RESUMO
Intestinal metaplasia (IM) is a pre-malignant condition of the gastric mucosa associated with increased gastric cancer (GC) risk. Analyzing 1,256 gastric samples (1,152 IMs) across 692 subjects from a prospective 10-year study, we identify 26 IM driver genes in diverse pathways including chromatin regulation (ARID1A) and intestinal homeostasis (SOX9). Single-cell and spatial profiles highlight changes in tissue ecology and IM lineage heterogeneity, including an intestinal stem-cell dominant cellular compartment linked to early malignancy. Expanded transcriptome profiling reveals expression-based molecular subtypes of IM associated with incomplete histology, antral/intestinal cell types, ARID1A mutations, inflammation, and microbial communities normally associated with the healthy oral tract. We demonstrate that combined clinical-genomic models outperform clinical-only models in predicting IMs likely to transform to GC. By highlighting strategies for accurately identifying IM patients at high GC risk and a role for microbial dysbiosis in IM progression, our results raise opportunities for GC precision prevention and interception.
Assuntos
Lesões Pré-Cancerosas , Neoplasias Gástricas , Humanos , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Estudos Prospectivos , Mucosa Gástrica/patologia , Genômica , Metaplasia/genética , Lesões Pré-Cancerosas/genéticaRESUMO
OBJECTIVE: Gastric cancer (GC) is a leading cause of cancer mortality, with ARID1A being the second most frequently mutated driver gene in GC. We sought to decipher ARID1A-specific GC regulatory networks and examine therapeutic vulnerabilities arising from ARID1A loss. DESIGN: Genomic profiling of GC patients including a Singapore cohort (>200 patients) was performed to derive mutational signatures of ARID1A inactivation across molecular subtypes. Single-cell transcriptomic profiles of ARID1A-mutated GCs were analysed to examine tumour microenvironmental changes arising from ARID1A loss. Genome-wide ARID1A binding and chromatin profiles (H3K27ac, H3K4me3, H3K4me1, ATAC-seq) were generated to identify gastric-specific epigenetic landscapes regulated by ARID1A. Distinct cancer hallmarks of ARID1A-mutated GCs were converged at the genomic, single-cell and epigenomic level, and targeted by pharmacological inhibition. RESULTS: We observed prevalent ARID1A inactivation across GC molecular subtypes, with distinct mutational signatures and linked to a NFKB-driven proinflammatory tumour microenvironment. ARID1A-depletion caused loss of H3K27ac activation signals at ARID1A-occupied distal enhancers, but unexpectedly gain of H3K27ac at ARID1A-occupied promoters in genes such as NFKB1 and NFKB2. Promoter activation in ARID1A-mutated GCs was associated with enhanced gene expression, increased BRD4 binding, and reduced HDAC1 and CTCF occupancy. Combined targeting of promoter activation and tumour inflammation via bromodomain and NFKB inhibitors confirmed therapeutic synergy specific to ARID1A-genomic status. CONCLUSION: Our results suggest a therapeutic strategy for ARID1A-mutated GCs targeting both tumour-intrinsic (BRD4-assocatiated promoter activation) and extrinsic (NFKB immunomodulation) cancer phenotypes.
Assuntos
Neoplasias Gástricas , Fatores de Transcrição , Humanos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/terapia , Neoplasias Gástricas/patologia , Proteínas Nucleares/genética , Epigenômica , Mutação , Microambiente Tumoral/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ciclo Celular/genéticaRESUMO
OBJECTIVE: Gastric cancer (GC) comprises multiple molecular subtypes. Recent studies have highlighted mesenchymal-subtype GC (Mes-GC) as a clinically aggressive subtype with few treatment options. Combining multiple studies, we derived and applied a consensus Mes-GC classifier to define the Mes-GC enhancer landscape revealing disease vulnerabilities. DESIGN: Transcriptomic profiles of ~1000 primary GCs and cell lines were analysed to derive a consensus Mes-GC classifier. Clinical and genomic associations were performed across >1200 patients with GC. Genome-wide epigenomic profiles (H3K27ac, H3K4me1 and assay for transposase-accessible chromatin with sequencing (ATAC-seq)) of 49 primary GCs and GC cell lines were generated to identify Mes-GC-specific enhancer landscapes. Upstream regulators and downstream targets of Mes-GC enhancers were interrogated using chromatin immunoprecipitation followed by sequencing (ChIP-seq), RNA sequencing, CRISPR/Cas9 editing, functional assays and pharmacological inhibition. RESULTS: We identified and validated a 993-gene cancer-cell intrinsic Mes-GC classifier applicable to retrospective cohorts or prospective single samples. Multicohort analysis of Mes-GCs confirmed associations with poor patient survival, therapy resistance and few targetable genomic alterations. Analysis of enhancer profiles revealed a distinctive Mes-GC epigenomic landscape, with TEAD1 as a master regulator of Mes-GC enhancers and Mes-GCs exhibiting preferential sensitivity to TEAD1 pharmacological inhibition. Analysis of Mes-GC super-enhancers also highlighted NUAK1 kinase as a downstream target, with synergistic effects observed between NUAK1 inhibition and cisplatin treatment. CONCLUSION: Our results establish a consensus Mes-GC classifier applicable to multiple transcriptomic scenarios. Mes-GCs exhibit a distinct epigenomic landscape, and TEAD1 inhibition and combinatorial NUAK1 inhibition/cisplatin may represent potential targetable options.
Assuntos
Elementos Facilitadores Genéticos , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Gástricas , Humanos , Cisplatino/metabolismo , Cisplatino/uso terapêutico , Estudos Prospectivos , Proteínas Quinases/genética , Proteínas Repressoras , Estudos Retrospectivos , Neoplasias Gástricas/genéticaRESUMO
Mutations in the DNA mismatch repair gene MSH2 are causative of microsatellite instability (MSI) in multiple cancers. Here, we discovered that besides its well-established role in DNA repair, MSH2 exerts a novel epigenomic function in gastric cancer. Unbiased CRISPR-based mass spectrometry combined with genome-wide CRISPR functional screening revealed that in early-stage gastric cancer MSH2 genomic binding is not randomly distributed but rather is associated specifically with tumor-associated super-enhancers controlling the expression of cell adhesion genes. At these loci, MSH2 genomic binding was required for chromatin rewiring, de novo enhancer-promoter interactions, maintenance of histone acetylation levels, and regulation of cell adhesion pathway expression. The chromatin function of MSH2 was independent of its DNA repair catalytic activity but required MSH6, another DNA repair gene, and recruitment to gene loci by the SWI/SNF chromatin remodeler SMARCA4/BRG1. Loss of MSH2 in advanced gastric cancers was accompanied by deficient cell adhesion pathway expression, epithelial-mesenchymal transition, and enhanced tumorigenesis in vitro and in vivo. However, MSH2-deficient gastric cancers also displayed addiction to BAZ1B, a bromodomain-containing family member, and consequent synthetic lethality to bromodomain and extraterminal motif (BET) inhibition. Our results reveal a role for MSH2 in gastric cancer epigenomic regulation and identify BET inhibition as a potential therapy in MSH2-deficient gastric malignancies. SIGNIFICANCE: DNA repair protein MSH2 binds and regulates cell adhesion genes by enabling enhancer-promoter interactions, and loss of MSH2 causes deficient cell adhesion and bromodomain and extraterminal motif inhibitor synthetic lethality in gastric cancer.
Assuntos
Reparo de Erro de Pareamento de DNA , Neoplasias Gástricas , Adesão Celular/genética , Cromatina/genética , DNA Helicases/genética , Reparo de Erro de Pareamento de DNA/genética , Proteínas de Ligação a DNA/genética , Mutação em Linhagem Germinativa , Humanos , Proteína 1 Homóloga a MutL/genética , Proteína 2 Homóloga a MutS/genética , Proteínas Nucleares/genética , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Fatores de Transcrição/genéticaRESUMO
Gastric cancer heterogeneity represents a barrier to disease management. We generated a comprehensive single-cell atlas of gastric cancer (>200,000 cells) comprising 48 samples from 31 patients across clinical stages and histologic subtypes. We identified 34 distinct cell-lineage states including novel rare cell populations. Many lineage states exhibited distinct cancer-associated expression profiles, individually contributing to a combined tumor-wide molecular collage. We observed increased plasma cell proportions in diffuse-type tumors associated with epithelial-resident KLF2 and stage-wise accrual of cancer-associated fibroblast subpopulations marked by high INHBA and FAP coexpression. Single-cell comparisons between patient-derived organoids (PDO) and primary tumors highlighted inter- and intralineage similarities and differences, demarcating molecular boundaries of PDOs as experimental models. We complemented these findings by spatial transcriptomics, orthogonal validation in independent bulk RNA-sequencing cohorts, and functional demonstration using in vitro and in vivo models. Our results provide a high-resolution molecular resource of intra- and interpatient lineage states across distinct gastric cancer subtypes. SIGNIFICANCE: We profiled gastric malignancies at single-cell resolution and identified increased plasma cell proportions as a novel feature of diffuse-type tumors. We also uncovered distinct cancer-associated fibroblast subtypes with INHBA-FAP-high cell populations as predictors of poor clinical prognosis. Our findings highlight potential origins of deregulated cell states in the gastric tumor ecosystem. This article is highlighted in the In This Issue feature, p. 587.
Assuntos
Fibroblastos Associados a Câncer , Neoplasias Gástricas , Fibroblastos Associados a Câncer/patologia , Ecossistema , Humanos , Análise de Célula Única , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Transcriptoma , Microambiente Tumoral/genéticaRESUMO
OBJECTIVE: To date, there are no predictive biomarkers to guide selection of patients with gastric cancer (GC) who benefit from paclitaxel. Stomach cancer Adjuvant Multi-Institutional group Trial (SAMIT) was a 2×2 factorial randomised phase III study in which patients with GC were randomised to Pac-S-1 (paclitaxel +S-1), Pac-UFT (paclitaxel +UFT), S-1 alone or UFT alone after curative surgery. DESIGN: The primary objective of this study was to identify a gene signature that predicts survival benefit from paclitaxel chemotherapy in GC patients. SAMIT GC samples were profiled using a customised 476 gene NanoString panel. A random forest machine-learning model was applied on the NanoString profiles to develop a gene signature. An independent cohort of metastatic patients with GC treated with paclitaxel and ramucirumab (Pac-Ram) served as an external validation cohort. RESULTS: From the SAMIT trial 499 samples were analysed in this study. From the Pac-S-1 training cohort, the random forest model generated a 19-gene signature assigning patients to two groups: Pac-Sensitive and Pac-Resistant. In the Pac-UFT validation cohort, Pac-Sensitive patients exhibited a significant improvement in disease free survival (DFS): 3-year DFS 66% vs 40% (HR 0.44, p=0.0029). There was no survival difference between Pac-Sensitive and Pac-Resistant in the UFT or S-1 alone arms, test of interaction p<0.001. In the external Pac-Ram validation cohort, the signature predicted benefit for Pac-Sensitive (median PFS 147 days vs 112 days, HR 0.48, p=0.022). CONCLUSION: Using machine-learning techniques on one of the largest GC trials (SAMIT), we identify a gene signature representing the first predictive biomarker for paclitaxel benefit. TRIAL REGISTRATION NUMBER: UMIN Clinical Trials Registry: C000000082 (SAMIT); ClinicalTrials.gov identifier, 02628951 (South Korean trial).
Assuntos
Adenocarcinoma , Neoplasias Gástricas , Adenocarcinoma/patologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Intervalo Livre de Doença , Humanos , Aprendizado de Máquina , Paclitaxel/uso terapêutico , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologiaRESUMO
OBJECTIVES: Epigenomic alterations in cancer interact with the immune microenvironment to dictate tumour evolution and therapeutic response. We aimed to study the regulation of the tumour immune microenvironment through epigenetic alternate promoter use in gastric cancer and to expand our findings to other gastrointestinal tumours. DESIGN: Alternate promoter burden (APB) was quantified using a novel bioinformatic algorithm (proActiv) to infer promoter activity from short-read RNA sequencing and samples categorised into APBhigh, APBint and APBlow. Single-cell RNA sequencing was performed to analyse the intratumour immune microenvironment. A humanised mouse cancer in vivo model was used to explore dynamic temporal interactions between tumour kinetics, alternate promoter usage and the human immune system. Multiple cohorts of gastrointestinal tumours treated with immunotherapy were assessed for correlation between APB and treatment outcomes. RESULTS: APBhigh gastric cancer tumours expressed decreased levels of T-cell cytolytic activity and exhibited signatures of immune depletion. Single-cell RNAsequencing analysis confirmed distinct immunological populations and lower T-cell proportions in APBhigh tumours. Functional in vivo studies using 'humanised mice' harbouring an active human immune system revealed distinct temporal relationships between APB and tumour growth, with APBhigh tumours having almost no human T-cell infiltration. Analysis of immunotherapy-treated patients with GI cancer confirmed resistance of APBhigh tumours to immune checkpoint inhibition. APBhigh gastric cancer exhibited significantly poorer progression-free survival compared with APBlow (median 55 days vs 121 days, HR 0.40, 95% CI 0.18 to 0.93, p=0.032). CONCLUSION: These findings demonstrate an association between alternate promoter use and the tumour microenvironment, leading to immune evasion and immunotherapy resistance.
Assuntos
Neoplasias Gastrointestinais , Neoplasias Gástricas , Animais , Epigênese Genética , Epigenômica , Neoplasias Gastrointestinais/genética , Neoplasias Gastrointestinais/terapia , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Imunoterapia , Camundongos , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/terapia , Microambiente TumoralRESUMO
BACKGROUND: Enhancers are distal cis-regulatory elements required for cell-specific gene expression and cell fate determination. In cancer, enhancer variation has been proposed as a major cause of inter-patient heterogeneity-however, most predicted enhancer regions remain to be functionally tested. METHODS: We analyzed 132 epigenomic histone modification profiles of 18 primary gastric cancer (GC) samples, 18 normal gastric tissues, and 28 GC cell lines using Nano-ChIP-seq technology. We applied Capture-based Self-Transcribing Active Regulatory Region sequencing (CapSTARR-seq) to assess functional enhancer activity. An Activity-by-contact (ABC) model was employed to explore the effects of histone acetylation and CapSTARR-seq levels on enhancer-promoter interactions. RESULTS: We report a comprehensive catalog of 75,730 recurrent predicted enhancers, the majority of which are GC-associated in vivo (> 50,000) and associated with lower somatic mutation rates inferred by whole-genome sequencing. Applying CapSTARR-seq to the enhancer catalog, we observed significant correlations between CapSTARR-seq functional activity and H3K27ac/H3K4me1 levels. Super-enhancer regions exhibited increased CapSTARR-seq signals compared to regular enhancers, even when decoupled from native chromatin contexture. We show that combining histone modification and CapSTARR-seq functional enhancer data improves the prediction of enhancer-promoter interactions and pinpointing of germline single nucleotide polymorphisms (SNPs), somatic copy number alterations (SCNAs), and trans-acting TFs involved in GC expression. We identified cancer-relevant genes (ING1, ARL4C) whose expression between patients is influenced by enhancer differences in genomic copy number and germline SNPs, and HNF4α as a master trans-acting factor associated with GC enhancer heterogeneity. CONCLUSIONS: Our results indicate that combining histone modification and functional assay data may provide a more accurate metric to assess enhancer activity than either platform individually, providing insights into the relative contribution of genetic (cis) and regulatory (trans) mechanisms to GC enhancer functional heterogeneity.
Assuntos
Elementos Facilitadores Genéticos , Epigenômica , Neoplasias Gástricas/genética , Fatores de Ribosilação do ADP/genética , Fatores de Ribosilação do ADP/metabolismo , Acetilação , Linhagem Celular Tumoral , Proliferação de Células , Cromatina , Regulação Neoplásica da Expressão Gênica , Genômica , Histonas/metabolismo , Humanos , Proteína 1 Inibidora do Crescimento/genética , Proteína 1 Inibidora do Crescimento/metabolismo , Oncogenes , Regiões Promotoras Genéticas , RNA-Seq , Transcriptoma , Sequenciamento Completo do GenomaRESUMO
BACKGROUND: CIMP (CpG island methylator phenotype) is an epigenetic molecular subtype, observed in multiple malignancies and associated with the epigenetic silencing of tumor suppressors. Currently, for most cancers including gastric cancer (GC), mechanisms underlying CIMP remain poorly understood. We sought to discover molecular contributors to CIMP in GC, by performing global DNA methylation, gene expression, and proteomics profiling across 14 gastric cell lines, followed by similar integrative analysis in 50 GC cell lines and 467 primary GCs. RESULTS: We identify the cystathionine beta-synthase enzyme (CBS) as a highly recurrent target of epigenetic silencing in CIMP GC. Likewise, we show that CBS epimutations are significantly associated with CIMP in various other cancers, occurring even in premalignant gastroesophageal conditions and longitudinally linked to clinical persistence. Of note, CRISPR deletion of CBS in normal gastric epithelial cells induces widespread DNA methylation changes that overlap with primary GC CIMP patterns. Reflecting its metabolic role as a gatekeeper interlinking the methionine and homocysteine cycles, CBS loss in vitro also causes reductions in the anti-inflammatory gasotransmitter hydrogen sulfide (H2S), with concomitant increase in NF-κB activity. In a murine genetic model of CBS deficiency, preliminary data indicate upregulated immune-mediated transcriptional signatures in the stomach. CONCLUSIONS: Our results implicate CBS as a bi-faceted modifier of aberrant DNA methylation and inflammation in GC and highlights H2S donors as a potential new therapy for CBS-silenced lesions.
Assuntos
Ilhas de CpG/genética , Cistationina beta-Sintase/genética , Metilação de DNA/genética , Inflamação/genética , Mutação/genética , Neoplasias Gástricas/genética , Animais , Linhagem Celular Tumoral , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Deleção de Genes , Humanos , Intestinos/patologia , Metaplasia , Camundongos Transgênicos , Fenótipo , Proteoma/metabolismo , Transcriptoma/genéticaRESUMO
Gastric cancer cases are often diagnosed at an advanced stage with poor prognosis. Platinum-based chemotherapy has been internationally accepted as first-line therapy for inoperable or metastatic gastric cancer. To achieve greater benefits, selection of patients eligible for this treatment is critical. Although gene expression profiling has been widely used as a genomic classifier to identify molecular subtypes of gastric cancer and to stratify patients for different chemotherapy regimens, its prediction accuracy can be improved. Adenosine-to-inosine (A-to-I) RNA editing has emerged as a new player contributing to gastric cancer development and progression, offering potential clinical utility for diagnosis and treatment. Using a systematic computational approach followed by both in vitro validations and in silico validations in The Cancer Genome Atlas (TCGA), we conducted a transcriptome-wide RNA editing analysis of a cohort of 104 patients with advanced gastric cancer and identified an RNA editing (GCRE) signature to guide gastric cancer chemotherapy. RNA editing events stood as a prognostic and predictive biomarker in advanced gastric cancer. A GCRE score based on the GCRE signature consisted of 50 editing sites associated with 29 genes, predicting response to chemotherapy with a high accuracy (84%). Of note, patients demonstrating higher editing levels of this panel of sites presented a better overall response. Consistently, gastric cancer cell lines with higher editing levels showed higher chemosensitivity. Applying the GCRE score on TCGA dataset confirmed that responders had significantly higher levels of editing in advanced gastric cancer. Overall, this newly defined GCRE signature reliably stratifies patients with advanced gastric cancer and predicts response from chemotherapy. SIGNIFICANCE: This study describes a novel A-to-I RNA editing signature as a prognostic and predictive biomarker in advanced gastric cancer, providing a new tool to improve patient stratification and response to therapy.
Assuntos
Adenocarcinoma/tratamento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biomarcadores Tumorais/genética , Regulação Neoplásica da Expressão Gênica , Recidiva Local de Neoplasia/tratamento farmacológico , Edição de RNA , Neoplasias Gástricas/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/patologia , Ensaios Clínicos como Assunto , Estudos de Coortes , Perfilação da Expressão Gênica , Humanos , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Prognóstico , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Taxa de SobrevidaRESUMO
BACKGROUND: Deregulated gene expression is a hallmark of cancer; however, most studies to date have analyzed short-read RNA sequencing data with inherent limitations. Here, we combine PacBio long-read isoform sequencing (Iso-Seq) and Illumina paired-end short-read RNA sequencing to comprehensively survey the transcriptome of gastric cancer (GC), a leading cause of global cancer mortality. RESULTS: We performed full-length transcriptome analysis across 10 GC cell lines covering four major GC molecular subtypes (chromosomal unstable, Epstein-Barr positive, genome stable and microsatellite unstable). We identify 60,239 non-redundant full-length transcripts, of which > 66% are novel compared to current transcriptome databases. Novel isoforms are more likely to be cell line and subtype specific, expressed at lower levels with larger number of exons, with longer isoform/coding sequence lengths. Most novel isoforms utilize an alternate first exon, and compared to other alternative splicing categories, are expressed at higher levels and exhibit higher variability. Collectively, we observe alternate promoter usage in 25% of detected genes, with the majority (84.2%) of known/novel promoter pairs exhibiting potential changes in their coding sequences. Mapping these alternate promoters to TCGA GC samples, we identify several cancer-associated isoforms, including novel variants of oncogenes. Tumor-specific transcript isoforms tend to alter protein coding sequences to a larger extent than other isoforms. Analysis of outcome data suggests that novel isoforms may impart additional prognostic information. CONCLUSIONS: Our results provide a rich resource of full-length transcriptome data for deeper studies of GC and other gastrointestinal malignancies.
Assuntos
Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Transcriptoma , Proteínas Adaptadoras de Transdução de Sinal , Processamento Alternativo , Linhagem Celular Tumoral , Éxons , Perfilação da Expressão Gênica , Genoma , Humanos , Fases de Leitura Aberta , Isoformas de Proteínas , Análise de Sequência de RNARESUMO
Transcriptional reactivation of telomerase catalytic subunit (TERT) is a frequent hallmark of cancer, occurring in 90% of human malignancies. However, specific mechanisms driving TERT reactivation remain obscure for many tumor types and in particular gastric cancer (GC), a leading cause of global cancer mortality. Here, through comprehensive genomic and epigenomic analysis of primary GCs and GC cell lines, we identified the transcription factor early B cell factor 1 (EBF1) as a TERT transcriptional repressor and inactivation of EBF1 function as a major cause of TERT upregulation. Abolishment of EBF1 function occurs through 3 distinct (epi)genomic mechanisms. First, EBF1 is epigenetically silenced via DNA methyltransferase, polycomb-repressive complex 2 (PRC2), and histone deacetylase activity in GCs. Second, recurrent, somatic, and heterozygous EBF1 DNA-binding domain mutations result in the production of dominant-negative EBF1 isoforms. Third, more rarely, genomic deletions and rearrangements proximal to the TERT promoter remobilize or abolish EBF1-binding sites, derepressing TERT and leading to high TERT expression. EBF1 is also functionally required for various malignant phenotypes in vitro and in vivo, highlighting its importance for GC development. These results indicate that multimodal genomic and epigenomic alterations underpin TERT reactivation in GC, converging on transcriptional repressors such as EBF1.
Assuntos
Epigenômica , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/metabolismo , Neoplasias Gástricas/metabolismo , Telomerase/biossíntese , Transativadores/metabolismo , Linhagem Celular Tumoral , Humanos , Mutação , Proteínas de Neoplasias/genética , Elementos de Resposta , Neoplasias Gástricas/genética , Telomerase/genética , Transativadores/genéticaRESUMO
OBJECTIVE: Gastric cancer (GC) is a leading cause of cancer mortality. Previous studies have shown that hepatocyte nuclear factor-4α (HNF4α) is specifically overexpressed in GC and functionally required for GC development. In this study, we investigated, on a genome-wide scale, target genes of HNF4α and oncogenic pathways driven by HNF4α and HNF4α target genes. DESIGN: We performed HNF4α chromatin immunoprecipitation followed by sequencing across multiple GC cell lines, integrating HNF4α occupancy data with (epi)genomic and transcriptome data of primary GCs to define HNF4α target genes of in vitro and in vivo relevance. To investigate mechanistic roles of HNF4α and HNF4α targets, we performed cancer metabolic measurements, drug treatments and functional assays including murine xenograft experiments. RESULTS: Gene expression analysis across 19 tumour types revealed HNF4α to be specifically upregulated in GCs. Unbiased pathway analysis revealed organic acid metabolism as the top HNF4α-regulated pathway, orthogonally supported by metabolomic analysis. Isocitrate dehydrogenase 1 (IDH1) emerged as a convergent HNF4α direct target gene regulating GC metabolism. We show that wild-type IDH1 is essential for GC cell survival, and that certain GC cells can be targeted by IDH1 inhibitors. CONCLUSIONS: Our results highlight a role for HNF4α in sustaining GC oncogenic metabolism, through the regulation of IDH1. Drugs targeting wild-type IDH1 may thus have clinical utility in GCs exhibiting HNF4α overexpression, expanding the role of IDH1 in cancer beyond IDH1/2 mutated malignancies.
Assuntos
Fator 4 Nuclear de Hepatócito/genética , Isocitrato Desidrogenase/metabolismo , Neoplasias Gástricas/genética , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Feminino , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Marcação de Genes/métodos , Fator 4 Nuclear de Hepatócito/metabolismo , Humanos , Isocitrato Desidrogenase/antagonistas & inibidores , Camundongos Endogâmicos NOD , Terapia de Alvo Molecular/métodos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Regiões Promotoras Genéticas/genética , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Regulação para Cima/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
OBJECTIVE: Genomic structural variations (SVs) causing rewiring of cis-regulatory elements remain largely unexplored in gastric cancer (GC). To identify SVs affecting enhancer elements in GC (enhancer-based SVs), we integrated epigenomic enhancer profiles revealed by paired-end H3K27ac ChIP-sequencing from primary GCs with tumour whole-genome sequencing (WGS) data (PeNChIP-seq/WGS). DESIGN: We applied PeNChIP-seq to 11 primary GCs and matched normal tissues combined with WGS profiles of >200 GCs. Epigenome profiles were analysed alongside matched RNA-seq data to identify tumour-associated enhancer-based SVs with altered cancer transcription. Functional validation of candidate enhancer-based SVs was performed using CRISPR/Cas9 genome editing, chromosome conformation capture assays (4C-seq, Capture-C) and Hi-C analysis of primary GCs. RESULTS: PeNChIP-seq/WGS revealed ~150 enhancer-based SVs in GC. The majority (63%) of SVs linked to target gene deregulation were associated with increased tumour expression. Enhancer-based SVs targeting CCNE1, a key driver of therapy resistance, occurred in 8% of patients frequently juxtaposing diverse distal enhancers to CCNE1 proximal regions. CCNE1-rearranged GCs were associated with high CCNE1 expression, disrupted CCNE1 topologically associating domain (TAD) boundaries, and novel TAD interactions in CCNE1-rearranged primary tumours. We also observed IGF2 enhancer-based SVs, previously noted in colorectal cancer, highlighting a common non-coding genetic driver alteration in gastric and colorectal malignancies. CONCLUSION: Integrated paired-end NanoChIP-seq and WGS of gastric tumours reveals tumour-associated regulatory SV in regions associated with both simple and complex genomic rearrangements. Genomic rearrangements may thus exploit enhancer-hijacking as a common mechanism to drive oncogene expression in GC.
Assuntos
Adenocarcinoma/metabolismo , Ciclina E/metabolismo , Elementos Facilitadores Genéticos/genética , Fator de Crescimento Insulin-Like II/metabolismo , Proteínas Oncogênicas/metabolismo , Neoplasias Gástricas/metabolismo , Adenocarcinoma/genética , Variação Estrutural do Genoma/genética , Humanos , Neoplasias Gástricas/genética , Sequenciamento Completo do GenomaRESUMO
BACKGROUND: Epstein-Barr virus-associated gastric cancer (EBVaGC) has traditionally been associated with high expression of PD-L1 and immune infiltration. Correlations between PD-L1 and other immune-related gene (IRG) expressions in EBVaGC have not been previously described. METHODS: We performed NanoString® transcriptomic profiling and PD-L1 immunohistochemistry (IHC) (using the FDA approved Dako PD-L1 IHC 22C3) on EBVaGC samples from gastric cancer patients undergoing primary tumor resections at Samsung Medical Centre, South Korea. For controls, EBV-negative samples from the previously reported Asian Cancer Research Group (EBVnegACRG) cohort were used. Genes tested included PD-L1 and other IRGs related to intra-tumoral cytolytic activity, cytokines and immune checkpoints. Samples with PD-L1 expression > 34th percentile were defined as PD-L1high and the remaining as PD-L1low. RESULTS: We identified 71 cases of EBVaGC and 193 EBV-negative ACRG samples as controls. EBVaGC showed higher expression of all queried immune genes compared to EBVnegACRG samples (p < 0.01). PD-L1 immunohistochemistry expression correlated with PD-L1 transcript expression (r = 0.63, p < 0.001). Tumor-infiltrating lymphocyte patterns were also found to be different between PD-L1low and PD-L1high groups. PD-L1low EBVaGC samples (n = 24, 34%) had consistently decreased expression of all other immune genes, such as CD8A, GZMA and PRF1 and PD-1 (p < 0.001). PD-L1low EBVaGC samples were also associated with worse disease-free survival (HR 5.03, p = 0.032) compared to PD-L1high EBVaGC samples. CONCLUSIONS: A substantial proportion of EBVaGC does not express high levels of PD-L1 and other immune genes. EBVaGCs which have lower transcriptomic expression of PD-L1 tend to have a similarly low expression of other immune genes, IHC scores and a poorer prognosis.
Assuntos
Infecções por Vírus Epstein-Barr/imunologia , Regulação Neoplásica da Expressão Gênica , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/virologia , Adulto , Idoso , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Biomarcadores Tumorais/genética , Antígenos CD8/genética , Estudos de Coortes , Intervalo Livre de Doença , Infecções por Vírus Epstein-Barr/complicações , Feminino , Perfilação da Expressão Gênica , Herpesvirus Humano 4/patogenicidade , Humanos , Imuno-Histoquímica , Linfócitos do Interstício Tumoral/patologia , Masculino , Pessoa de Meia-Idade , Perforina/genética , Receptor de Morte Celular Programada 1/genética , Neoplasias Gástricas/genéticaRESUMO
Intestinal metaplasia (IM) is a pre-malignant condition of the gastric mucosa associated with increased gastric cancer (GC) risk. We performed (epi)genomic profiling of 138 IMs from 148 cancer-free patients, recruited through a 10-year prospective study. Compared with GCs, IMs exhibit low mutational burdens, recurrent mutations in certain tumor suppressors (FBXW7) but not others (TP53, ARID1A), chromosome 8q amplification, and shortened telomeres. Sequencing identified more IM patients with active Helicobacter pylori infection compared with histopathology (11%-27%). Several IMs exhibited hypermethylation at DNA methylation valleys; however, IMs generally lack intragenic hypomethylation signatures of advanced malignancy. IM patients with shortened telomeres and chromosomal alterations were associated with subsequent dysplasia or GC; conversely patients exhibiting normal-like epigenomic patterns were associated with regression.
Assuntos
Mucosa Gástrica/patologia , Infecções por Helicobacter/genética , Metaplasia/genética , Lesões Pré-Cancerosas/genética , Neoplasias Gástricas/etiologia , Adulto , Idoso , Metilação de DNA , Progressão da Doença , Epigenômica , Feminino , Mucosa Gástrica/microbiologia , Genômica , Infecções por Helicobacter/microbiologia , Humanos , Masculino , Metaplasia/microbiologia , Pessoa de Meia-Idade , Lesões Pré-Cancerosas/patologia , Neoplasias Gástricas/genética , Neoplasias Gástricas/microbiologiaRESUMO
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RESUMO
The switch between quiescence and proliferation is central for neurogenesis and its alteration is linked to neurodevelopmental disorders such as microcephaly. However, intrinsic mechanisms that reactivate Drosophila larval neural stem cells (NSCs) to exit from quiescence are not well established. Here we show that the spindle matrix complex containing Chromator (Chro) functions as a key intrinsic regulator of NSC reactivation downstream of extrinsic insulin/insulin-like growth factor signalling. Chro also prevents NSCs from re-entering quiescence at later stages. NSC-specific in vivo profiling has identified many downstream targets of Chro, including a temporal transcription factor Grainy head (Grh) and a neural stem cell quiescence-inducing factor Prospero (Pros). We show that spindle matrix proteins promote the expression of Grh and repress that of Pros in NSCs to govern their reactivation. Our data demonstrate that nuclear Chro critically regulates gene expression in NSCs at the transition from quiescence to proliferation.The spindle matrix proteins, including Chro, are known to regulate mitotic spindle assembly in the cytoplasm. Here the authors show that in Drosophila larval brain, Chro promotes neural stem cell (NSC) reactivation and prevents activated NSCs from entering quiescence, and that Chro carries out such a role by regulating the expression of key transcription factors in the nucleus.