bims-meluca 2019-06-23 papers

on Metabolism of non-small cell lung carcinoma

Issue of 2019–06–23
ten papers selected by
the Muñoz-Pinedo/Nadal (PReTT) lab, L’Institut d’Investigació Biomèdica de Bellvitge and Cristina Muñoz Pinedo, L’Institut d’Investigació Biomèdica de Bellvitge

Lung adenocarcinoma-intrinsic GBE1 signaling inhibits anti-tumor immunity.
Hypoxia inducible factor-1α/B-cell lymphoma 2 signaling impacts radiosensitivity of H1299 non-small cell lung cancer cells in a normoxic environment.
18F-FDG PET metabolic-to-morphological volume ratio predicts PD-L1 tumour expression and response to PD-1 blockade in non-small-cell lung cancer.
CHTM1 regulates cancer cell sensitivity to metabolic stress via p38-AIF1 pathway.
Cytochalasin H Inhibits Angiogenesis via the Suppression of HIF-1α Protein Accumulation and VEGF Expression through PI3K/AKT/P70S6K and ERK1/2 Signaling Pathways in Non-Small Cell Lung Cancer Cells.
Prognostic Value of Ferritin-to-Hemoglobin Ratio in Patients with Advanced Non-Small-Cell Lung Cancer.
High Folic Acid Supplementation Reprograms Glycolytic and Lactate-generating Metabolism to Promote Anchorage-independent Tumor Spheroid Formation in NSCLC Cells (P05-001-19).
Effect of butein and glucose on oxidative stress and p38 activation marker in non-small cell lung cancer cell.
Paradoxical relationship between body mass index and bone mineral density in patients with non-small cell lung cancer with brain metastasis.
Cholesterol Biosynthesis Is Required for Cell Death in Response to G9A Inhibitor in Lung Cancer (P05-010-19).

Mol Cancer. 2019 Jun 20. 18(1): 108

Lung adenocarcinoma-intrinsic GBE1 signaling inhibits anti-tumor immunity.

Lifeng Li, Li Yang, Shiqi Cheng, Zhirui Fan, Zhibo Shen, Wenhua Xue, Yujia Zheng, Feng Li, Dong Wang, Kai Zhang, Jingyao Lian, Dan Wang, Zijia Zhu, Jie Zhao, Yi Zhang.

   BACKGROUND: Changes in glycogen metabolism is an essential feature among the various metabolic adaptations used by cancer cells to adjust to the conditions imposed by the tumor microenvironment. Our previous study showed that glycogen branching enzyme (GBE1) is downstream of the HIF1 pathway in hypoxia-conditioned lung cancer cells. In the present study, we investigated whether GBE1 is involved in the immune regulation of the tumor microenvironment in lung adenocarcinoma (LUAD).
METHODS: We used RNA-sequencing analysis and the multiplex assay to determine changes in GBE1 knockdown cells. The role of GBE1 in LUAD was evaluated both in vitro and in vivo.
RESULTS: GBE1 knockdown increased the expression of chemokines CCL5 and CXCL10 in A549 cells. CD8 expression correlated positively with CCL5 and CXCL10 expression in LUAD. The supernatants from the GBE1 knockdown cells increased recruitment of CD8+ T lymphocytes. However, the neutralizing antibodies of CCL5 or CXCL10 significantly inhibited cell migration induced by shGBE1 cell supernatants. STING/IFN-I pathway mediated the effect of GBE1 knockdown for CCL5 and CXCL10 upregulation. Moreover, PD-L1 increased significantly in shGBE1 A549 cells compared to those in control cells. Additionally, in LUAD tumor tissues, a negative link between PD-L1 and GBE1 was observed. Lastly, blockade of GBE1 signaling combined with anti-PD-L1 antibody significantly inhibited tumor growth in vivo.
CONCLUSIONS: GBE1 blockade promotes the secretion of CCL5 and CXCL10 to recruit CD8+ T lymphocytes to the tumor microenvironment via the IFN-I/STING signaling pathway, accompanied by upregulation of PD-L1 in LUAD cells, suggesting that GBE1 could be a promising target for achieving tumor regression through cancer immunotherapy in LUAD.

Keywords:  Anti-tumor immunity; GBE1; PD-L1; STING pathway; T cell infiltration; Type I interferon

DOI:  https://doi.org/10.1186/s12943-019-1027-x
Radiat Environ Biophys. 2019 Jun 15.

Hypoxia inducible factor-1α/B-cell lymphoma 2 signaling impacts radiosensitivity of H1299 non-small cell lung cancer cells in a normoxic environment.

Gang Wang, Liang Xiao, Fen Wang, Jing Yang, Li Yang, Ye Zhao, Wensen Jin.

  Hypoxia inducible factor-1α (HIF-1α) is a critical transcriptional factor for the response of cells to hypoxic microenvironment and its expression induces resistance of hypoxic non-small-cell lung cancer (NSCLC) cells to radiotherapy. This study investigated how the activation of HIF-1α/B-cell lymphoma 2 (BCL-2) signaling under normoxic conditions impacted radiosensitivity of NSCLC cells. The recombinant pcDNA3.0-EGFP plasmids with wild-type or mutant HIF-1α complementary DNA (cDNA) were transfected into H1299 cells, an NSCLC cell line, establishing two H1299 sublines with high expression of HIF-1α. Compared with the levels of HIF-1α and BCL-2 proteins in non-transfected cells, increased levels of both proteins were found in transfected cells. Moreover, the expression of HIF-1α in non-transfected cells induced by chloride cobalt (CoCl2), a commonly used mimetic hypoxia reagent, was concomitant with the enhancement of BCL-2 expression. Conversely, reduction of HIF-1α expression by an inhibitor decreased the levels of BCL-2 proteins. The results revealed that the stabilization and expression of HIF-1α promoted the accumulation of BCL-2 proteins in H1299 cells. Subsequent experiments showed that intracellular HIF-1α/BCL-2 signaling was triggered in a normoxic environment after H1299 cells were exposed to irradiation, causing an elevated radioresistance. In contrast, blockage of HIF-1α/BCL-2 signaling leads to an elevated radiosensitivity. Proliferation of cells assay showed that, under normoxic conditions, population doubling times (PDTs) of irradiated cells were prolonged by suppression of HIF-1α/BCL-2 signaling. It is, therefore, indicated that HIF-1α/BCL-2 signaling activated by ionizing radiation reduces the radiosensitivity of H1299 cells independent of the hypoxic environment.

Keywords:  B-cell lymphoma 2; Hypoxia inducible factor-1α; Non-small cell lung cancer; Radiosensitivity

DOI:  https://doi.org/10.1007/s00411-019-00802-4
Eur J Nucl Med Mol Imaging. 2019 Jun 18.

18F-FDG PET metabolic-to-morphological volume ratio predicts PD-L1 tumour expression and response to PD-1 blockade in non-small-cell lung cancer.

Mario Jreige, Igor Letovanec, Kariman Chaba, Stephanie Renaud, Sylvie Rusakiewicz, Valerie Cristina, Solange Peters, Thorsten Krueger, Laurence de Leval, Lana E Kandalaft, Marie Nicod-Lalonde, Pedro Romero, John O Prior, George Coukos, Niklaus Schaefer.

   PURPOSE: Anti-PD-1/PD-L1 blockade can restore tumour-specific T-cell immunity and is an emerging therapy in non-small-cell lung cancer (NSCLC). We investigated the correlation between 18F-FDG PET/CT-based markers and tumour tissue expression of PD-L1, necrosis and clinical outcome in patients receiving checkpoint inhibitor treatment.
METHODS: PD-Li expression in biopsy or resection specimens from 49 patients with confirmed NSCLC was investigated by immunohistochemistry. Maximum standardized uptake value (SUVmax), mean SUV (SUVmean), metabolic tumour volume (MTV) and total lesion glycolysis (TLG) were obtained from 18F-FDG PET/CT images. The ratio of metabolic to morphological lesion volumes (MMVR) and its association with PD-L1 expression in each lesion were calculated. The associations between histologically reported necrosis and 18F-FDG PET imaging patterns and radiological outcome (evaluated by iRECIST) following anti-PD-1/PD-L1 therapy were also analysed. In 14 patients, the association between necrosis and MMVR and tumour immune contexture were analysed by multiple immunofluorescent (IF) staining for CD8, PD-1, granzyme B (GrzB) and NFATC2.
RESULTS: In total, 25 adenocarcinomas and 24 squamous cell carcinomas were analysed. All tumours showed metabolic 18F-FDG PET uptake. MMVR was correlated inversely with PD-L1 expression in tumour cells. Furthermore, PD-L1 expression and low MMVR were significantly correlated with clinical benefit. Necrosis was correlated negatively with MMVR. Multiplex IF staining showed a greater frequency of activated CD8+ cells in necrotic tumours than in nonnecrotic tumours in both stromal and epithelial tumour compartments.
CONCLUSION: This study introduces MMVR as a new imaging biomarker and its ability to noninvasively capture increased PD-L1 tumour expression and predict clinical benefit from checkpoint blockade in NSCLC should be further evaluated.

Keywords:  FDG; Imaging; NSCLC; PD-L1; PET/CT

DOI:  https://doi.org/10.1007/s00259-019-04348-x
J Exp Clin Cancer Res. 2019 Jun 20. 38(1): 271

CHTM1 regulates cancer cell sensitivity to metabolic stress via p38-AIF1 pathway.

Mansi Babbar, Ying Huang, Christopher M Curtiss, M Saeed Sheikh.

   BACKGROUND: Recently, we have reported the characterization of a novel protein named Coiled-coil Helix Tumor and Metabolism 1 (CHTM1). CHTM1 localizes to both cytosol and mitochondria. Sequence corresponding to CHTM1 is also annotated in the database as CHCHD5. CHTM1 is deregulated in human breast and colon cancers and its deficiency in human cancer cells leads to defective lipid metabolism and poor growth under glucose/glutamine starvation.
METHODS: Human cancer cell lines and tissue specimens were used. CHTM1 knockdown was done via lentiviral approach. CHTM1-expresssion constructs were developed and mutants were generated via site-directed mutagenesis approach. Western blotting, immunostaining, immunohistochemistry, cell fractionation and luciferase assays were performed. Reactive oxygen species and reactive nitrogen species were also measured.
RESULTS: Here we report that CHTM1 deficiency sensitizes human lung cancer cells to metabolic stress-induced cell death mediated by glucose/glutamine deprivation and metformin treatment. CHTM1 interacts with Apoptosis Inducing Factor 1 (AIF1) that is one of the important death inducing molecules. CHTM1 appears to negatively regulate AIF1 by preventing AIF1 translocation to cytosol/nucleus and thereby inhibit AIF1-mediated caspase-independent cell death. Our results also indicate that p38, a stress kinase, plays a critical role in metabolic stress-induced cell death in CHTM1-deficient cells. Furthermore, p38 appears to enhance AIF1 translocation from mitochondria to cytosol particularly in metabolically stressed CHTM1-deficient cells and CHTM1 negatively regulates p38 kinase activity. The expression status of CHTM1 in lung cancer patient samples is also investigated and our results indicate that CHTM1 levels are increased in the majority of lung tumors when compared to their matching normal tissues.
CONCLUSION: Thus, CHTM1 appears to be an important metabolic marker that regulates cancer cell survival under metabolic stress conditions, and has the potential to be developed as a predictive tumor marker.

Keywords:  AIF; CHCHD5; Cancer metabolism; Cell death; Lung cancer; Metabolic stress; p38

DOI:  https://doi.org/10.1186/s13046-019-1253-5
J Cancer. 2019 ;10(9): 1997-2005

Cytochalasin H Inhibits Angiogenesis via the Suppression of HIF-1α Protein Accumulation and VEGF Expression through PI3K/AKT/P70S6K and ERK1/2 Signaling Pathways in Non-Small Cell Lung Cancer Cells.

Yuefan Ma, Zihan Xiu, Zhiyuan Zhou, Bingyu Huang, Jiao Liu, Xiaofeng Wu, Sanzhong Li, Xudong Tang.

  Our previous study has demonstrated that cytochalasin H (CyH) isolated from mangrove-derived endophytic fungus induces apoptosis and inhibits migration in A549 non-small cell lung cancer (NSCLC) cells. In this study, we further explored the effect of CyH on angiogenesis in NSCLC cells and the underlying molecular mechanisms. A549 and H460 NSCLC cells were treated with different concentrations of CyH for 24 h. The effects of CyH on NSCLC angiogenesis in vitro and in vivo were investigated. Hypoxia inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF) expression in xenografted NSCLC of nude mice was analyzed by immunohistochemistry. ELISA was used to analyze the concentration of VEGF in the conditioned media derived from treated and untreated NSCLC cells. Western blot was performed to detect the levels of HIF-1α, p-AKT, p-P70S6K, and p-ERK1/2 proteins, and RT-qPCR was used to determine the levels of HIF-1α and VEGF mRNA in A549 and H460 cells. Our results showed that CyH significantly inhibited angiogenesis in vitro and in vivo, and suppressed the hemoglobin content and HIF-1α and VEGF protein expression in xenografted NSCLC tissues of nude mice. Meanwhile, CyH inhibited the secretion of VEGF protein and the expression of HIF-1α protein in A549 and H460 cells. Moreover, CyH had a significant inhibitory effect on VEGF mRNA expression but had no effect on HIF-1α mRNA expression, and CyH inhibited HIF-1α protein expression by promoting the degradation of HIF-1α protein in A549 and H460 cells. Additionally, CyH dramatically inhibited AKT, P70S6K, and ERK1/2 activation in A549 and H460 cells. Taken together, our results suggest that CyH can inhibit NSCLC angiogenesis by the suppression of HIF-1α protein accumulation and VEGF expression through PI3K/AKT/P70S6K and ERK1/2 signaling pathways.

Keywords:  ERK1/2; PI3K/AKT/P70S6K; angiogenesis; cytochalasin H (CyH); hypoxia inducible factor-1α (HIF-1α); mangrove; non-small cell lung cancer (NSCLC); vascular endothelial growth factor (VEGF)

DOI:  https://doi.org/10.7150/jca.29933
J Cancer. 2019 ;10(7): 1717-1725

Prognostic Value of Ferritin-to-Hemoglobin Ratio in Patients with Advanced Non-Small-Cell Lung Cancer.

Sookyung Lee, Hyeonjin Jeon, Bumsang Shim.

   BACKGROUND: Among commonly used biomarkers that reflect overall health in patients with cancer, hemoglobin is an iron-containing, oxygen-carrying protein in red blood cells, and serum ferritin is an iron-storage protein. This study investigated the ability of the ferritin-to-hemoglobin ratio to predict survival in patients with advanced non-small-cell lung cancer (NSCLC).
METHODS: The medical records of patients with pathologically confirmed advanced NSCLC were retrospectively reviewed. The ferritin level, hemoglobin level, and ferritin-to-hemoglobin ratio at the initiation of treatment were investigated. After descriptive analysis of the ferritin-to-hemoglobin ratio, the optimal diagnostic cutoff value for survival was determined using receiver operating characteristic analysis. After dichotomizing patients according to the optimal cutoff value, the prognostic effect of the ferritin-to-hemoglobin ratio was assessed. Overall survival (OS) was calculated using Kaplan-Meier analysis and compared using log-rank tests. Cox proportional hazards regression was used to evaluate the prognostic effect with respect to survival.
RESULTS: Of the enrolled patients, 91.3% had stage IV NSCLC, 42.0% had an Eastern Cooperative Oncology Group-performance status (ECOG-PS) score of 2, and 56.5% previously underwent systemic chemotherapy. The median OS of enrolled patients was 11.5 months. The range of the ferritin-to-hemoglobin ratio was 0.6-294.2, and the optimal cutoff value of the ferritin-to-hemoglobin ratio for survival was 13.0 (sensitivity, 58.5%; specificity, 80.0%; area under the curve = 0.68; P = 0.004). The median OS of patients with a low ferritin-to-hemoglobin ratio (<13.0) was 19.7 months, whereas that of patients with a high ferritin-to-hemoglobin ratio (≥13.0) was 8.5 months (P < 0.001). After eliminating confounding factors such as age, sex, ECOG-PS, histologic type, and C-reactive protein level, a high ferritin-to-hemoglobin ratio was significantly associated with poor survival. The multivariate proportional hazards model revealed that the ferritin-to-hemoglobin ratio was an independent prognostic marker for survival (hazard ratio, 1.91; 95% confidence interval, 1.27-2.88; P = 0.002).
CONCLUSION: The ferritin-to-hemoglobin ratio, a potential parameter of tumor progression, was a significant prognostic factor for OS, with a direct correlation to survival time in patients with advanced NSCLC.

Keywords:  ferritin; hemoglobinn; non-small cell lung cancer; prognosis; survival

DOI:  https://doi.org/10.7150/jca.26853
Curr Dev Nutr. 2019 Jun;pii: nzz030.P05-001-19. [Epub ahead of print]3(Suppl 1):

High Folic Acid Supplementation Reprograms Glycolytic and Lactate-generating Metabolism to Promote Anchorage-independent Tumor Spheroid Formation in NSCLC Cells (P05-001-19).

Yi-Wen Chen, Rwei-Fen S Huang.

Objectives: Aims of this study were to investigate the effect of HF on anchorage-independent tumor spheroid formation and its' working mechanisms of lung malignant tumor cells.
Methods: Human NSCLC cells A549 were cultured in control (C) medium (2.2 μM folic acid) or HF medium (10, 30, 50 μM folic acid) for 4 days. Cells were harvested to explore the self-renewal capacity of cancer cells by observing the anchorage-independent tumor spheroid formation. Meanwhile, the L-lactate assay were conducted to evaluate the lactate-generating state and the cell protein extract for the western blotting analysis to realize expression of energy metabolism related proteins in cells.
Results: The results showed that numbers of tumor spheroids in HF group were significantly higher than the control group. Besides, compared to C group, HF50 group showed significantly reduced lactate release into medium with highly accumulation in cellular lactate levels. The glycolytic-related protein expression of hexokinase II (HKII), lactate dehydrogenase (LDH) were increased and glucose transporter 1 (GLUT1), pyruvate dehydrogenase (PDH) were decreased in HF group. The signaling-related protein expression of insulin receptor substract-1 (IRS-1), hypoxia-inducible factor-1 alpha (HIF-1α) were increased and PI3 kinase (PI3K), AMP-activated protein kinase (AMPK) were decreased in HF group.
Conclusions: Collectively, HF supplementation may reprogram glycolytic metabolism and increase anchorage-independent tumor spheroids formation to mediate malignant progress of NSCLC.
Funding Sources: Ministry of Science and Technology, Taiwan, R.O.C.

DOI: https://doi.org/10.1093/cdn/nzz030.P05-001-19
Hum Exp Toxicol. 2019 Jun 19. 960327119851250

Effect of butein and glucose on oxidative stress and p38 activation marker in non-small cell lung cancer cell.

C Zhang, X Zong, Y Han.

   INTRODUCTION: Tumor microenvironment is known to alter the anticancer drug efficiency. One of the factors that get altered in cancer microenvironment is glucose concentration. Butein, an active principle from plant, known to have anticancer effect against different types of tumor. The objective of the study is to determine the effect of butein on glucose exposed non-small cell lung cancer cells (NSCLCCs).
METHODS: The current study deals with the effect of butein (6.25-50μM) on NSCLCCs treated with different concentrations (0-40 mM) of glucose.
RESULTS AND DISCUSSION: Glucose concentration, 0 mM and 40 mM, was found to be lethal at 72 h. Viable cell numbers were statistically increased in 5-mM, 10-mM, and 20-mM glucose-treated cells. Butein at 12.5 µM inhibits (p < 0.05) glucose-induced cell proliferation. Butein inhibits glucose-induced proliferation through DNA damage and oxidative stress. Mitochondrial reactive oxygen species (ROS) level was elevated in 20-mM glucose-treated cells when compared to 5-mM glucose-treated cells, whereas butein treatment further increases glucose-induced mitochondrial ROS. Pharmacological inhibitor of glycolysis, such as 2-deoxy glucose (2-DG), was found to inhibit (p < 0.05) glucose-induced cells proliferation. Furthermore, 2-DG and butein showed synergistic anticancer effect. Butein treatment increases p38 phosphorylation. Inhibition of p38 phosphorylation and antioxidant pretreatment partially revert the glucose-induced cell proliferation. However, inhibition of p38 phosphorylation combined with antioxidant pretreatment completely reverts the anticancer effect of butein. The present study concludes through the evidence that butein could serve as a potential anticancer compound in tumor microenvironment.

Keywords:  Lung cancer; butein; microenvironment; oxidative stress; p38

DOI:  https://doi.org/10.1177/0960327119851250
PLoS One. 2019 ;14(6): e0218825

Paradoxical relationship between body mass index and bone mineral density in patients with non-small cell lung cancer with brain metastasis.

Min Woo Nam, Jae Min Kim, Jin Hwan Cheong, Je Il Ryu, Myung-Hoon Han.

BACKGROUND AND PURPOSE: Low body mass index (BMI) at presentation has been reported to be associated with higher incidence and mortality of lung cancer, but studies on the relationship between brain metastasis and BMI at presentation are lacking. This study aimed to evaluate the association between brain metastasis and BMI and bone mineral density (BMD) in NSCLC.
METHODS: We retrospectively enrolled patients with non-small cell lung cancer who underwent brain magnetic resonance imaging with contrast within 3 months of diagnosis. The BMI was collected, and the BMD was measured in Hounsfield unit (HU) on initial staging computed tomography scans. The independent relationship between BMI and BMD was assessed using multivariable linear regression according to the presence of brain metastasis.
RESULTS: A total of 356 consecutive NSCLC patients were enrolled in the study over a 8-year period in a single institution. Lower BMI with higher BMD was an independent predictive factor for brain metastasis in patients with NSCLC, relative to the other group (HR, 2.03; 95% CI, 1.21 to 3.40; P = 0.007). We also found a significant negative correlation between BMI and BMD among patients with NSCLC with brain metastases (B, -3.343; 95% confidence interval, -6.352 to -0.333; P = 0.030).
CONCLUSIONS: Brain metastasis may possibly be associated with lower BMI and higher BMD in NSCLC patients. We expect that these results may facilitate future predictions of brain metastases during the clinical course of NSCLC and enhance our understanding of the underlying mechanisms that link brain metastases and lung cancer.

DOI: https://doi.org/10.1371/journal.pone.0218825
Curr Dev Nutr. 2019 Jun;pii: nzz030.P05-010-19. [Epub ahead of print]3(Suppl 1):

Cholesterol Biosynthesis Is Required for Cell Death in Response to G9A Inhibitor in Lung Cancer (P05-010-19).

Ha Eun Kim, Seo Yoon Choi, Yoon Jung Park.

Objectives: G9A is a eukaryotic histone methyltransferase that contributes to transcriptional silencing of tumor suppressor genes by modulating histone 3 lysine 9 methylation (H3K9m). It has been recently suggested as a promising therapeutic target for breast cancer and neuroblastoma. This study was aimed to investigate the effect of G9A inhibition and its cellular metabolic mechanisms.
Methods: Using OncomineTM, G9A overexpression in lung cancer was assessed. Cell viability upon treatment of G9A inhibitor (BIX01294, BIX) and siG9A was measured by MTT and IncuCyteR assays. Additionally, apoptosis and autophagy were analyzed through western blots. In order to identify targets, transcriptomes using RNA sequencing was conducted upon BIX treatment. Further functional relevance of targets was validated using Chromatin IP and recovery tests.
Results: BIX-mediated inhibition of G9A reduced cell viability of lung cancer cells via induction of autophagy. Through RNA sequencing, we found that G9A inhibition mainly affected cholesterol biosynthesis pathway. BIX directly induced the expression of SREBF2 gene, by lowering H3K9me1 and H3K9me2 at the promoter. A cholesterol biosynthesis inhibitor, 25-HC, partially recovered BIX-induced cell death by attenuating autophagy. Our data suggests that cholesterol metabolism can be a potential therapeutic target by G9A inhibition and its induction of autophagic cell death.
Conclusions: Our data suggests that cholesterol metabolism can be a potential therapeutic target by G9A inhibition and its induction of autophagic cell death.
Funding Sources: NRF of Korea grant (2018R1D1A1B07051274); BK21 Plus Project (22A20130012143).

DOI: https://doi.org/10.1093/cdn/nzz030.P05-010-19