ABSTRACT
Esophageal squamous-cell carcinoma (ESCC) is commonly treated with radiotherapy; however, radioresistance hinders its clinical effectiveness, and the underlying mechanism remains elusive. Here, we develop patient-derived xenografts (PDXs) from 19 patients with ESCC to investigate the mechanisms driving radioresistance. Using RNA sequencing, cytokine arrays, and single-cell RNA sequencing, we reveal an enrichment of cancer-associated fibroblast (CAF)-derived collagen type 1 (Col1) and tumor-cell-derived CXCL1 in non-responsive PDXs. Col1 not only promotes radioresistance by augmenting DNA repair capacity but also induces CXCL1 secretion in tumor cells. Additionally, CXCL1 further activates CAFs via the CXCR2-STAT3 pathway, establishing a positive feedback loop. Directly interfering with tumor-cell-derived CXCL1 or inhibiting the CXCL1-CXCR2 pathway effectively restores the radiosensitivity of radioresistant xenografts in vivo. Collectively, our study provides a comprehensive understanding of the molecular mechanisms underlying radioresistance and identifies potential targets to improve the efficacy of radiotherapy for ESCC.
Subject(s)
Cancer-Associated Fibroblasts , Carcinoma, Squamous Cell , Esophageal Neoplasms , Esophageal Squamous Cell Carcinoma , Radiation Tolerance , Humans , Cancer-Associated Fibroblasts/metabolism , Cancer-Associated Fibroblasts/radiation effects , Carcinoma, Squamous Cell/genetics , Carcinoma, Squamous Cell/radiotherapy , Carcinoma, Squamous Cell/metabolism , Cell Line, Tumor , Chemokine CXCL1/metabolism , Collagen/metabolism , Esophageal Neoplasms/genetics , Esophageal Neoplasms/radiotherapy , Esophageal Neoplasms/metabolism , Esophageal Squamous Cell Carcinoma/radiotherapy , Esophageal Squamous Cell Carcinoma/genetics , Esophageal Squamous Cell Carcinoma/metabolismABSTRACT
OBJECTIVES: The disposable esophagogastroduodenoscopy (EGD) system is a novel endoscopic device which is highly portable and is designed to eliminate the risk of cross-infection caused by reusable EGD. This study aimed to investigate the feasibility and safety of disposable EGD in emergency, bedside, and intraoperative settings. METHODS: This was a prospective, single-center, noncomparative study. Disposable EGD was used for emergency, bedside, and intraoperative endoscopies in 30 patients. The primary end-point was the technical success rate of the disposable EGD. Secondary end-points included technical performance indicators including clinical operability, image quality score, procedure time, the incidence of device malfunction and/or failure, and the incidence of adverse events. RESULTS: A total of 30 patients underwent diagnosis and/or treatment with disposable EGD. Therapeutic EGD was performed on 13/30 patients, including hemostasis (n = 3), foreign body retrieval (n = 6), nasoenteric tube placement (n = 3), and percutaneous endoscopic gastrostomy (n = 1). The technical success rate was 100%: all procedures and indicated interventions were completed without changing to a conventional upper endoscope. The mean image quality score obtained immediately after procedure completion was 3.72 ± 0.56. The mean (± SD) procedure time was 7.4 (± 7.6) min. There were no device malfunctions or failures, device-related adverse events, or overall adverse events. CONCLUSION: The disposable EGD may be a feasible alternative to the traditional EGD in emergency, bedside, and intraoperative settings. Preliminary data show that it is a safe and effective tool for diagnosis and treatment in emergency and bedside upper gastrointestinal cases. TRIAL REGISTRATION: Chinese Clinical Trial Registry (Trial ID: ChiCTR2100051452, https://www.chictr.org.cn/showprojen.aspx?proj=134284).
Subject(s)
Endoscopy, Digestive System , Endoscopy , Humans , Pilot Projects , Prospective Studies , Endoscopy, Digestive System/methods , Intubation, GastrointestinalABSTRACT
Esophageal squamous-cell carcinoma (ESCC) develops through multistage epithelial cancer formation, i.e., from normal epithelium, low- and high-grade intraepithelial neoplasia to invasive carcinoma. However, how the precancerous lesions progress to carcinoma remains elusive. Here, we report a comprehensive single-cell RNA sequencing and spatial transcriptomic study of 79 multistage esophageal lesions from 29 patients with ESCC. We reveal a gradual and significant loss of ANXA1 expression in epithelial cells due to its transcription factor KLF4 suppression along the lesion progression. We demonstrate that ANXA1 is a ligand to formyl peptide receptor type 2 (FPR2) on fibroblasts that maintain fibroblast homeostasis. Loss of ANXA1 leads to uncontrolled transformation of normal fibroblasts into cancer-associated fibroblasts (CAFs), which can be enhanced by secreted TGF-ß from malignant epithelial cells. Given the role of CAFs in cancer, our study underscores ANXA1/FPR2 signaling as an important crosstalk mechanism between epithelial cells and fibroblasts in promoting ESCC.
Subject(s)
Carcinoma in Situ , Esophageal Neoplasms , Precancerous Conditions , Humans , Esophageal Neoplasms/genetics , Epithelial Cells , FibroblastsABSTRACT
Objectives: Adenocarcinoma at the gastroesophageal junction (ACGEJ) refers to a malignant tumor that occurs at the esophagogastric junction. Despite some progress in targeted therapies for HER2, FGFR2, EGFR, MET, Claudin 18.2 and immune checkpoints in ACGEJ tumors, the 5-year survival rate of patients remains poor. Thus, it is urgent to explore genomic alterations and neoantigen characteristics of tumors and identify CD8+ T-cell infiltration-associated genes to find potential therapeutic targets and develop a risk model to predict ACGEJ patients' overall survival (OS). Methods: Whole-exome sequencing (WES) was performed on 55 paired samples from Chinese ACGEJ patients. Somatic mutations and copy number variations were detected by Strelka2 and FACETS, respectively. SigProfiler and SciClone were employed to decipher the mutation signature and clonal structure of each sample, respectively. Neoantigens were predicted using the MuPeXI pipeline. RNA sequencing (RNA-seq) data of ACGEJ samples from our previous studies and The Cancer Genome Atlas (TCGA) were used to identify genes significantly associated with CD8+ T-cell infiltration by weighted gene coexpression network analysis (WGCNA). To construct a risk model, we conducted LASSO and univariate and multivariate Cox regression analyses. Results: Recurrent MAP2K7, RNF43 and RHOA mutations were found in ACGEJ tumors. The COSMIC signature SBS17 was associated with ACGEJ progression. CCNE1 and VEGFA were identified as putative CNV driver genes. PI3KCA and TP53 mutations conferred selective advantages to cancer cells. The Chinese ACGEJ patient neoantigen landscape was revealed for the first time, and 58 potential neoantigens common to TSNAdb and IEDB were identified. Compared with Siewert type II samples, Siewert type III samples had significant enrichment of the SBS17 signature, a lower TNFRSF14 copy number, a higher proportion of samples with complex clonal architecture and a higher neoantigen load. We identified 10 important CD8+ T-cell infiltration-related Hub genes (CCL5, CD2, CST7, GVINP1, GZMK, IL2RB, IKZF3, PLA2G2D, P2RY10 and ZAP70) as potential therapeutic targets from the RNA-seq data. Seven CD8+ T-cell infiltration-related genes (ADAM28, ASPH, CAMK2N1, F2R, STAP1, TP53INP2, ZC3H3) were selected to construct a prognostic model. Patients classified as high risk based on this model had significantly worse OS than low-risk patients, which was replicated in the TCGA-ACGEJ cohort. Conclusions: This study provides new neoantigen-based immunotherapeutic targets for ACGEJ treatment and effective disease prognosis biomarkers.
ABSTRACT
Evidence points toward the differentiation state of cells as a marker of cancer risk and progression. Measuring the differentiation state of single cells in a preneoplastic population could thus enable novel strategies for early detection and risk prediction. Recent maps of somatic mutagenesis in normal tissues from young healthy individuals have revealed cancer driver mutations, indicating that these do not correlate well with differentiation state and that other molecular events also contribute to cancer development. We hypothesized that the differentiation state of single cells can be measured by estimating the regulatory activity of the transcription factors (TF) that control differentiation within that cell lineage. To this end, we present a novel computational method called CancerStemID that estimates a stemness index of cells from single-cell RNA sequencing data. CancerStemID is validated in two human esophageal squamous cell carcinoma (ESCC) cohorts, demonstrating how it can identify undifferentiated preneoplastic cells whose transcriptomic state is overrepresented in invasive cancer. Spatial transcriptomics and whole-genome bisulfite sequencing demonstrated that differentiation activity of tissue-specific TFs was decreased in cancer cells compared with the basal cell-of-origin layer and established that differentiation state correlated with differential DNA methylation at the promoters of these TFs, independently of underlying NOTCH1 and TP53 mutations. The findings were replicated in a mouse model of ESCC development, and the broad applicability of CancerStemID to other cancer-types was demonstrated. In summary, these data support an epigenetic stem-cell model of oncogenesis and highlight a novel computational strategy to identify stem-like preneoplastic cells that undergo positive selection. SIGNIFICANCE: This study develops a computational strategy to dissect the heterogeneity of differentiation states within a preneoplastic cell population, allowing identification of stem-like cells that may drive cancer progression.
Subject(s)
Esophageal Neoplasms , Esophageal Squamous Cell Carcinoma , Animals , Biomarkers, Tumor/genetics , DNA Methylation , Esophageal Neoplasms/pathology , Esophageal Squamous Cell Carcinoma/genetics , Gene Expression Regulation, Neoplastic , Humans , MiceABSTRACT
This study investigates aberrant DNA methylations as potential diagnosis and prognosis markers for esophageal squamous-cell carcinoma (ESCC), which if diagnosed at advanced stages has <30% five-year survival rate. Comparing genome-wide methylation sites of 91 ESCC and matched adjacent normal tissues, we identified 35,577 differentially methylated CpG sites (DMCs) and characterized their distribution patterns. Integrating whole-genome DNA and RNA-sequencing data of the same samples, we found multiple dysregulated transcription factors and ESCC-specific genomic correlates of identified DMCs. Using featured DMCs, we developed a 12-marker diagnostic panel with high accuracy in our dataset and the TCGA ESCC dataset, and a 4-marker prognostic panel distinguishing high-risk patients. In-vitro experiments validated the functions of 4 marker host genes. Together these results provide additional evidence for the important roles of aberrant DNA methylations in ESCC development and progression. Our DMC-based diagnostic and prognostic panels have potential values for clinical care of ESCC, laying foundations for developing targeted methylation assays for future non-invasive cancer detection methods.
Subject(s)
Carcinoma, Squamous Cell , Esophageal Neoplasms , Carcinoma, Squamous Cell/diagnosis , Carcinoma, Squamous Cell/genetics , Carcinoma, Squamous Cell/pathology , CpG Islands/genetics , DNA , DNA Methylation/genetics , Esophageal Neoplasms/diagnosis , Esophageal Neoplasms/genetics , Esophageal Neoplasms/pathology , Humans , PrognosisABSTRACT
Esophageal squamous-cell carcinoma (ESCC), one of the most prevalent and lethal malignant disease, has a complex but unknown tumor ecosystem. Here, we investigate the composition of ESCC tumors based on 208,659 single-cell transcriptomes derived from 60 individuals. We identify 8 common expression programs from malignant epithelial cells and discover 42 cell types, including 26 immune cell and 16 nonimmune stromal cell subtypes in the tumor microenvironment (TME), and analyse the interactions between cancer cells and other cells and the interactions among different cell types in the TME. Moreover, we link the cancer cell transcriptomes to the somatic mutations and identify several markers significantly associated with patients' survival, which may be relevant to precision care of ESCC patients. These results reveal the immunosuppressive status in the ESCC TME and further our understanding of ESCC.
Subject(s)
Esophageal Neoplasms/genetics , Esophageal Squamous Cell Carcinoma/genetics , Neoplasm Proteins/genetics , Stromal Cells/immunology , Transcription, Genetic , Adult , Aged , B-Lymphocytes/immunology , B-Lymphocytes/pathology , Epithelial Cells/immunology , Epithelial Cells/pathology , Esophageal Neoplasms/immunology , Esophageal Neoplasms/mortality , Esophageal Neoplasms/pathology , Esophageal Squamous Cell Carcinoma/immunology , Esophageal Squamous Cell Carcinoma/mortality , Esophageal Squamous Cell Carcinoma/pathology , Female , Fibroblasts/immunology , Fibroblasts/pathology , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Humans , Male , Middle Aged , Myeloid Cells/immunology , Myeloid Cells/pathology , Neoplasm Proteins/classification , Neoplasm Proteins/immunology , Prognosis , Single-Cell Analysis , Stromal Cells/pathology , Survival Analysis , T-Lymphocytes/immunology , T-Lymphocytes/pathology , Tumor Microenvironment/genetics , Tumor Microenvironment/immunology , Whole Genome SequencingABSTRACT
Radiotherapy remains the mainstay for treatment of various types of human cancer; however, the clinical efficacy is often limited by radioresistance, in which the underlying mechanism is largely unknown. Here, using esophageal squamous cell carcinoma (ESCC) as a model, we demonstrate that guanine nucleotide exchange factor 2 (VAV2), which is overexpressed in most human cancers, plays an important role in primary and secondary radioresistance. We have discovered for the first time that VAV2 is required for the Ku70/Ku80 complex formation and participates in non-homologous end joining repair of DNA damages caused by ionizing radiation. We show that VAV2 overexpression substantially upregulates signal transducer and activator of transcription 1 (STAT1) and the STAT1 inhibitor Fludarabine can significantly promote the sensitivity of radioresistant patient-derived ESCC xenografts in vivo in mice to radiotherapy. These results shed new light on the mechanism of cancer radioresistance, which may be important for improving clinical radiotherapy.
Subject(s)
DNA Repair , Esophageal Neoplasms , Esophageal Squamous Cell Carcinoma , Gene Expression Regulation, Neoplastic/radiation effects , Proto-Oncogene Proteins c-vav/metabolism , Radiation Tolerance , Animals , Esophageal Neoplasms/genetics , Esophageal Neoplasms/metabolism , Esophageal Neoplasms/radiotherapy , Esophageal Squamous Cell Carcinoma/genetics , Esophageal Squamous Cell Carcinoma/metabolism , Esophageal Squamous Cell Carcinoma/radiotherapy , Humans , Mice , Mice, Inbred NOD , Mice, SCID , Proto-Oncogene Proteins c-vav/geneticsABSTRACT
Adenocarcinoma at the gastroesophageal junction (ACGEJ) has dismal clinical outcomes, and there are currently few specific effective therapies because of limited knowledge on its genomic and transcriptomic alterations. The present study investigates genomic and transcriptomic changes in ACGEJ from Chinese patients and analyzes their drug vulnerabilities and associations with the survival time. Here we show that the major genomic changes of Chinese ACGEJ patients are chromosome instability promoted tumorigenic focal copy-number variations and COSMIC Signature 17-featured single nucleotide variations. We provide a comprehensive profile of genetic changes that are potentially vulnerable to existing therapeutic agents and identify Signature 17-correlated IFN-α response pathway as a prognostic marker that might have practical value for clinical prognosis of ACGEJ. These findings further our understanding on the molecular biology of ACGEJ and may help develop more effective therapeutic strategies.
Subject(s)
Adenocarcinoma/genetics , Adenocarcinoma/metabolism , Esophagogastric Junction/metabolism , Genomics , Transcriptome , Adenocarcinoma/drug therapy , Aged , Asian People/genetics , Chromosomal Instability , DNA Copy Number Variations , Esophageal Neoplasms/genetics , Esophageal Neoplasms/metabolism , Female , Genes, Neoplasm/genetics , Humans , Interferon-alpha/metabolism , Kaplan-Meier Estimate , Mutation , Prognosis , Stomach Neoplasms/genetics , Stomach Neoplasms/metabolismABSTRACT
Esophageal squamous cell carcinoma (ESCC) is prevalent in some geographical regions of the world. ESCC development presents a multistep pathogenic process from inflammation to invasive cancer; however, what is critical in these processes and how they evolve is largely unknown, obstructing early diagnosis and effective treatment. Here, we create a mouse model mimicking human ESCC development and construct a single-cell ESCC developmental atlas. We identify a set of key transitional signatures associated with oncogenic evolution of epithelial cells and depict the landmark dynamic tumorigenic trajectories. An early downregulation of CD8+ response against the initial tissue damage accompanied by the transition of immune response from type 1 to type 3 results in accumulation and activation of macrophages and neutrophils, which may create a chronic inflammatory environment that promotes carcinogen-transformed epithelial cell survival and proliferation. These findings shed light on how ESCC is initiated and developed.
Subject(s)
Esophageal Neoplasms/genetics , Esophageal Neoplasms/metabolism , Gene Expression Regulation, Neoplastic , Single-Cell Analysis/methods , Transcriptome , Animals , Cell Line, Tumor , Cell Proliferation , Cell Transformation, Neoplastic/genetics , Cell Transformation, Neoplastic/metabolism , Disease Models, Animal , Down-Regulation , Epithelial Cells/pathology , Esophageal Neoplasms/immunology , Esophageal Neoplasms/pathology , Esophageal Squamous Cell Carcinoma , Female , Fibroblasts/metabolism , Fibroblasts/pathology , Humans , Mice , Mice, Inbred C57BL , T-Lymphocytes , Transcription Factors , Tumor MicroenvironmentABSTRACT
Rationale: Whole-genome sequencing has identified many amplified genes in esophageal squamous-cell carcinoma (ESCC). This study investigated the role and clinical relevance of these genes in ESCC. Methods: We collected ESCC and non-tumor esophageal tissues from 225 individuals who underwent surgery. Clinical data were collected and survival time was measured from the date of diagnosis to the date of last follow-up or death. Patient survival was compared with immunohistochemical staining score using Kaplan-Meier methods and hazard ratios were calculated by Cox models. Cells with gene overexpression and knockout were analyzed in proliferation, migration and invasion assays. Cells were also analyzed for levels of intracellular lactate, NADPH, ATP and mRNA and protein expression patterns. Protein levels in cell line and tissue samples were measured by immunoblotting or immunohistochemistry. ESCC cell were grown as xenograft tumors in nude mice. Primary ESCC in genetically engineered mice and patient-derived xenograft mouse models were established for test of therapeutic effects. Results: We show that TP53-induced glycolysis and apoptosis regulator (TIGAR) is a major player in ESCC progression and chemoresistance. TIGAR reprograms glucose metabolism from glycolysis to the glutamine pathway through AMP-activated kinase, and its overexpression is correlated with poor disease outcomes. Tigar knockout mice have reduced ESCC tumor burden and growth rates. Treatment of TIGAR-overexpressing ESCC cell xenografts and patient-derived tumor xenografts in mice with combination of glutaminase inhibitor and chemotherapeutic agents achieves significant more efficacy than chemotherapy alone. Conclusion: These findings shed light on an important role of TIGAR in ESCC and might provide evidence for targeted treatment of TIGAR-overexpressing ESCC.
Subject(s)
Antineoplastic Agents/therapeutic use , Apoptosis Regulatory Proteins/genetics , Esophageal Neoplasms/drug therapy , Esophageal Squamous Cell Carcinoma/drug therapy , Neoplasm Proteins/genetics , Phosphoric Monoester Hydrolases/genetics , Animals , Apoptosis Regulatory Proteins/drug effects , Cell Line, Tumor , Cell Proliferation , Disease Progression , Drug Delivery Systems , Esophageal Neoplasms/genetics , Esophageal Neoplasms/mortality , Esophageal Squamous Cell Carcinoma/genetics , Esophageal Squamous Cell Carcinoma/mortality , Female , Glutaminase/antagonists & inhibitors , Humans , Mice , Mice, Inbred BALB C , Mice, Nude , Neoplasm Proteins/drug effects , Oncogenes , Phosphoric Monoester Hydrolases/drug effects , Survival Rate , Xenograft Model Antitumor AssaysABSTRACT
Esophageal squamous-cell carcinoma (ESCC) is one of the most lethal malignancies in the world and occurs at particularly higher frequency in China. While several genome-wide association studies (GWAS) of germline variants and whole-genome or whole-exome sequencing studies of somatic mutations in ESCC have been published, there is no comprehensive database publically available for this cancer. Here, we developed the Chinese Cancer Genomic Database-Esophageal Squamous Cell Carcinoma (CCGD-ESCC) database, which contains the associations of 69,593 single nucleotide polymorphisms (SNPs) with ESCC risk in 2022 cases and 2039 controls, survival time of 1006 ESCC patients (survival GWAS) and gene expression (expression quantitative trait loci, eQTL) in 94 ESCC patients. Moreover, this database also provides the associations between 8833 somatic mutations and survival time in 675 ESCC patients. Our user-friendly database is a resource useful for biologists and oncologists not only in identifying the associations of genetic variants or somatic mutations with the development and progression of ESCC but also in studying the underlying mechanisms for tumorigenesis of the cancer. CCGD-ESCC is freely accessible at http://db.cbi.pku.edu.cn/ccgd/ESCCdb.