ABSTRACT
Ferroptosis is a recently discovered form of cell death that plays an important role in tumor growth and holds promise as a target for antitumor therapy. However, evidence in the regulation of ferroptosis in lung adenocarcinoma (LUAD) remains elusive. Here, we show that retinoic acid receptor alpha (RARA) is upregulated with the treatment of ferroptosis inducers (FINs). Pharmacological activation of RARA increases the resistance of LUAD to ferroptosis according to cell viability and lipid peroxidation assays, while RARA inhibitor or knockdown (KD) does the opposite. Through transcriptome sequencing in RARA-KD cells and chromatin immunoprecipitation (CHIP)-Seq data, we identify thioredoxin (TXN) and protein phosphatase 1 F (PPM1F) as downstream targets of RARA, both of which inhibit ferroptosis. We confirm that RARA binds to the promotor region of TXN and PPM1F and promotes their transcription by CHIP-qPCR and dual-luciferase assays. Overexpression of TXN and PPM1F reverses the effects of RARA knockdown on ferroptosis in vitro and vivo. Clinically, RARA knockdown or inhibitor increases cells' sensitivity to pemetrexed and cisplatin (CDDP). Immunohistochemistry (IHC) of LUAD from our cohort shows the same expression tendency of RARA and the downstream targets. Our study uncovers that RARA inhibits ferroptosis in LUAD by promoting TXN and PPM1F, and inhibiting RARA-TXN/PPM1F axis represents a promising strategy for improving the efficacy of FINs or chemotherapy in the treatment of LUAD patients.
Subject(s)
Adenocarcinoma of Lung , Ferroptosis , Lung Neoplasms , Thioredoxins , Ferroptosis/drug effects , Ferroptosis/genetics , Humans , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/drug therapy , Adenocarcinoma of Lung/pathology , Adenocarcinoma of Lung/metabolism , Thioredoxins/metabolism , Thioredoxins/genetics , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Animals , Mice , Cell Line, Tumor , Gene Expression Regulation, Neoplastic/drug effects , Mice, Nude , Female , MaleABSTRACT
EGF receptor (EGFR) tyrosine kinase inhibitors (TKIs) have achieved clinical success in lung adenocarcinoma (LUAD). However, tumors often show profound but transient initial response and then gain resistance. We identify transcription factor ZNF263 as being significantly decreased in osimertinib-resistant or drug-tolerant persister LUAD cells and clinical residual tumors. ZNF263 overexpression improves the initial response of cells and delays the formation of persister cells with osimertinib treatment. We further show that ZNF263 binds and recruits DNMT1 to the EGFR gene promoter, suppressing EGFR transcription with DNA hypermethylation. ZNF263 interacts with nuclear EGFR, impairing the EGFR-STAT5 interaction to enhance AURKA expression. Overexpressing ZNF263 also makes tumor cells with wild-type EGFR expression or refractory EGFR mutations more susceptible to EGFR inhibition. More importantly, lentivirus or adeno-associated virus (AAV)-mediated ZNF263 overexpression synergistically suppresses tumor growth and regrowth with osimertinib treatment in xenograft animal models. These findings suggest that enhancing ZNF263 may achieve complete response in LUAD with EGFR-targeted therapies.
Subject(s)
Acrylamides , Adenocarcinoma of Lung , Aniline Compounds , Indoles , Lung Neoplasms , Pyrimidines , Animals , Humans , Transcription Factors/genetics , Neoplasm, Residual , Adenocarcinoma of Lung/drug therapy , Adenocarcinoma of Lung/genetics , ErbB Receptors/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , DNA-Binding ProteinsABSTRACT
BACKGROUND: Lung cancer is one of the most common tumors in the world, and metastasis is one of the major causes of tumor-related death in lung cancer patients. Tumor-associated macrophages (TAMs) are a major component of the tumor microenvironment (TME) and are frequently associated with tumor metastasis in human cancers. However, the regulatory mechanisms of TAMs in lung cancer metastasis remain unclear. METHODS: Single-cell sequencing analysis of lung cancer and normal tissues from public databases and from 14 patients who underwent surgery at Zhongshan Hospital was performed. In vitro co-culture experiments were performed to evaluate the effects of TAMs on lung cancer migration and invasion. Changes in the expression of IL-6, STAT3, C/EBPΒ, and EMT pathway were verified using RT-qPCR, western blotting, and immunofluorescence. Dual luciferase reporter assays and ChIP were used to reveal potential regulatory sites on the transcription factor sets. In addition, the effects of TAMs on lung cancer progression and metastasis were confirmed by in vivo models. RESULTS: TAM infiltration is associated with tumor progression and poor prognosis. IL-6 secreted by TAMs can activate the JAK2/STAT3 pathway through autocrine secretion, and STAT3 acts as a transcription factor to activate the expression of C/EBPß, which further promotes the transcription and expression of IL-6, forming positive feedback loops for IL6-STAT3-C/EBPß-IL6 in TAMs. IL-6 secreted by TAMs promotes lung cancer progression and metastasis in vivo and in vitro by activating the EMT pathway, which can be attenuated by the use of JAK2/STAT3 pathway inhibitors or IL-6 monoclonal antibodies. CONCLUSIONS: Our data suggest that TAMs promote IL-6 expression by forming an IL6-STAT3-C/EBPß-IL6 positive feedback loop. Released IL-6 can induce the EMT pathway in lung cancer to enhance migration, invasion, and metastasis. The use of IL-6-neutralizing antibody can partially counteract the promotion of LUAD by TAMs. A novel mechanism of macrophage-promoted tumor progression was revealed, and the IL6-STAT3-C/EBPß-IL6 signaling cascade may be a potential therapeutic target against lung cancer.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Humans , Interleukin-6/metabolism , Tumor-Associated Macrophages/metabolism , Cell Line, Tumor , Feedback , Adenocarcinoma of Lung/genetics , Lung Neoplasms/genetics , Transcription Factors/metabolism , STAT3 Transcription Factor/genetics , STAT3 Transcription Factor/metabolism , Tumor Microenvironment , Epithelial-Mesenchymal TransitionABSTRACT
AIMS: Lung cancer is the leading cause of cancer mortality and lung adenocarcinoma (LUAD) accounts for more than half of all lung cancer cases. Tumor elimination is mostly hindered by drug resistance and the mechanisms remain to be explored in LUAD. METHODS: CRISPR screens in cell and murine models and single-cell RNA sequencing were conducted, which identified MAF bZIP transcription factor F (MAFF) as a critical factor regulating tumor growth and treatment resistance in LUAD. RNA and ChIP sequencing analyses were performed for transcriptional target expression and specific binding sites of MAFF. Functions of MAFF in inhibiting tumor growth and promoting cisplatin or irradiation efficacy were investigated using cellular and xenograft models. RESULTS: Patients with lung adenocarcinoma and reduced MAFF expression had worse clinical outcomes. MAFF inhibited tumor cell proliferation by regulating the expression of SLC7A11, CDK6, and CDKN2C, promoting ferroptosis and preventing cell cycle progression from G1 to S. MAFF also conferred tumor cells vulnerable to cisplatin-based or ionizing radiation treatments. MAFF reduction was a final event in the acquisition of cisplatin resistance of LUAD cells. The intracellular cAMP/PKA/CREB1 pathway upregulated MAFF in response to cisplatin-based or ionizing radiation treatments. CONCLUSIONS: MAFF suppresses tumor growth, and pharmacological agonists targeting MAFF may improve cisplatin or irradiation therapies for lung adenocarcinoma patients.
Subject(s)
Adenocarcinoma of Lung , Ferroptosis , Lung Neoplasms , Humans , Animals , Mice , Cisplatin/pharmacology , Cisplatin/therapeutic use , Ferroptosis/genetics , Cell Line, Tumor , Adenocarcinoma of Lung/drug therapy , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/radiotherapy , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/radiotherapy , Cell Proliferation , Cell Cycle , Nuclear Proteins/metabolism , Nuclear Proteins/therapeutic use , MafF Transcription FactorABSTRACT
Metformin's effect on tumor treatment was complex, because it significantly reduced cancer cell proliferation in vitro, but made no difference in prognosis in several clinical cohorts. Our transcriptome sequencing results revealed that tumor-associated macrophage (TAM) infiltration significantly increased in active lung adenocarcinoma (LUAD) patients with long-term metformin use. We further identified that the tumor suppressive effect of metformin was more significant in mice after the depletion of macrophages, suggesting that TAMs might play an important role in metformin's effects in LUAD. Combining 10X Genomics single-cell sequencing of tumor samples, transcriptome sequencing of metformin-treated TAMs, and the ChIP-Seq data of the Encode database, we identified and validated that metformin significantly increased the expression and secretion of S100A9 of TAMs through AMPK-CEBP/ß pathway. For the downstream, S100A9 binds to RAGE receptors on the surface of LUAD cells, and then activates the NF-κB pathway to promote EMT and progression of LUAD, counteracting the inhibitory effect of metformin on LUAD cells. In cell-derived xenograft models (CDX) and patient-derived xenograft models (PDX) models, our results showed that neutralizing antibodies targeting TAM-secreted S100A9 effectively enhanced the tumor suppressive effect of metformin in treating LUAD. Our results will enable us to better comprehend the complex role of metformin in LUAD, and advance its clinical application in cancer treatment.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Metformin , Animals , Humans , Mice , Adenocarcinoma of Lung/drug therapy , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/metabolism , Calgranulin B/genetics , Disease Models, Animal , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/metabolism , Metformin/pharmacology , Metformin/therapeutic use , Prognosis , Tumor-Associated Macrophages/metabolismABSTRACT
Background: N6-methyladenosine (m6A) RNA methylation plays a crucial role in important genomic processes in a variety of malignancies. However, the characterization of m6A with infiltrating immune cells in the tumor microenvironment (TME) in esophageal squamous carcinoma (ESCC) remains unknown. Methods: The single-cell transcriptome data from five ESCC patients in our hospital were analyzed, and TME clusters associated with prognosis and immune checkpoint genes were investigated. Cell isolation and qPCR were conducted to validate the gene characterization in different cells. Results: According to distinct biological processes and marker genes, macrophages, T cells, and B cells clustered into three to four different subgroups. In addition, we demonstrated that m6A RNA methylation regulators were strongly related to the clinical and biological features of ESCC. Analysis of transcriptome data revealed that m6A-mediated TME cell subsets had high predictive value and showed a close relationship with immune checkpoint genes. The validation results from qPCR demonstrated the characteristics of essential genes. CellChat analysis revealed that RNA from TME cells m6A methylation-associated cell subtypes had substantial and diversified interactions with cancer cells. Further investigation revealed that MIF- (CD74+CXCR4) and MIF- (CD74+CD44) ligand-receptor pairings facilitated communication between m6A-associated subtypes of TME cells and cancer cells. Conclusion: Overall, our study demonstrated for the first time the function of m6A methylation-mediated intercellular communication in the microenvironment of tumors in controlling tumor development and anti-tumor immune regulation in ESCC.
ABSTRACT
BACKGROUND: Neoadjuvant chemotherapy (NACT) becomes the first-line option for advanced tumors, while patients who are not sensitive to it may not benefit. Therefore, it is important to screen patients suitable for NACT. METHODS: Single-cell data of lung adenocarcinoma (LUAD) and esophageal squamous carcinoma (ESCC) before and after cisplatin-containing (CDDP) NACT and cisplatin IC50 data of tumor cell lines were analyzed to establish a CDDP neoadjuvant chemotherapy score (NCS). Differential analysis, GO, KEGG, GSVA and logistic regression models were performed by R. Survival analysis were applied to public databases. siRNA knockdown in A549, PC9, TE1 cell lines, qRT-PCR, western-blot, cck8 and EdU experiments were used for further verification in vitro. RESULTS: 485 genes were expressed differentially in tumor cells before and after neoadjuvant treatment for LUAD and ESCC. After combining the CDDP-associated genes, 12 genes, CAV2, PHLDA1, DUSP23, VDAC3, DSG2, SPINT2, SPATS2L, IGFBP3, CD9, ALCAM, PRSS23, PERP, were obtained and formed the NCS score. The higher the score, the more sensitive the patients were to CDDP-NACT. The NCS divided LUAD and ESCC into two groups. Based on differentially expressed genes, a model was constructed to predict the high and low NCS. CAV2, PHLDA1, ALCAM, CD9, IGBP3 and VDAC3 were significantly associated with prognosis. Finally, we demonstrated that the knockdown of CAV2, PHLDA1 and VDAC3 in A549, PC9 and TE1 significantly increased the sensitivity to cisplatin. CONCLUSIONS: NCS scores and related predictive models for CDDP-NACT were developed and validated to assist in selecting patients who might benefit from it.
ABSTRACT
PURPOSE: The tumor immune microenvironment (TME) plays a vital role in tumorigenesis, progression, and treatment. Macrophages, as an important component of the tumor microenvironment, play an essential role in antitumor immunity and TME remodeling. In this study, we aimed to explore the different functions of different origins macrophages in TME and their value as potential predictive markers of prognosis and treatment. METHODS: We performed single-cell analysis using 21 lung adenocarcinoma (LUAD), 12 normal, and four peripheral blood samples from our data and public databases. A prognostic prediction model was then constructed using 502 TCGA patients and explored the potential factors affecting prognosis. The model was validated using data from 4 different GEO datasets with 544 patients after integration. RESULTS: According to the source of macrophages, we classified macrophages into alveolar macrophages (AMs) and interstitial macrophages (IMs). AMs mainly infiltrated in normal lung tissue and expressed proliferative, antigen-presenting, scavenger receptors genes, while IMs occupied the majority in TME and expressed anti-inflammatory, lipid metabolism-related genes. Trajectory analysis revealed that AMs rely on self-renew, whereas IMs originated from monocytes in the blood. Cell-to-cell communication showed that AMs interacted mainly with T cells through the MHC I/II signaling pathway, while IMs mostly interacted with tumor-associated fibrocytes and tumor cells. We then constructed a risk model based on macrophage infiltration and showed an excellent predictive power. We further revealed the possible reasons for its potential prognosis prediction by differential genes, immune cell infiltration, and mutational differences. CONCLUSION: In conclusion, we investigated the composition, expression differences, and phenotypic changes of macrophages from different origins in lung adenocarcinoma. In addition, we developed a prognostic prediction model based on different macrophage subtype infiltration, which can be used as a valid prognostic biomarker. New insights were provided into the role of macrophages in the prognosis and potential treatment of LUAD patients.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Humans , Transcriptome/genetics , Macrophages , Adenocarcinoma of Lung/genetics , Monocytes , Lung Neoplasms/genetics , Tumor Microenvironment/geneticsABSTRACT
Background: Lung adenocarcinoma is one of the most common tumors, and cisplatin is frequently used in treating lung adenocarcinoma patients. This study aimed to look into the roles and mechanisms of HNF4G in cisplatin resistance of lung adenocarcinoma. Materials & Methods: Cisplatin resistance and gene expression data of 542 cell lines from the CTRP and CCLE databases were analyzed. HNF4G expression was detected in the lung adenocarcinoma cell lines after treatment with various concentrations of cisplatin. Cisplatin sensitivity curves were detected in cells that overexpressed or knocked down HNF4G. The ChIP-Seq data were then analyzed to identify the targets of HNF4G involved in cisplatin resistance. Expression and phosphorylation of the MAPK6/Akt pathway were detected after HNF4G was overexpressed or knocked down. Finally, ChIP-qPCR and dual-luciferase assays were used to investigate the regulation of HNF4G on MAPK6. Results: In cell lines, high expression of HNF4G was significantly positively correlated with cisplatin resistance, and lung adenocarcinoma patients who had high HNF4G expression had a poor prognosis. Cisplatin treatment increased HNF4G expression, and overexpression of HNF4G significantly increased the resistance to cisplatin in A549 and HCC827 cells, whereas knockdown of HNF4G had the opposite effect. HNF4G overexpression increased MAPK6 expression and activated the MAPK6/Akt pathway, while an Akt inhibitor reduced the effects of HNF4G on cisplatin resistance. HNF4G bound to the MAPK6 promoter region, promoting MAPK6 expression, according to ChIP-qPCR and luciferase assays. Conclusion: By binding to the MAPK6 promoter region, HNF4G promotes MAPK6 expression and subsequent Akt phosphorylation, resulting in resistance to cisplatin in lung adenocarcinoma.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Humans , Cisplatin/pharmacology , Proto-Oncogene Proteins c-akt/metabolism , Lung Neoplasms/drug therapy , Signal Transduction , Cell Line, Tumor , Drug Resistance, Neoplasm/genetics , Adenocarcinoma of Lung/drug therapy , Hepatocyte Nuclear Factor 4/geneticsABSTRACT
Background: This study identified the expression and prognosis significance of secretory or membrane-associated proteins in KRAS lung adenocarcinoma (LUAD) and depicted the characteristics between the immune cell infiltration and the expression of these genes. Methods: Gene expression data of LUAD samples (n = 563) were accessed from The Cancer Genome Atlas (TCGA). The expression of secretory or membrane-associated proteins was compared among the KRAS-mutant, wild-type, and normal groups, as well as the subgroup of the KRAS-mutant group. We identified the survival-related differentially expressed secretory or membrane-associated proteins and conducted the functional enrichment analysis. Then, the characterization and association between their expression and the 24 immune cell subsets were investigated. We also constructed a scoring model to predict KRAS mutation by LASSO and logistic regression analysis. Results: Secretory or membrane-associated genes with differential expression (n = 74) across three groups (137 KRAS LUAD, 368 wild-type LUAD, and 58 normal groups) were identified, and the results of GO and KEGG indicated that they were strongly associated with immune cell infiltrations. Among them, ten genes were significantly related to the survival of patients with KRAS LUAD. The expression of IL37, KIF2, INSR, and AQP3 had the most significant correlations with immune cell infiltration. In addition, eight DEGs from the KRAS subgroups were highly correlated with immune infiltrations, especially TNFSF13B. Using LASSO-logistic regression, a KRAS mutation prediction model based on the 74 differentially expressed secretory or membrane-associated genes was built, and the accuracy was 0.79. Conclusion: The research investigated the relationship between the expression of KRAS-related secretory or membrane-associated proteins in LUAD patients with prognostic prediction and immune infiltration characterization. Our study demonstrated that secretory or membrane-associated genes were closely associated with the survival of KRAS LUAD patients and were strongly correlated to immune cell infiltration.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Humans , Proto-Oncogene Proteins p21(ras)/genetics , Adenocarcinoma of Lung/genetics , Biological Transport , Mutation , Lung Neoplasms/genetics , Prognosis , Interleukin-1ABSTRACT
Studies have shown that tumors with ground-glass opacity (GGO) components are associated with favorable outcomes. However, this view should be confirmed in an international cohort. We aimed to verify the impact of a GGO component on clinical (c)-stage IA lung adenocarcinoma and to describe the biological discrepancies between the part-solid and pure-solid groups. We evaluated 1333 cases of surgically resected c-stage IA lung adenocarcinomas, including 484 part-solid and 849 pure-solid tumors. Furthermore, we matched the solid size between the 2 groups and examined 470 patients. We compared the prognoses between the 2 groups before and after matching. The prognostic and biological differences were described before and after matching. Compared with the pure-solid group, the part-solid group was associated with favorable outcomes [5-year overall survival (OS) 99.4% vs 87.6%, P < 0.001; 5-year recurrence-free survival (RFS) 96.9% vs 82.2%, P < 0.001]. Similar results were obtained after matching (5-year OS 98.9% vs 92.2%, P = 0.012; 5-year RFS 95.0% vs 88.5%, P = 0.007). Multivariable analyses revealed that GGO component appearance was a factor of better OS and RFS. The part-solid tumor, regardless of the size of the solid component, had a similar outcome to the pure-solid tumor of c-stage T1a classification. Also, more epidermal growth factor receptor, human epidermal growth factor receptor-2 mutations, and receptor tyrosine kinase ROS-1-positive were observed in the part-solid group. In comparison, more wild types and Kirsten-Ras were observed in the pure-solid group. Adenocarcinomas with a GGO component were associated with superior outcomes. The GGO component should be considereda new clinical T descriptor. Early-stage lung adenocarcinomas with and without a GGO component may be 2 distinct tumor types.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Humans , Lung Neoplasms/diagnostic imaging , Lung Neoplasms/surgery , Neoplasm Staging , Treatment Outcome , Retrospective Studies , Adenocarcinoma of Lung/diagnostic imaging , Adenocarcinoma of Lung/surgery , Prognosis , ErbB ReceptorsABSTRACT
INTRODUCTION: Surgery is the most effective treatment for early-stage lung cancer. This study will propose a personalized plan for mediastinal lymph node dissection in early-stage lung adenocarcinoma to reduce the risk of surgery and improve the quality of life. METHODS: This study retrospectively analyzed the patients underwent lobectomy and lymph node dissection in the Department of Thoracic Surgery, Zhongshan Hospital, Fudan University. Clinical stage I lung adenocarcinoma patients with solid component ratio (CTR) between 0.5 and 1 were included. Patients were divided into systematic (S-MLND) and lobe-specific (L-MLND) mediastinal lymph node dissection groups. The days of hospitalization, the presence or absence of complications, the recurrence-free survival rate, and the overall survival rate were calculated to evaluate the postoperative quality and operation risk of the patients. RESULTS: 210 patients (138 L-MLND and 72 S-MLND) were included. 2 lymph node metastases appeared in the S-MLND group while none in the L-MLND group (P = .049). No differences were shown in age, tumor site, size, solid component, degree of tumor invasion, and stage. The proportion of patients with severe postoperative cough and the length of hospital stay in the L-MLND group decreased. The 5-year OS of the entire cohort was 98.1%, 98.6% in L-MLND, compared with 97.2% in S-MLND; RFS was 94.8%, 95.7% in L-MLND, compared with 93.0% in S-MLND. CONCLUSION: For cIA lung adenocarcinoma, according to the Thin-slice CT within 1 month before the operation, if the main lesion was less than 3 cm and CTR over 0.5, L-MLND is as effective as S-MLND.
Subject(s)
Adenocarcinoma of Lung , Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Humans , Carcinoma, Non-Small-Cell Lung/pathology , Lung Neoplasms/pathology , Retrospective Studies , Quality of Life , Neoplasm Staging , Lymph Node Excision , Adenocarcinoma of Lung/pathology , Lymph Nodes/pathology , PneumonectomyABSTRACT
Background: The mitochondrion and its associated genes were heavily implicated in developing and therapy tumors as the primary cellular organelle in charge of metabolic reprogramming and ferroptosis. Our work focuses on discovering new potential targets while analyzing the multi-omics data of mitochondria-related genes in lung adenocarcinoma (LUAD). Methods: The Cancer Genome Atlas (TCGA) database provided multi-omics data for LUAD patients. Based on the expression profile of the genes associated with mitochondria, the patients were grouped by the unsupervised clustering method. R was used to explore the differential expressed protein-code gene, miRNA, and lncRNA, as well as their enriched functions and ceRNA networks. Additionally, the discrepancy between immune infiltration and genetic variation was comprehensively characterized. Our clinical samples and in vitro experiments investigated the hub gene determined by LASSO and batch analysis. Results: Two clusters are distinguished using unsupervised consensus clustering based on mitochondrial heterogeneity. The integrated analysis emphasized that patients in cluster B had a worse prognosis, higher mutation frequencies, and less immune cell infiltration. The hub genes DARS2 and COX5B are identified by further analysis using LASSO penalization. In vitro experiments indicated that DARS2 and COX5B knockdown inhibited tumor cell proliferation. The specimen of our hospital cohort conducted the immunohistochemistry analysis and validated that DARS2 and COX5B's expression was significantly higher in the tumor than in adjacent normal tissue and correlated to LUAD patients' prognosis. Conclusion: Our observations implied that LUAD patients' tumors had distinct mitochondrial function heterogeneity with different clinical and molecular characteristics. DARS2 and COX5B might be critical genes involved in mitochondrial alterations and potential therapeutic targets.
Subject(s)
DNA Copy Number Variations , Lung Neoplasms , B7-H1 Antigen , Humans , Polymerase Chain Reaction , RNA, MessengerABSTRACT
Background: The tumor immune microenvironment (TIME) played an important role in immunotherapy prognosis and treatment response. Immune cells constitute a large part of the tumor microenvironment and regulate tumor progression. Our research is dedicated to studying the infiltrating immune cell in lung adenocarcinoma (LUAD) and seeking potential targets. Methods: The scRNA-seq data were collected from our FDZSH and two public datasets. The code for cell-type mapping algorithms was downloaded from the CIBERSORTx portal. The bioinformatics data of LUAD patients could be approached from The Cancer Genome Atlas (TCGA) portal. Weighted gene coexpression network analysis (WGCNA) and least absolute shrinkage and selection operator (LASSO) analyses were performed to construct a risk model. TIMER2 and TIDE helped with the immune infiltration estimation, while PROGENy helped the cancer-related pathways' enrichment analysis. GSE31210 dataset and IMVigor ICB therapy cohort validated our findings as the external validation datasets. Results: We clustered the scRNA-seq dataset (integrating our FDZSH datasets and other public datasets) into 23 subpopulations. After curated cell annotation, we implemented Cibersort and WGCNA analysis to anchor the brown module and natural killer cell cluster1 due to the most relationship with tumor trait. The overlap of the brown module gene, natural killer cell signature, and DEGs of tumor and adjacent normal samples was screened by LASSO Cox regression. The obtained 5-gene risk model showed an excellent prognostic performance in the validation dataset. Furthermore, there was a correlation between risk score and tumor-infiltrating immune cells and tumor genomics abnormity. Patients with higher risk scores had a significantly lower immunotherapy response rate. Conclusion: Our observations implied that immune cells played a pivotal role in TIME and established a 5-gene signature (including IDH2, ADRB2, SFTPC, CCDC69, and CCND2) on the basement of nature killer markers targeted by WGCNA analysis. The significance of clinical outcome and immunotherapy response prediction was validated robustly.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Biomarkers, Tumor/genetics , Humans , Kaplan-Meier Estimate , Lung Neoplasms/genetics , Prognosis , Sequence Analysis, RNA , Tumor Microenvironment/geneticsABSTRACT
OBJECTIVES: Platinum-based chemotherapies are currently the first-line treatment of non-small cell lung cancer. This study will improve our understanding of the causes of resistance to cisplatin, especially in lung adenocarcinoma (LUAD) and provide a reference for therapeutic decisions in clinical practice. METHODS: Cancer Cell Line Encyclopedia (CCLE), The Cancer Genome Atlas (TCGA) and Zhongshan hospital affiliated to Fudan University (zs-cohort) were used to identify the multi-omics differences related to platinum chemotherapy. Cisplatin-resistant mRNA and miRNA models were constructed by Logistic regression, classification and regression tree and C4.5 decision tree classification algorithm with previous feature selection performed via least absolute shrinkage and selection operator (LASSO). qRT-PCR and western-blotting of A549 and H358 cells, as well as single-cell Seq data of tumor samples were applied to verify the tendency of certain genes. RESULTS: 661 cell lines were divided into three groups according to the IC50 value of cisplatin, and the top 1/3 (220) with a small IC50 value were defined as the sensitive group while the last 1/3 (220) were enrolled in the insensitive group. TP53 was the most common mutation in the insensitive group, in contrast to TTN in the sensitive group. 1348 mRNA, 80 miRNA, and 15 metabolites were differentially expressed between 2 groups (P < 0.05). According to the LASSO penalized logistic modeling, 6 of the 1348 mRNAs, FOXA2, BATF3, SIX1, HOXA1, ZBTB38, IRF5, were selected as the associated features with cisplatin resistance and for the contribution of predictive mRNA model (all of adjusted P-values < 0.001). Three of 6 (BATF3, IRF5, ZBTB38) genes were finally verified in cell level and patients in zs-cohort. CONCLUSIONS: Somatic mutations, mRNA expressions, miRNA expressions, metabolites and methylation were related to the resistance of cisplatin. The models we created could help in the prediction of the reaction and prognosis of patients given platinum-based chemotherapies.
Subject(s)
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , MicroRNAs , Carcinoma, Non-Small-Cell Lung/drug therapy , Cisplatin/pharmacology , Cisplatin/therapeutic use , Homeodomain Proteins , Humans , Interferon Regulatory Factors , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Machine Learning , MicroRNAs/genetics , RNA, Messenger/genetics , RNA, Messenger/therapeutic useABSTRACT
Background: The role of ferroptosis in esophageal squamous cell carcinoma (ESCC) is still unclear. Methods: The association of iron metabolism and ferroptosis-related genes with the prognosis, copy number variation (CNV), TMB, and immune cell infiltration of ESCC was explored using data from the GEO and TCGA database and validated by immunofluorescence in 112 ESCC patients from our center. The potential anti-cancer drugs and compounds from the GDSC and the Connectivity Map database were also screened. Results: A total of 117 iron metabolism and ferroptosis-related genes were identified. We found the expressions of PRNP, SLC3A2, SLC39A8, and SLC39A14 negatively related to the prognosis of ESCC patients, while ATP6V0A1 and LCN2 were opposite, which was validated in 112 ESCC samples from our center. And a prognostic signature was constructed based on their expressions and Cox regression coefficient (ß). The low-score group exhibited a significantly worse OS. Besides, analysis of 179 ESCC samples from GSE53625 revealed that patients of poorly differentiation, more than 60 years, T4 stage, advanced N stage, advanced stage, and adjuvant therapy also exhibited a significantly shorter OS, based on which a nomogram to predict the OS was established. Moreover, the low-score group exhibited significantly higher CNV and TMB and more frequent mutations of TP53, MUC16, and NOTCH1. Higher proportion of Macrophages M2, and lower proportion of T cells follicular helper were observed in the low-score group. We discovered that AZD7762, Sunitinib, Cytarabine, Docetaxel, Vinblastine, and Elesclomol exhibited lower IC50 in the low-score group. And 20 potential compounds were identified from the CMap database. Conclusions: Six iron metabolism and ferroptosis-related genes were associated with the prognosis, CNV, TMB, and immune cell infiltration of ESCC. Some potential anti-cancer drugs and compounds may be helpful for OS.
ABSTRACT
BACKGROUND: Single-cell transcriptomics has been used to investigate various tumors to elucidate the molecular distinction of all cell type compositions of a complex mix. AIMS: This study aimed to investigate malignant-cell-specific genes to explore diagnostic and therapeutic biomarkers using single-cell transcriptomic data of lung adenocarcinoma. MATERIALS & METHODS: 10X single-cell RNA-seq data of fourteen patients with lung adenocarcinoma were analyzed. Genes that expressed differentially and those with higher confidence to distinguish tumor cells from normal cells were picked out using the ROC curves. The LASSO regression method was used to select most markedly correlated genes to predict the malignancy of every single cell within a model. We also conducted further experiments to determine their roles in lung cancer in vitro. RESULTS: Twenty two thousand four hundred and ninety one tumor and 181 666 normal single cells were analyzed where 369 genes were found to be specifically expressed in single malignant cells. Seventy of them, encoding secreted or membrane-bound proteins, showed involvement in cell-to-cell communications in tumor biology. KRT18 and the other six genes were identified as predictors to distinguish single malignant cells and were integrated to construct an accurate (96.1%) predicting model. Notably, IRX2, SPINK13, and CAPN8 outperformed the other four genes. Further experiments confirmed the upregulation of them in lung adenocarcinoma at both tissue and cell levels. Proliferative capacities of lung adenocarcinoma cells were attenuated by knocking-down of either of them. However, targeting CAPN8, IRX2, or SPINK13 hardly exerted a cytotoxic effect on these cells. DISCUSSION: Apart from the current model, similar tools were still warranted using single-cell RNA-seq data of more types of tumors. The three genes identified as potential therapeutic targets in the present study still need to be validated in more in lung cancer. CONCLUSION: Our model can aid the analyses of single-cell sequencing data. CAPN8, IRX2, and SPINK13 may serve as novel targets of targeted and immune-based therapies in lung adenocarcinoma.