CD14(+)S100A9(+) monocytic myeloid-derived suppressor cells and their clinical relevance in non-small cell lung cancer.

RATIONALE: Myeloid-derived suppressor cells (MDSCs) are a heterogeneous family of myeloid cells that suppress T-cell immunity in tumor-bearing hosts. Their clinical relevance remains unclear. OBJECTIVES: To identify subtypes of myeloid-derived suppressor cells in patients with non-small cell lung cancer (NSCLC) and their clinical relevance. METHODS: CD11b(+)CD14(-) and CD11b(+)CD14(+) cells, determined and phenotyped by fluorescence-activated cell sorter analysis, in the peripheral blood mononuclear cells (PBMCs) of treatment-naive patients with advanced NSCLC were correlated with clinical data. T-cell activation in response to CD3/CD28 costimulation was determined by carboxy-fluorescein diacetate succinimidyl ester (CFSE) staining and ELISA analysis of IFN-γ. The percentage of CD11b(+)CD14(+)S100A9(+) cells in PBMCs was correlated with and tested as a predictor for treatment response in a cohort of patients prospectively receiving first-line cisplatin-based chemotherapy. MEASUREMENTS AND MAIN RESULTS: Patients with NSCLC had a significantly higher ratio of CD11b(+)CD14(+) cells than healthy subjects, which was correlated with poor performance status and poor response to chemotherapy. The depletion of these cells in the PBMC reversed the suppression of CD8(+) and CD4(+) T cells. Isolated CD11b(+)CD14(+) cells suppressed CD8(+) T-cell proliferation and IFN-γ production, and the former effect was attenuated by the inducible nitric oxide synthase (iNOS) inhibitor aminoguanidine hydrochloride, arginase inhibitor N-hydroxy-nor-l-arginine (nor-NOHA), and blocking antibodies for IL-4Rα(+) and IL-10. CD11b(+)CD14(+) cells were monocyte-like, expressing CD33(+), CD15(-/low), IL-4Rα(+), and S100A9(+) and producing iNOS, arginase, and several cytokines. The ratio of S100A9(+) cells positively correlated with the suppressive ability of the CD11b(+)CD14(+) cells, was associated with poor response to chemotherapy, and predicted shorter progression-free survival. CONCLUSIONS: CD14(+)S100A9(+) inflammatory monocytes in patients with NSCLC are a distinct subset of MDSCs, which suppress T cells by arginase, iNOS, and the IL-13/IL-4Rα axis. The amount of these inflammatory monocytes is associated with poor response to chemotherapy. Clinical trial registered with www.clinicaltrials.gov (NCT 01204307).

Reduced nuclear factor-κB repressing factor: a link toward systemic inflammation in COPD.

Chronic systemic inflammation is implicated in the systemic manifestations and, probably, the excess mortality risk of chronic obstructive pulmonary disease (COPD). The role of nuclear factor (NF)-κB repressing factor (NRF), a DNA-binding, protein-inhibiting NF-κB response gene, in human diseases has not been explored. We hypothesised that the NRF-negative regulatory mechanism is impaired in COPD peripheral blood mononuclear cells (PBMCs) leading to excessive interleukin (IL)-8/CXCL8 production. NRF expression, NF-κB activation, IL-8/CXCL8 release and intracellular oxidative stress were assessed in PBMCs of normal subjects and stable COPD patients. Primary PBMCs with NRF overexpression, NRF knockdown and exposure to H(2)O(2) were used to elucidate the mechanisms. Stable COPD patients, especially those with severe COPD, showed decreased NRF expression, enhanced NF-κB activation and increased IL-8/CXCL8 release in PBMCs compared with normal subjects. This was associated with reduced NRF and increased RNA polymerase II occupancy at the IL-8/CXCL8 promoter. NRF knockdown enhanced IL-8/CXCL8 production in normal PBMCs, whilst NRF overexpression attenuated IL-8/CXCL8 production. Intracellular oxidative stress was increased in COPD PBMCs. H(2)O(2)-decreased NRF expression and -enhanced IL-8/CXCL8 production was augmented in COPD PBMCs. NRF expression is reduced in PBMCs of stable COPD patients, probably through oxidative stress, leading to increased production of IL-8/CXCL8 and potentially chronic systemic inflammation.

Neutrophil elastase represses IL-8/CXCL8 synthesis in human airway smooth muscle cells through induction of NF-kappa B repressing factor.

NF-kappaB repressing factor (NRF), a nuclear inhibitor of NF-kappaB, is constitutively expressed and is implicated in the basal silencing of specific NF-kappaB targeting genes, including IFN-beta, IL-8/CXCL8, and iNOS. Little is known about the regulation of NRF and its role in response to stimuli. Airway smooth muscle (ASM) is a rich source of inflammatory mediators that may regulate the development and progression of airway inflammation. We have previously reported that NE activates NF-kappaB in primary human ASM (hASM), leading to induction of TGF-beta1. In this study, we describe that, instead of inducing the NF-kappaB response gene IL-8/CXCL8, NE suppressed IL-8/CXCL8 release and mRNA expression in hASM cells. Transcriptional blockade studies using actinomycin D revealed a similar degradation rate of IL-8/CXCL8 mRNA in the presence or absence of NE, suggesting an involvement at the transcription level. Mechanistically, the NE repressive effect was mediated by inducing NRF, as shown by RT-PCR and Western blotting, which was subsequently recruited to the native IL-8/CXCL8 promoter leading to removal of RNA polymerase II from the promoter, as demonstrated by chromatin immunoprecipitation assays. Knockdown of NRF by small interfering RNA prevented NE-induced suppression of IL-8/CXCL8 expression. In contrast, NE did not induce NRF expression in A549 and Beas-2B cells, where NE only stimulates NF-kappaB activation and IL-8/CXCL8 induction. Forced expression of NRF in A549 cells by an NRF expression plasmid suppressed IL-8/CXCL8 expression. Hence, we describe a novel negative regulatory mechanism of NE-induced NRF, which is restricted to hASM and mediates the suppression of IL-8/CXCL8 expression.

Chemotherapy Immunophenoprofiles in Non-Small-Cell Lung Cancer by Personalized Membrane Proteomics.

OBJECTIVES: No study has addressed how the immune status at the molecular level is affected by first-line pemetrexed and cisplatin (PEM-CIS) combination therapy in patients with non-small-cell lung cancer (NSCLC). Thus, we aimed to identify the immune status from membrane proteome alterations in patients with NSCLC upon PEM-CIS treatment. METHODS: The paired peripheral blood mononuclear cells (PBMCs) were collected from four patients with lung adenocarcinoma before and after the first regimen of PEM-CIS treatment and applied quantitative membrane proteomics analysis. RESULT: In the personalized PBMC membrane proteome profiles, 2424 proteins were identified as displaying patient-specific responsive patterns. We discovered an elevated neutrophil activity and a more suppressive T-cell phenotype with the downregulation of cytotoxic T lymphocyte antigen 4 degradation and the upregulation of type 2 T-helper and T-regulatory cells in the patient with the highest progression-free survival (PFS) of 14.5 months. Patients with a PFS of 2 months showed higher expressions of T-cell subsets, MHC class II pathways, and T-cell receptor signaling, which indicated an activated immune status. CONCLUSION AND CLINICAL RELEVANCE: Without the additional isolation of specific immune cell populations, our study demonstrated that PEM-CIS chemotherapy altered patients' immune system in terms of neutrophils, T cells, and antigen presentation pathways.

Correction: S100A9+ MDSC and TAM-mediated EGFR-TKI resistance in lung adenocarcinoma: the role of RELB.

[This corrects the article DOI: 10.18632/oncotarget.24146.].

S100A9+ MDSC and TAM-mediated EGFR-TKI resistance in lung adenocarcinoma: the role of RELB.

BACKGROUND: Monocytic myeloid-derived suppressor cells (MDSCs), particularly the S100A9+ subset, has been shown initial clinical relevance. However, its role in EGFR-mutated lung adenocarcinoma, especially to EGFR-tyrosine kinase inhibitor (EGFR-TKI) is not clear. In a clinical setting of EGFR mutated lung adenocarcinoma, a role of the MDSC apart from T cell suppression was also investigated. RESULTS: Blood monocytic S100A9+ MDSC counts were higher in lung cancer patients than healthy donors, and were associated with poor treatment response and shorter progression-free survival (PFS). S100A9+ MDSCs in PBMC were well correlated to tumor infiltrating CD68+ and S100A9+ cells, suggesting an origin of TAMs. Patient's MDMs, mostly from S100A9+ MDSC, similar to primary alveolar macrophages from patients, both expressed S100A9 and CD206, attenuated EGFR-TKI cytotoxicity. Microarray analysis identified up-regulation of the RELB signaling genes, confirmed by Western blotting and functionally by RELB knockdown. CONCLUSIONS: In conclusion, blood S100A9+ MDSC is a predictor of poor treatment response to EGFR-TKI, possibly via its derived TAMs through activation of the non-canonical NF-κB RELB pathway. METHODS: Patients with activating EGFR mutation lung adenocarcinoma receiving first line EGFR TKIs were prospectively enrolled. Peripheral blood mononuclear cells (PBMCs) were collected for MDSCs analysis and for monocyte-derived macrophages (MDMs) and stored tissue for TAM analysis by IHC. A transwell co-culture system of MDMs/macrophages and H827 cells was used to detect the effect of macrophages on H827 and microarray analysis to explore the underlying molecular mechanisms, functionally confirmed by RNA interference.

Noncanonical NF-κB mediates the Suppressive Effect of Neutrophil Elastase on IL-8/CXCL8 by Inducing NKRF in Human Airway Smooth Muscle.

Neutrophil elastase (NE) suppresses IL-8/CXCL8 in human airway smooth muscle cells (hASM) while stimulating its production in respiratory epithelial cells. This differential effect is mediated by the selective induction of NKRF and dysregulation in chronic inflammatory diseases. We hypothesized that the differential activation of NF-κB subunits confer the opposite effect of NKRF on IL-8/CXCL8 in primary hASM and A549 cells stimulated with NE. The events occurring at the promoters of NKRF and IL-8/CXCL8 were observed by ChIP assays, and the functional role of RelB was confirmed by knockdown and overexpression. Although p65 was stimulated in both cell types, RelB was only activated in NE-treated hASM, as confirmed by NF-κB DNA binding ELISA, Western blotting and confocal microscopy. Knockdown of RelB abolished the induction of NKRF and converted the suppression of IL-8/CXCL8 to stimulation. The forced expression of RelB induced NKRF production in hASM and A549 cells. NE activated the NIK/IKK1/RelB non-canonical NF-κB pathway in hASM but not in A549. The nuclear-translocated RelB was recruited to the NKRF promoter around the putative κB site, accompanied by p52 and RNA polymerase II. In conclusion, NFRF is a novel RelB-response gene, and NE is a stimulator of the non-canonical RelB/NF-κB pathway in hASM.

SUV39H1 Reduction Is Implicated in Abnormal Inflammation in COPD.

Chronic obstructive pulmonary disease(COPD) is characterized by enhanced chronic inflammation in the airways, lung parenchyma, and circulation. We investigated whether SUV39H1, a histone methyltransferase, is causatively implicated in the abnormal inflammation observed in COPD. The SUV39H1 and H3K9me3 levels were reduced in peripheral blood mononuclear cells(PBMCs), primary human small airway epithelial cells(HSAEpCs) and lung tissues from COPD patients, which were correlated with poor lung function and the serum IL-8 and IL-6 levels. A specific SUV39H1 inhibitor, chaetocin, induced a distinct COPD panel of inflammatory cytokines in normal PBMCs. Mechanistically, chaetocin reduced the SUV39H1 and H3K9me3 levels in the native IL-8 promoter in normal HSAEpCs, which mimicked unstimulated COPD HSAEpCs and led to decreased HP-1α levels and increased RNA polymerase II levels. SUV39H1 knockdown reproduced the pattern of COPD inflammation, whereas SUV39H1 overexpression in COPD HSAEpCs rescued the H3K9me3 levels and suppressed inflammation. In COPD mice, chaetocin further repressed the SUV39H1/H3K9me3 levels and enhanced inflammation. SUV39H1 epigenetically controls a distinct panel of pro-inflammatory cytokines. Its reduction in COPD leads to a loss of the repressive chromatin mark H3K9me3 and confers an abnormal inflammatory response to stimulators. SUV39H1 and its regulatory pathways are potential therapeutic targets for COPD.

Corrigendum: SUV39H1 Reduction Is Implicated in Abnormal Inflammation in COPD.

This corrects the article DOI: 10.1038/srep46667.

Regulation of AKR1B1 by thyroid hormone and its receptors.

The objective of this study was to identify genes regulated by thyroid hormone (T(3)) mediated by its receptor (TR) and associated with tumorigenesis. The gene encoding aldo-keto reductase family 1, member B1 (AKR1B1), as previously identified by c-DNA microarray, is known to be up-regulated by T(3) treatment. Enzyme AKR1B1 was elevated roughly 3-fold in HepG2-TRalpha1 cells at the protein level and 4.6-fold increase at the mRNA level after 48 h T(3) treatment. Similar findings were obtained from thyroidectomized rats after T(3) application. To identify and localize the critical TR element (TRE), series deletion of the promoter mutant were constructed and electrophoretic mobility shift assays were carried out. The TRE on the AKR1B1 promoter was localized to the -1099/-1028 region. Further, this study demonstrated that AKR1B1 over-expression in some types of hepatocellular carcinomas (HCCs) is TR-dependent and might play a crucial role in the development of HCC. Thus, T(3) regulates AKR1B1 gene expression via a TRE-dependant mechanism and associates liver cancer.