bims-meluca 2019-03-24 papers

on Metabolism of non-small cell lung carcinoma

Issue of 2019–03–24
nine 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

Cisplatin resistance involves a metabolic reprogramming through ROS and PGC-1α in NSCLC which can be overcome by OXPHOS inhibition.
Hypoxic non-small-cell lung cancer cell-derived exosomal miR-21 promotes resistance of normoxic cell to cisplatin.
Elevated heme synthesis and uptake underpin intensified oxidative metabolism and tumorigenic functions in non-small cell lung cancer cells.
S100A4 alters metabolism and promotes invasion of lung cancer cells by up-regulating mitochondrial complex I protein NDUFS2.
NF-κB upregulates glutamine-fructose-6-phosphate transaminase 2 to promote migration in non-small cell lung cancer.
Metabolic dependence of cyclosporine A on cell proliferation of human non‑small cell lung cancer A549 cells and its implication in post‑transplant malignancy.
Hypoxia-inducible factor-1 signaling pathway influences the sensitivity of HCC827 cells to gefitinib.
IKKβ targeting reduces KRAS-induced lung cancer angiogenesis in vitro and in vivo: A potential anti-angiogenic therapeutic target.
Serum Metabolomics Study of Non-Smoking Female Patients with Non-Small Cell Lung Cancer Using a Gas Chromatography-Mass Spectrometry.

Free Radic Biol Med. 2019 Mar 14. pii: S0891-5849(18)31582-X. [Epub ahead of print]135 167-181

Cisplatin resistance involves a metabolic reprogramming through ROS and PGC-1α in NSCLC which can be overcome by OXPHOS inhibition.

Alberto Cruz-Bermúdez, Raquel Laza-Briviesca, Ramiro J Vicente-Blanco, Aránzazu García-Grande, Maria José Coronado, Sara Laine-Menéndez, Sara Palacios-Zambrano, M Rocío Moreno-Villa, Asunción Martin Ruiz-Valdepeñas, Cristina Lendinez, Atocha Romero, Fernando Franco, Virginia Calvo, Cristina Alfaro, Paloma Martin Acosta, Clara Salas, José Miguel Garcia, Mariano Provencio.

   BACKGROUND: Platinum-based chemotherapy remains the standard of care for most lung cancer cases. However chemoresistance is often developed during the treatment, limiting clinical utility of this drug. Recently, the ability of tumor cells to adapt their metabolism has been associated to resistance to therapies. In this study, we first described the metabolic reprogramming of Non-Small Cell Lung Cancer (NSCLC) in response to cisplatin treatment.
METHODS: Cisplatin-resistant versions of the A549, H1299, and H460 cell lines were generated by continuous drug exposure. The long-term metabolic changes, as well as, the early response to cisplatin treatment were analyzed in both, parental and cisplatin-resistant cell lines. In addition, four Patient-derived xenograft models treated with cisplatin along with paired pre- and post-treatment biopsies from patients were studied. Furthermore, metabolic targeting of these changes in cell lines was performed downregulating PGC-1α expression through siRNA or using OXPHOS inhibitors (metformin and rotenone).
RESULTS: Two out of three cisplatin-resistant cell lines showed a stable increase in mitochondrial function, PGC1-α and mitochondrial mass with reduced glycolisis, that did not affect the cell cycle. This phenomenon was confirmed in vivo. Post-treatment NSCLC tumors showed an increase in mitochondrial mass, PGC-1α, and a decrease in the GAPDH/MT-CO1 ratio. In addition, we demonstrated how a ROS-mediated metabolism reprogramming, involving PGC-1α and increased mitochondrial mass, is induced during short-time cisplatin exposure. Moreover, we tested how cells with increased PGC-1a induced by ZLN005 treatment, showed reduced cisplatin-driven apoptosis. Remarkably, the long-term metabolic changes, as well as the metabolic reprogramming during short-time cisplatin exposure can be exploited as an Achilles' heel of NSCLC cells, as demonstrated by the increased sensitivity to PGC-1α interference or OXPHOS inhibition using metformin or rotenone.
CONCLUSION: These results describe a new cisplatin resistance mechanism in NSCLC based on a metabolic reprogramming that is therapeutically exploitable through PGC-1α downregulation or OXPHOS inhibitors.

Keywords:  Chemoresistance; Chemotherapy; Metabolism; Metformin; NSCLC

DOI:  https://doi.org/10.1016/j.freeradbiomed.2019.03.009
Onco Targets Ther. 2019 ;12 1947-1956

Hypoxic non-small-cell lung cancer cell-derived exosomal miR-21 promotes resistance of normoxic cell to cisplatin.

Caijun Dong, Xingwang Liu, Huisheng Wang, Jutao Li, Lei Dai, Jun Li, Zhen Xu.

   Purpose: To explore the effects of hypoxic non-small-cell lung cancer (NSCLC)-derived exosomes on NSCLC resistance to cisplatin.
Materials and methods: Exosomes were isolated by differential centrifugation and characterized by transmission electron microscope and Western blotting. Quantitative real-time PCR was used to measure miR-21 levels. MTT assays and colony formation assays were performed to investigate the effects of hypoxia-induced exosomes on cisplatin resistance.
Results: Hypoxic NSCLC cell-derived exosomes facilitate normoxic cell resistance to cisplatin. In addition, hypoxia enhanced the miR-21 expression in NSCLC cells and cell-derived exosomes. Interestingly, changes in miR-21 levels in the hypoxia-induced exosomes affected the sensitivity of recipient cells to cisplatin. Mechanically, exosomal miR-21 promoted cisplatin resistance by downregulating phosphatase and tensin homolog (PTEN). The expression of miR-21 in tumor cell lines and clinical NSCLC tumor samples was positively correlated with hypoxia-inducible factor-1α and negatively correlated with PTEN. Moreover, high miR-21 expression was associated with shorter median survival period in patients undergoing pharmacotherapy, but no association was observed in patients who were not under pharmacotherapy.
Conclusion: Exosomal miR-21 derived from hypoxic NSCLC cells may promote cisplatin resistance, which indicates that exosomal miR-21 might be a potential biomarker and therapeutic target to address NSCLC chemoresistance.

Keywords:  PTEN; cisplatin resistance; exosomes; miR-21; non-small-cell lung cancer

DOI:  https://doi.org/10.2147/OTT.S186922
Cancer Res. 2019 Mar 22. pii: canres.2156.2018. [Epub ahead of print]

Elevated heme synthesis and uptake underpin intensified oxidative metabolism and tumorigenic functions in non-small cell lung cancer cells.

Sagar Sohoni, Poorva Ghosh, Tianyuan Wang, Sarada Preeta Kalainayakan, Chantal Vidal, Sanchareeka Dey, Purna Chaitanya Konduri, Li Zhang.

Tumors of human non-small cell lung cancer (NSCLC) are heterogeneous but exhibit elevated glycolysis and glucose oxidation relative to benign lung. Heme is a central molecule for oxidative metabolism and ATP generation via mitochondrial oxidative phosphorylation (OXPHOS). Here we showed that levels of heme synthesis and uptake, mitochondrial heme, oxygen-utilizing hemoproteins, oxygen consumption, ATP generation, and key mitochondrial biogenesis regulators were enhanced in NSCLC cells relative to non-tumorigenic cells. Likewise, proteins and enzymes relating to heme and mitochondrial functions were upregulated in human NSCLC tissues relative to normal tissues. Engineered heme-sequestering peptides (HSP) reduced heme uptake, intracellular heme levels, and tumorigenic functions of NSCLC cells. Addition of heme largely reversed the effect of HSP on tumorigenic functions. Furthermore, HSP2 significantly suppressed the growth of human NSCLC xenograft tumors in mice. HSP2-treated tumors exhibited reduced oxygen consumption rates and ATP levels. To further verify the importance of heme in promoting tumorigenicity, we generated NSCLC cell lines with increased heme synthesis or uptake by overexpressing either the rate-limiting heme synthesis enzyme ALAS1 or uptake protein SLC48A1, respectively. These cells exhibited enhanced migration and invasion and accelerated tumor growth in mice. Notably, tumors formed by cells with increased heme synthesis or uptake also displayed elevated oxygen consumption rates and ATP levels. These data show that elevated heme flux and function underlie enhanced OXPHOS and tumorigenicity of NSCLC cells. Targeting heme flux and function offers a potential strategy for developing therapies for lung cancer.

DOI: https://doi.org/10.1158/0008-5472.CAN-18-2156
J Biol Chem. 2019 Mar 18. pii: jbc.RA118.004365. [Epub ahead of print]

S100A4 alters metabolism and promotes invasion of lung cancer cells by up-regulating mitochondrial complex I protein NDUFS2.

Lili Liu, Lei Qi, Teresa Knifley, Dava W Piecoro, Piotr Rychahou, Jinpeng Liu, Mihail I Mitov, Jeremiah Martin, Chi Wang, Jianrong Wu, Heidi L Weiss, D Allan Butterfield, B Mark Evers, Kathleen L O'Connor, Min Chen.

  It is generally accepted that alterations in metabolism are critical for the metastatic process; however, the mechanisms by which these metabolic changes are controlled by the major drivers of the metastatic process remain elusive. Here, we found that S100 calcium-binding protein A4 (S100A4), a major metastasis-promoting protein, confers metabolic plasticity to drive tumor invasion and metastasis of non-small cell lung cancer cells. Investigating how S100A4 regulates metabolism, we found that S100A4 depletion decreases oxygen consumption rates, mitochondrial activity, and ATP production and also shifts cell metabolism to higher glycolytic activity. We further identified that the 49 KD mitochondrial complex I subunit NADH dehydrogenase (ubiquinone) Fe-S protein 2 (NDUFS2) is regulated in an S100A4-dependent manner and that S100A4 and NDUFS2 exhibit co-occurrence at significant levels in various cancer types as determined by database-driven analysis of genomes in clinical samples using cBioPortal for Cancer Genomics. Importantly, we noted that S100A4 or NDUFS2 silencing inhibits mitochondrial complex I activity, reduces cellular ATP level, decreases invasive capacity in three-dimensional (3D) growth, and dramatically decreases metastasis rates as well as tumor growth in vivo. Finally, we provide evidence that cells depleted in S100A4 or NDUFS2 shift their metabolism toward glycolysis by up-regulating hexokinase expression and that suppressing S100A4 signaling sensitizes lung cancer cells to glycolysis inhibition. Our findings uncover a novel S100A4 function and highlight its importance in controlling NDUFS2 expression to regulate the plasticity of mitochondrial metabolism and thereby promote the invasive and metastatic capacity in lung cancer.

Keywords:  Fibroblast specific protein-1; NADH:ubiquinone oxidoreductase core subunit S2 (NDUFS2); S100 calcium-binding protein A4 (S100A4); S100 proteins; energy metabolism; glycolysis; invasion; lung cancer; metastasis; metastasis-1; mitochondria; mitochondrial complex I; mitochondrial respiratory chain complex

DOI:  https://doi.org/10.1074/jbc.RA118.004365
Cell Commun Signal. 2019 Mar 18. 17(1): 24

NF-κB upregulates glutamine-fructose-6-phosphate transaminase 2 to promote migration in non-small cell lung cancer.

Szymon J Szymura, Jacob P Zaemes, David F Allison, Sheena H Clift, Jaclyn M D'Innocenzi, Lisa G Gray, Brian D McKenna, Benjamin B Morris, Stefan Bekiranov, Robin D LeGallo, David R Jones, Marty W Mayo.

   BACKGROUND: Epithelial-to-mesenchymal transition (EMT) results in changes that promote de-differentiation, migration, and invasion in non-small cell lung cancer (NSCLC). While it is recognized that EMT promotes altered energy utilization, identification of metabolic pathways that link EMT with cancer progression is needed. Work presented here indicates that mesenchymal NSCLC upregulates glutamine-fructose-6-phosphate transaminase 2 (GFPT2). GFPT2 is the rate-limiting enzyme in the synthesis of uridine diphosphate N-acetylglucosamine (UDP-GlcNAc). UDP-GlcNAc is the obligate activator of O-linked N-acetylglucosamine transferase (OGT).
METHODS: Analysis of our transcriptomic data indicates that GFPT2 is one of the most significantly upregulated metabolic genes in mesenchymal NSCLC. Ectopic GFPT2 expression, as well as gene silencing strategies were used to determine the importance of this metabolic enzyme in regulating EMT-driven processes of cell motility and invasion.
RESULTS: Our work demonstrates that GFPT2 is transcriptionally upregulated by NF-κB and repressed by the NAD+-dependent deacetylase SIRT6. Depletion of GFPT2 expression in NSCLC highlights its importance in regulating cell migration and invasion during EMT.
CONCLUSIONS: Consistent with GFPT2 promoting cancer progression, we find that elevated GFPT2 expression correlates with poor clinical outcome in NSCLC. Modulation of GFPT2 activity offers a potentially important therapeutic target to combat NSCLC disease progression.

Keywords:  Cell migration; Epithelial-mesenchymal transition; Glutamine-fructose-6-phosphate transaminase 2 (GFPT2); Non-small cell lung cancer (NSCLC); Nuclear factor kappa B (NF-κB); Sirtuin-6 (SIRT6)

DOI:  https://doi.org/10.1186/s12964-019-0335-5
Oncol Rep. 2019 Mar 18.

Metabolic dependence of cyclosporine A on cell proliferation of human non‑small cell lung cancer A549 cells and its implication in post‑transplant malignancy.

Xinghua Qin, Ziwei Chen.

Cyclosporine A (CsA), a widely used immunosuppressant to prevent organ transplant rejection, is associated with an increased cancer risk following transplantation, particularly in the lung. However, the underlying mechanisms remain unclear. In the present study, using human non‑small cell lung cancer A549 cells, it was determined that CsA (0.1 or 1 µM) promoted cell proliferation with glucose alone as the energy source. CsA treatment increased the phosphorylation of protein kinase B (Akt) and consequently the expression of Cyclin D1. Inhibiting Akt signaling with the phosphatidylinositol 3‑kinase inhibitor wortmannin prevented this effect. Mechanistically, CsA treatment increased reactive oxygen species (ROS) generation, and the intracellular ROS scavenger N‑acetyl‑cysteine (NAC) attenuated CsA‑induced cell proliferation as well as the activation of Akt/Cyclin D1 signaling. However, notably, it was demonstrated that CsA treatment decreased cell proliferation and Akt phosphorylation under normal lipid loading. Further investigation indicated that palmitic acid induced excessive generation of ROS, while CsA treatment further stimulated this ROS production. Scavenging intracellular ROS with NAC attenuated the CsA‑mediated inhibition of cell proliferation. Collectively, the results indicated a pleiotropic effect of CsA in the regulation of A549 cell proliferation under different metabolic conditions. This indicated that CsA administration may contribute to increased post‑transplant cancer risk in organ recipients.

DOI: https://doi.org/10.3892/or.2019.7076
Oncol Lett. 2019 Apr;17(4): 4034-4043

Hypoxia-inducible factor-1 signaling pathway influences the sensitivity of HCC827 cells to gefitinib.

Qian Jin, Jianying Zhou, Xianrong Xu, Feihua Huang, Weihua Xu.

  The majority of patients with non-small cell lung cancer (NSCLC) with activating epidermal growth factor receptor (EGFR) mutations inevitably progress in stage despite an initial substantial and rapid response to EGFR-tyrosine kinase inhibitors (EGFR-TKIs). Previous research indicates that hypoxia may be associated with resistance to EGFR-TKIs in EGFR mutation-positive NSCLC. Therefore, the present study regulated the activity of hypoxia-inducible factor-1 (HIF-1) signaling pathway to observe if it is able to alter the sensitivity of lung cancer cells to gefitinib. The present study selected 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1) and dimethyloxalylglycine (DMOG) as a HIF-1 signaling pathway inhibitor and activator, respectively, on HCC827 cells. Cells were incubated with different treatments for different durations: A blank control, DMOG, gefitinib, or DMOG and gefitinib combined, for 36 and 48 h; and then a blank control, YC-1, gefitinib, or YC-1 and gefitinib combined, for 16 and 28 h. A western blot analysis assay was performed to evaluate the protein expression levels of HIF-1α and phosphorylated hepatocyte growth factor receptor (p-MET), an MTT assay was used to determine cell proliferation, a colony formation assay was used to investigate the colony-forming ability and a wound healing assay was used to test the cell migration ability. Additionally, Pearson's correlation analysis was used to evaluate the correlation between p-Met and HIF-1α expression levels. Finally, it was identified that gefitinib and DMOG combined notably improve the growth and cell migration ability of HCC827 cells, compared with gefitinib alone. When gefitinib and YC-1 were combined, the inhibiting effect on the growth and cell migration ability of HCC827 cells was substantially enhanced, compared with the control cells. Pearson's correlation analysis revealed that the p-Met expression level had a strong positive correlation with HIF-1α expression levels. Thus, it was concluded that the HIF-1 signaling pathway influences the sensitivity of HCC827 cells to gefitinib. The positive correlation between p-Met and HIF-1α expression levels may be the underlying mechanism of the HIF-1 signaling pathway influencing the sensitivity of HCC827 cells to gefitinib.

Keywords:  3-(5′-hydroxymethyl-2′-furyl)-1-benzylindazole; gefitinib; hypoxia-inducible factor-1; oxalylglycine; phosphorylated hepatocyte growth factor receptor

DOI:  https://doi.org/10.3892/ol.2019.10025
Lung Cancer. 2019 Apr;pii: S0169-5002(19)30343-5. [Epub ahead of print]130 169-178

IKKβ targeting reduces KRAS-induced lung cancer angiogenesis in vitro and in vivo: A potential anti-angiogenic therapeutic target.

Tatiana Correa Carneiro-Lobo, Luiza Coimbra Scalabrini, Leila da Silva Magalhães, Laura B Cardeal, Felipe Silva Rodrigues, Edmilson Ozorio Dos Santos, Albert S Baldwin, Elena Levantini, Ricardo J Giordano, Daniela Sanchez Bassères.

   OBJECTIVES: The ability of tumor cells to drive angiogenesis is an important cancer hallmark that positively correlates with metastatic potential and poor prognosis. Therefore, targeting angiogenesis is a rational therapeutic approach and dissecting proangiogenic pathways is important, particularly for malignancies driven by oncogenic KRAS, which are widespread and lack effective targeted therapies. Based on published studies showing that oncogenic RAS promotes angiogenesis by upregulating the proangiogenic NF-κB target genes IL-8 and VEGF, that NF-κB activation by KRAS requires the IKKβ kinase, and that targeting IKKβ reduces KRAS-induced lung tumor growth in vivo, but has limited effects on cell growth in vitro, we hypothesized that IKKβ targeting would reduce lung tumor growth by inhibiting KRAS-induced angiogenesis.
MATERIALS AND METHODS: To test this hypothesis, we targeted IKKβ in KRAS-mutant lung cancer cell lines either by siRNA-mediated transfection or by treatment with Compound A (CmpdA), a highly specific IKKβ inhibitor, and used in vitro and in vivo assays to evaluate angiogenesis.
RESULTS AND CONCLUSIONS: Both pharmacological and siRNA-mediated IKKβ targeting in lung cells reduced expression and secretion of NF-κB-regulated proangiogenic factors IL-8 and VEGF. Moreover, conditioned media from IKKβ-targeted lung cells reduced human umbilical vein endothelial cell (HUVEC) migration, invasion and tube formation in vitro. Furthermore, siRNA-mediated IKKβ inhibition reduced xenograft tumor growth and vascularity in vivo. Finally, IKKβ inhibition also affects endothelial cell function in a cancer-independent manner, as IKKβ inhibition reduced pathological retinal angiogenesis in a mouse model of oxygen-induced retinopathy. Taken together, these results provide a novel mechanistic understanding of how the IKKβ pathway affects human lung tumorigenesis, indicating that IKKβ promotes KRAS-induced angiogenesis both by cancer cell-intrinsic and cancer cell-independent mechanisms, which strongly suggests IKKβ inhibition as a promising antiangiogenic approach to be explored for KRAS-induced lung cancer therapy.

Keywords:  Angiogenesis; IKKβ; IL-8; KRAS; Lung cancer; Therapeutic target; VEGF

DOI:  https://doi.org/10.1016/j.lungcan.2019.02.027
J Proteome Res. 2019 Mar 20.

Serum Metabolomics Study of Non-Smoking Female Patients with Non-Small Cell Lung Cancer Using a Gas Chromatography-Mass Spectrometry.

Ying Mu, Yang Zhou, Yanfeng Wang, Wei Li, Lina Zhou, Xin Lu, Peng Gao, Mingyang Gao, Yanhui Zhao, Qi Wang, Yanfu Wang, Guowang Xu.

The incidence of non-smoking female patients with non-small cell lung cancer (NSCLC) is increasing in recent decades. However, the pathogenesis of patients is unclear and early diagnosis biomarkers are in urgent need. In this study, 136 non-smoking female subjects (65 patients with NSCLC, 6 patients with benign lung tumors and 65 healthy controls) were enrolled, and their metabolic profiling was investigated by using a pseudotargeted gas chromatography-mass spectrometry. A total of 56 annotated metabolites were found and verified to be significantly different in non-smoking female with NSCLC compared with the control. The metabolic profiling was featured by disturbed energy metabolism, amino acid metabolism, oxidative stress, lipid metabolism, etc. Cysteine, serine and 1-monooleoylglycerol were defined as the biomarker panel for the diagnosis of NSCLC patients. 98.5% and 91.4% of subjects were correctly distinguished in the discovery and validation sets, respectively. The biomarker panel was also useful for the diagnosis of in-situ malignancy patients with the accuracy of 97.7% and 97.8% in the discovery and validation sets, respectively. The study provides a biomarker panel for the auxiliary diagnosis of non-smoking female with NSCLC.

DOI: https://doi.org/10.1021/acs.jproteome.9b00069