, EL-overexpression in mice markedly decreased HDL-cholesterol and apolipoprotein A-I serum levels associated with a decreased CEC of serum. HDL lipid composition was altered, but HDL particle size and CEC were not affected. Our study highlights the multiple and complex effects of EL on HDL composition and function and may help to clarify the seemingly contradictory data found in published articles.

TR3 is involved in hypoxia-induced apoptosis resistance in lung cancer cells downstream of HIF-1α.

OBJECTIVES: Lung cancer is the leading cause of cancer death worldwide. Like in all solid tumors, hypoxia is common in lung cancer and contributes to apoptosis, and thus chemotherapy resistance. However, the underlying mechanisms are not entirely clear. TR3 (NR4A1, Nur77) is an orphan nuclear receptor that induces apoptosis and may mediate chemotherapy-induced apoptosis in cancer cells. MATERIALS AND METHODS: We used A549, H23 and H1299 cell lines to investigate how TR3-mediated apoptosis is affected by hypoxia in non-small cell lung cancer (NSCLC) cells. Cell culture, western blot analysis, apoptosis assay, and siRNA-mediated gene silencing were performed in this study. RESULTS AND CONCLUSION: The TR3 activator cytosporone B was used to investigate TR3-mediated apoptosis in NSCLC cells under normoxic and hypoxic conditions. Cytosporone B induced apoptosis in a concentration-dependent manner. Chronic moderate hypoxia induced a significant down-regulation of TR3. Accordingly, the cytosporone B effect was reduced under these conditions. Hypoxia-induced down-regulation of TR3 was mediated by hypoxia-inducible factor 1α. Our immunoblotting analysis and expression data from a public dataset suggest that TR3 is downregulated in NSCLC. In conclusion, our findings suggest that hypoxia-induced down-regulation of TR3 might play an important role for hypoxia-induced apoptosis resistance in NSCLC.

MicroRNAs as regulators of cisplatin-resistance in non-small cell lung carcinomas.

With more than 80% of all diagnosed lung cancer cases, non-small cell lung cancer (NSCLC) remains the leading cause of cancer death worldwide. Exact diagnosis is mostly very late and advanced-stage NSCLCs are inoperable at admission. Tailored therapies with tyrosine kinase inhibitors are only available for a minority of patients. Thus, chemotherapy is often the treatment of choice. As first-line chemotherapy for NSCLCs, platinum-based substances (e.g. cisplatin, CDDP) are mainly used. Unfortunately, the positive effects of CDDP are frequently diminished due to development of drug resistance and negative influence of microenvironmental factors like hypoxia. MicroRNAs (miRNAs) are small, non-coding molecules involved in the regulation of gene expression and modification of biological processes like cell proliferation, apoptosis and cell response to chemotherapeutics. Expression of miRNAs is often deregulated in lung cancer compared to corresponding non-malignant tissue. In this review we summarize the present knowledge about the effects of miRNAs on CDDP-resistance in NSCLCs. Further, we focus on miRNAs deregulated by hypoxia, which is an important factor in the development of CDDP-resistance in NSCLCs. This review will contribute to the general understanding of miRNA-regulated biological processes in NSCLC, with special focus on the role of miRNA in CDDP-resistance.

TASK-1 Regulates Apoptosis and Proliferation in a Subset of Non-Small Cell Lung Cancers.

Lung cancer is the leading cause of cancer deaths worldwide; survival times are poor despite therapy. The role of the two-pore domain K+ (K2P) channel TASK-1 (KCNK3) in lung cancer is at present unknown. We found that TASK-1 is expressed in non-small cell lung cancer (NSCLC) cell lines at variable levels. In a highly TASK-1 expressing NSCLC cell line, A549, a characteristic pH- and hypoxia-sensitive non-inactivating K+ current was measured, indicating the presence of functional TASK-1 channels. Inhibition of TASK-1 led to significant depolarization in these cells. Knockdown of TASK-1 by siRNA significantly enhanced apoptosis and reduced proliferation in A549 cells, but not in weakly TASK-1 expressing NCI-H358 cells. Na+-coupled nutrient transport across the cell membrane is functionally coupled to the efflux of K+ via K+ channels, thus TASK-1 may potentially influence Na+-coupled nutrient transport. In contrast to TASK-1, which was not differentially expressed in lung cancer vs. normal lung tissue, we found the Na+-coupled nutrient transporters, SLC5A3, SLC5A6, and SLC38A1, transporters for myo-inositol, biotin and glutamine, respectively, to be significantly overexpressed in lung adenocarcinomas. In summary, we show for the first time that the TASK-1 channel regulates apoptosis and proliferation in a subset of NSCLC.

JAZF1/SUZ12 gene fusion in endometrial stromal sarcomas.

Endometrial stromal sarcomas (ESSs) belong to the rarest uterine malignancies (prevalence category <1-9/1,000,000). According to the new 2014 World Health Organisation (WHO) classification, they are separated into four categories; benign endometrial stromal nodules (ESNs), low grade endometrial stromal sarcomas (LG-ESSs), high-grade endometrial stromal sarcomas (HG-ESSs) and undifferentiated uterine sarcomas (UUSs). Due to heterogeneous histopathologic appearance these tumors still represent diagnostic challenge, even for experienced pathologists. ESSs are genetically very heterogeneous and several chromosomal translocations and gene fusions have so far been identified in these malignancies. To date the JAZF1/SUZ12 gene fusion is by far the most frequent and seems to be the cytogenetic hallmark of ESN and LG-ESS. Based on present literature data this gene fusion is present in approximately 75% of ESN, 50% of LG-ESS and 15% of HG-ESS cases. The frequency of JAZF1/SUZ12 appearance varies between classic ESS and different morphologic variants. This gene fusion is suggested to become a specific diagnostic tool, especially in difficult borderline cases. In combination with the recently described YWHAE/FAM22 gene fusion the JAZF1/SUZ12 fusion could be used to differentiate between LG-ESS and HG-ESS. The purpose of this review is to summarize literature data published in last two and a half decades about this gene fusion, as a contribution to our understanding of ESS genetics and pathogenesis.