RESUMO
The majority of low-grade isocitrate dehydrogenase-mutant (IDHmt) gliomas undergo malignant progression (MP), but their underlying mechanism remains unclear. IDHmt gliomas exhibit global DNA methylation, and our previous report suggested that MP could be partly attributed to passive demethylation caused by accelerated cell cycles. However, during MP, there is also active demethylation mediated by ten-eleven translocation, such as DNA hydroxymethylation. Hydroxymethylation is reported to potentially contribute to gene expression regulation, but its role in MP remains under investigation. Therefore, we conducted a comprehensive analysis of hydroxymethylation during MP of IDHmt astrocytoma. Five primary/malignantly progressed IDHmt astrocytoma pairs were analyzed with oxidative bisulfite and the Infinium EPIC methylation array, detecting 5-hydroxymethyl cytosine at over 850,000 locations for region-specific hydroxymethylation assessment. Notably, we observed significant sharing of hydroxymethylated genomic regions during MP across the samples. Hydroxymethylated CpGs were enriched in open sea and intergenic regions (p < 0.001), and genes undergoing hydroxymethylation were significantly associated with cancer-related signaling pathways. RNA sequencing data integration identified 91 genes with significant positive/negative hydroxymethylation-expression correlations. Functional analysis suggested that positively correlated genes are involved in cell-cycle promotion, while negatively correlated ones are associated with antineoplastic functions. Analyses of The Cancer Genome Atlas clinical data on glioma were in line with these findings. Motif-enrichment analysis suggested the potential involvement of the transcription factor KLF4 in hydroxymethylation-based gene regulation. Our findings shed light on the significance of region-specific DNA hydroxymethylation in glioma MP and suggest its potential role in cancer-related gene expression and IDHmt glioma malignancy.
Assuntos
Neoplasias Encefálicas , Metilação de DNA , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Glioma , Isocitrato Desidrogenase , Fator 4 Semelhante a Kruppel , Mutação , Humanos , Isocitrato Desidrogenase/genética , Glioma/genética , Glioma/patologia , Glioma/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Ilhas de CpG/genética , Feminino , Masculino , Astrocitoma/genética , Astrocitoma/patologia , Astrocitoma/metabolismo , Pessoa de Meia-Idade , 5-Metilcitosina/análogos & derivados , 5-Metilcitosina/metabolismo , AdultoRESUMO
Ependymomas encompass multiple clinically relevant tumor types based on localization and molecular profiles. Tumors of the methylation class "spinal ependymoma" (SP-EPN) represent the most common intramedullary neoplasms in children and adults. However, their developmental origin is ill-defined, molecular data are scarce, and the potential heterogeneity within SP-EPN remains unexplored. The only known recurrent genetic events in SP-EPN are loss of chromosome 22q and NF2 mutations, but neither types and frequency of these alterations nor their clinical relevance have been described in a large, epigenetically defined series. Transcriptomic (n = 72), epigenetic (n = 225), genetic (n = 134), and clinical data (n = 112) were integrated for a detailed molecular overview on SP-EPN. Additionally, we mapped SP-EPN transcriptomes to developmental atlases of the developing and adult spinal cord to uncover potential developmental origins of these tumors. The integration of transcriptomic ependymoma data with single-cell atlases of the spinal cord revealed that SP-EPN display the highest similarities to mature adult ependymal cells. Unsupervised hierarchical clustering of transcriptomic data together with integrated analysis of methylation profiles identified two molecular SP-EPN subtypes. Subtype A tumors primarily carried previously known germline or sporadic NF2 mutations together with 22q loss (bi-allelic NF2 loss), resulting in decreased NF2 expression. Furthermore, they more often presented as multilocular disease and demonstrated a significantly reduced progression-free survival as compared to SP-EP subtype B. In contrast, subtype B predominantly contained samples without NF2 mutation detected in sequencing together with 22q loss (monoallelic NF2 loss). These tumors showed regular NF2 expression but more extensive global copy number alterations. Based on integrated molecular profiling of a large multi-center cohort, we identified two distinct SP-EPN subtypes with important implications for genetic counseling, patient surveillance, and drug development priorities.
Assuntos
Ependimoma , Neoplasias da Medula Espinal , Adulto , Criança , Humanos , Transcriptoma , Perfilação da Expressão Gênica , Mutação , Epigênese GenéticaRESUMO
Epstein-Barr virus (EBV) is one of the major drivers of gastric carcinogenesis. EBV infection is established before tumour initiation and is generally maintained throughout tumour development; however, the significance of EBV in tumour maintenance and progression remains to be elucidated. Here, we report eight cases of EBV-associated gastric carcinoma (EBVaGC) with intratumoural heterogenous expression of EBV-encoded small RNA (EBER), a highly expressed latent gene of EBV, and demonstrate clinicopathological characteristics of these rare cases. By performing detailed histological assessment of EBER-positive and -negative components of each case, detection of EBV genome in tumour cells by fluorescence in situ hybridisation, TP73 methylation analysis, whole exome sequencing, and targeted gene panel sequencing, we identified tumours in two patients to be collision tumours of different origins. In the other six patients, some genetic/epigenetic alterations were shared between EBER-positive and -negative components, suggesting that EBV was eliminated from tumour cells during progression. Interestingly, in both tumour types, programmed death ligand 1 and intratumoural infiltration of CD8+ T lymphocytes were lower in EBER-negative than in EBER-positive components, suggesting an immunogenic role of EBV. To the best of our knowledge, this study is the first to demonstrate the detailed histological features and genetic/epigenetic alterations in EBVaGC with heterogenous EBER expression; the loss of EBV may benefit tumour progression and immune evasion and might be clinically important for selecting treatment strategies for such cancers. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
Assuntos
Carcinoma , Infecções por Vírus Epstein-Barr , Neoplasias Gástricas , Humanos , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/metabolismo , Infecções por Vírus Epstein-Barr/complicações , Infecções por Vírus Epstein-Barr/genética , Infecções por Vírus Epstein-Barr/patologia , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Genoma Viral , Carcinoma/genética , RNA Viral/genética , Microambiente TumoralRESUMO
Comprehensive cancer genome profiling (CGP) has been nationally reimbursed in Japan since June 2019. Less than 10% of the patients have been reported to undergo recommended treatment. Todai OncoPanel (TOP) is a dual DNA-RNA panel as well as a paired tumor-normal matched test. Two hundred patients underwent TOP as part of Advanced Medical Care B with approval from the Ministry of Health, Labour and Welfare between September 2018 and December 2019. Tests were carried out in patients with cancers without standard treatment or when patients had already undergone standard treatment. Data from DNA and RNA panels were analyzed in 198 and 191 patients, respectively. The percentage of patients who were given therapeutic or diagnostic recommendations was 61% (120/198). One hundred and four samples (53%) harbored gene alterations that were detected with the DNA panel and had potential treatment implications, and 14 samples (7%) had a high tumor mutational burden. Twenty-two samples (11.1%) harbored 30 fusion transcripts or MET exon 14 skipping that were detected by the RNA panel. Of those 30 transcripts, 6 had treatment implications and 4 had diagnostic implications. Thirteen patients (7%) were found to have pathogenic or likely pathogenic germline variants and genetic counseling was recommended. Overall, 12 patients (6%) received recommended treatment. In summary, patients benefited from both TOP DNA and RNA panels while following the same indication as the approved CGP tests. (UMIN000033647).
Assuntos
Genômica , Neoplasias , Humanos , Japão , Neoplasias/tratamento farmacológico , Neoplasias/genética , Medicina de PrecisãoRESUMO
BACKGROUND: Tumors with a high number of mutations in the genome, or tumor mutational burden, are presumed to be more likely to respond to immune checkpoint inhibitors. However, the optimal method to calculate tumor mutational burden using comprehensive genomic profiling assays is unknown. METHODS: Todai OncoPanel is a dual panel of a deoxyribonucleic acid panel and a ribonucleic acid panel. Todai OncoPanel deoxyribonucleic acid panel version 6 is an improvement over version 3 with increased number of targeted genes and limited targeting of intronic regions. We calculated tumor mutational burden measured by Todai OncoPanel deoxyribonucleic acid panel versions 3 and 6 using three different calculation methods: all mutations within the targeted region (target tumor mutational burden), all mutations within the coding region (all coding tumor mutational burden) and non-synonymous mutations (non-synonymous coding tumor mutational burden). We then compared them with whole exosome sequencing tumor mutational burden. In addition, 16 lung cancer patients whose samples were analyzed using Todai OncoPanel deoxyribonucleic acid version 3 were treated with anti-PD-1 or PD-L1 antibody monotherapy. RESULTS: When compared with whole exosome sequencing tumor mutational burden as the standard, tumor mutational burden measured by Todai OncoPanel deoxyribonucleic acid version 3 resulted in accuracy of 71% for all three calculation methods. In version 6, accuracy was 96% for target tumor mutational burden and all coding tumor mutational burden and 91% for non-synonymous coding tumor mutational burden. Patients with either partial response or stable disease had higher non-synonymous coding tumor mutational burden (6.7/Mb vs. 1.6/Mb, P = 0.02) and higher PD-L1 expression (40% vs. 3%, P = 0.01) and a trend toward higher target tumor mutational burden (9.2/Mb vs. 2.4/Mb, P = 0.09) compared with patients with progressive disease. CONCLUSIONS: Increase in targeted gene number and limiting intronic regions improved tumor mutational burden measurement by Todai OncoPanel when compared with whole exosome sequencing tumor mutational burden. Target tumor mutational burden may be the method of choice to measure tumor mutational burden.
Assuntos
Antígeno B7-H1 , Neoplasias Pulmonares , Antígeno B7-H1/genética , Biomarcadores Tumorais/genética , DNA , Genômica , Humanos , Neoplasias Pulmonares/genética , Mutação , Carga TumoralRESUMO
The organoid culture technique has been recently applied to modeling carcinogenesis in several organs. To further explore its potential and gain novel insights into tumorigenesis, we here investigated whether pancreatic ductal adenocarcinoma (PDA) could be generated as subcutaneous tumors in immunocompromised nude mice, by genetic engineering of normal organoids. As expected, acute induction of KrasG12Din vitro occasionally led to development of tiny nodules compatible with early lesions known as pancreatic intraepithelial neoplasia (PanIN). KrasG12D-expressing cells were enriched after inoculation in the subcutis, yet proved rather declined during culture, suggesting that its advantage might depend on surrounding environments. Depletion of growth factors or concurrent Trp53 deletion resulted in its robust enrichment, invariably leading to development of PanIN or large high-grade adenocarcinoma, respectively, consistent with in vivo mouse studies for the same genotype. Progression from PanIN was also recapitulated by subsequent knockdown of common tumor suppressors, whereas the impact of Tgfbr2 deletion was only partially recapitulated, illustrating genotype-dependent requirement of the pancreatic niche for tumorigenesis. Intriguingly, analysis of tumor-derived organoids revealed that KrasG12D-expressing cells with spontaneous deletion of wild-type Kras were positively selected and exhibited an aging-related mutation signature in nude mice, mirroring the pathogenesis of human PDA, and that the sphere-forming potential and orthotopic tumorigenicity in syngenic mice were significantly augmented. These observations highlighted the relevance of the subcutis of nude mice in promoting PDA development despite its ectopic nature. Taken together, pancreatic carcinogenesis could be considerably recapitulated with organoids, which would probably serve as a novel disease model.
Assuntos
Carcinoma Ductal Pancreático/patologia , Transformação Celular Neoplásica/patologia , Mutação , Organoides/patologia , Ductos Pancreáticos/patologia , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Apoptose , Carcinoma Ductal Pancreático/genética , Proliferação de Células , Transformação Celular Neoplásica/genética , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Nus , Organoides/metabolismo , Ductos Pancreáticos/metabolismo , Neoplasias Pancreáticas/genética , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/genética , Ensaios Antitumorais Modelo de Xenoenxerto , Neoplasias PancreáticasRESUMO
Multiple hepatocellular carcinoma (HCC) is divided into two categories: intrahepatic metastasis (IM), which is a true relapse of HCC, and multicentric origin (MO), which is a second primary tumor. Clinical diagnosis of multiple HCC is usually made based on tumor location and/or time to recurrence; however, it is often difficult to distinguish the two types of multiple HCC. Using 41 matched pairs of multiple HCC specimens, we confirmed the accuracy of clinical diagnoses using exome sequence data and investigated the importance of discriminating the type of multiple HCC. Genomic analysis revealed that 18 (43.9%) patients diagnosed as having genomic IM had common mutations in a pair of HCC tumors with the main tumor of these patients being more progressive compared to those with genomic MO. The accuracy of clinical diagnosis based on lobe (Definition 1) and segment (Definition 2) were 68.3% and 78.0%, respectively. Intriguingly, recurrence ≥2 years after initial surgery for 3 patients was IM. The survival of patients with clinical IM was significantly shorter than for those with clinical MO based on both Definition 1 (P = 0.045) and Definition 2 (P = 0.043). However, mean survival was not different between the patients with genomic IM and those with MO (P = 0.364). Taken together, genomic analysis elucidated that liver cancer may spread more extensively and more slowly than previously thought. In addition, distinguishing multiple HCC as IM or MC may have provided biological information but was not of clinical importance with respect to patient prognosis.
Assuntos
Carcinoma Hepatocelular/genética , Sequenciamento do Exoma/métodos , Neoplasias Hepáticas/genética , Metástase Neoplásica/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Hepatocelular/cirurgia , Células Clonais/metabolismo , Feminino , Hepatectomia , Humanos , Neoplasias Hepáticas/cirurgia , Masculino , Pessoa de Meia-Idade , PrognósticoRESUMO
Birt-Hogg-Dubé (BHD) syndrome is a hereditary kidney cancer syndrome, which predisposes patients to develop kidney cancer, cutaneous fibrofolliculomas and pulmonary cysts. The responsible gene FLCN is a tumor suppressor for kidney cancer, which plays an important role in energy homeostasis through the regulation of mitochondrial oxidative metabolism. However, the process by which FLCN-deficiency leads to renal tumorigenesis is unclear. In order to clarify molecular pathogenesis of BHD-associated kidney cancer, we conducted whole-exome sequencing analysis using next-generation sequencing technology as well as metabolite analysis using liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry. Whole-exome sequencing analysis of BHD-associated kidney cancer revealed that copy number variations of BHD-associated kidney cancer are considerably different from those already reported in sporadic cases. In somatic variant analysis, very few variants were commonly observed in BHD-associated kidney cancer; however, variants in chromatin remodeling genes were frequently observed in BHD-associated kidney cancer (17/29 tumors, 59%). Metabolite analysis of BHD-associated kidney cancer revealed metabolic reprogramming toward upregulated redox regulation which may neutralize reactive oxygen species potentially produced from mitochondria with increased respiratory capacity under FLCN-deficiency. BHD-associated kidney cancer displays unique molecular characteristics that are completely different from sporadic kidney cancer, providing mechanistic insight into tumorigenesis under FLCN-deficiency as well as a foundation for development of novel therapeutics for kidney cancer.
Assuntos
Síndrome de Birt-Hogg-Dubé/patologia , Montagem e Desmontagem da Cromatina/genética , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Síndrome de Birt-Hogg-Dubé/genética , Variações do Número de Cópias de DNA , Mutação em Linhagem Germinativa , Humanos , Neoplasias Renais/patologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Sequenciamento do ExomaRESUMO
Tumor molecular profiling is becoming a standard of care for patients with cancer, but the optimal platform for cancer sequencing remains undetermined. We established a comprehensive assay, the Todai OncoPanel (TOP), which consists of DNA and RNA hybridization capture-based next-generation sequencing panels. A novel method for target enrichment, named the junction capture method, was developed for the RNA panel to accurately and cost-effectively detect 365 fusion genes as well as aberrantly spliced transcripts. The TOP RNA panel can also measure the expression profiles of an additional 109 genes. The TOP DNA panel was developed to detect single nucleotide variants and insertions/deletions for 464 genes, to calculate tumor mutation burden and microsatellite instability status, and to infer chromosomal copy number. Clinically relevant somatic mutations were identified in 32.2% (59/183) of patients by prospective TOP testing, signifying the clinical utility of TOP for providing personalized medicine to cancer patients.
Assuntos
Perfilação da Expressão Gênica , Neoplasias/genética , Transcriptoma , Processamento Alternativo , Biomarcadores Tumorais , Biópsia , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Neoplasias/diagnóstico , Neoplasias/metabolismo , Proteínas de Fusão Oncogênica/genética , Sequenciamento Completo do GenomaRESUMO
Recent studies have demonstrated that tumor-driving alterations are often different among gliomas that originated from different brain regions and have underscored the importance of analyzing molecular characteristics of gliomas stratified by brain region. Therefore, to elucidate molecular characteristics of diffuse cerebellar gliomas (DCGs), 27 adult, mostly glioblastoma cases were analyzed. Comprehensive analysis using whole-exome sequencing, RNA sequencing, and Infinium methylation array (n = 17) demonstrated their distinct molecular profile compared to gliomas in other brain regions. Frequent mutations in chromatin-modifier genes were identified including, noticeably, a truncating mutation in SETD2 (n = 4), which resulted in loss of H3K36 trimethylation and was mutually exclusive with H3F3A K27M mutation (n = 3), suggesting that epigenetic dysregulation may lead to DCG tumorigenesis. Alterations that cause loss of p53 function including TP53 mutation (n = 9), PPM1D mutation (n = 2), and a novel type of PPM1D fusion (n = 1), were also frequent. On the other hand, mutations and copy number changes commonly observed in cerebral gliomas were infrequent. DNA methylation profile analysis demonstrated that all DCGs except for those with H3F3A mutations were categorized in the "RTK I (PDGFRA)" group, and those DCGs had a gene expression signature that was highly associated with PDGFRA. Furthermore, compared with the data of 315 gliomas derived from different brain regions, promoter methylation of transcription factors genes associated with glial development showed a characteristic pattern presumably reflecting their tumor origin. Notably, SOX10, a key transcription factor associated with oligodendroglial differentiation and PDGFRA regulation, was up-regulated in both DCG and H3 K27M-mutant diffuse midline glioma, suggesting their developmental and biological commonality. In contrast, SOX10 was silenced by promoter methylation in most cerebral gliomas. These findings may suggest potential tailored targeted therapy for gliomas according to their brain region, in addition to providing molecular clues to identify the region-related cellular origin of DCGs.
Assuntos
Neoplasias Cerebelares/genética , Neoplasias Cerebelares/metabolismo , Glioma/genética , Glioma/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Cerebelares/patologia , Neoplasias Cerebelares/cirurgia , Cerebelo/diagnóstico por imagem , Cerebelo/metabolismo , Cerebelo/patologia , Cerebelo/cirurgia , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Predisposição Genética para Doença , Glioma/patologia , Glioma/cirurgia , Humanos , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Cancer microenvironment plays a vital role in cancer development and progression, and cancer-stromal interactions have been recognized as important targets for cancer therapy. However, identifying relevant and druggable cancer-stromal interactions is challenging due to the lack of quantitative methods to analyze whole cancer-stromal interactome. RESULTS: We present CASTIN (CAncer-STromal INteractome analysis), a novel framework for the evaluation of cancer-stromal interactome from RNA-Seq data using cancer xenograft models. For each ligand-receptor interaction which is derived from curated protein-protein interaction database, CASTIN summarizes gene expression profiles of cancer and stroma into three evaluation indices. These indices provide quantitative evaluation and comprehensive visualization of interactome, and thus enable to identify critical cancer-microenvironment interactions, which would be potential drug targets. We applied CASTIN to the dataset of pancreas ductal adenocarcinoma, and successfully characterized the individual cancer in terms of cancer-stromal relationships, and identified both well-known and less-characterized druggable interactions. CONCLUSIONS: CASTIN provides comprehensive view of cancer-stromal interactome and is useful to identify critical interactions which may serve as potential drug targets in cancer-microenvironment. CASTIN is available at: http://github.com/tmd-gpat/CASTIN .
Assuntos
Comunicação Celular , Neoplasias/etiologia , Neoplasias/metabolismo , Células Estromais/metabolismo , Microambiente Tumoral , Animais , Comunicação Celular/genética , Linhagem Celular Tumoral , Biologia Computacional , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica/métodos , Xenoenxertos , Humanos , Camundongos , Neoplasias/patologia , Mapeamento de Interação de Proteínas/métodos , Células Estromais/patologia , Transcriptoma , Microambiente Tumoral/genética , Fluxo de TrabalhoRESUMO
Anti-epidermal growth factor receptor (EGFR) treatment is an effective option for metastatic colorectal cancer (CRC) treatment. However, there are few reliable biomarkers to predict the clinical response to anti-EGFR treatment. We investigated the genome-wide DNA methylation status in metastatic colorectal cancer to identify associations between the methylation status and clinical response to anti-EGFR antibody. We retrospectively reviewed the medical records of 97 patients (45 patients for the first cohort and 52 patients for the second cohort) who received anti-EGFR treatment for KRAS wild-type metastatic CRC. Then we analyzed the associations between genome-wide DNA methylation status and clinical response to anti-EGFR treatment, and evaluated the predictive power and value of the methylation status statistically. As a result, each cohort was classified into highly methylated CRC and low methylated CRC subgroups by unsupervised clustering analyses. In the first cohort, clinical outcomes were significantly better in the low methylated CRC subgroup than in the highly methylated CRC subgroup (response rate, 35.7% vs 6.3%, P = 0.03; disease control rate, 75% vs 31.3%, P = 0.005; hazard ratio for progression-free survival, 0.27; 95% confidence interval, 0.13-0.57, P < 0.001; overall survival, 0.19; 95% confidence interval, 0.06-0.54, P < 0.001). These results were reproducible in the second cohort. The genome-wide methylation status was a predictive factor of progression-free survival and overall survival independently of RAS mutation status. In conclusion, we found that the genome-wide DNA methylation status is a powerful epigenetic predictor of anti-EGFR treatment in patients with KRAS wild-type metastatic colorectal cancer (UMIN000005490).
Assuntos
Adenocarcinoma/genética , Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Metilação de DNA/genética , Receptores ErbB/imunologia , Adenocarcinoma/mortalidade , Adulto , Idoso , Biomarcadores Tumorais/análise , Análise por Conglomerados , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/patologia , Intervalo Livre de Doença , Feminino , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Análise de Sequência com Séries de Oligonucleotídeos , Estudos RetrospectivosRESUMO
Whole-exome sequencing (Exome-seq) has been successfully applied in several recent studies. We here sequenced the exomes of 15 pancreatic tumor cell lines and their matched normal samples. We captured 162,073 exons of 16,954 genes and sequenced the targeted regions to a mean coverage of 56-fold. This study identified a total of 1517 somatic mutations and validated 934 mutations by transcriptome sequencing. We detected recurrent mutations in 56 genes. Among them, 41 have not been described. The mutation rates varied widely among cell lines. The diversity of the mutation rates was significantly correlated with the distinct MLH1 copy-number status. Exome-seq revealed intensive genomic instability in a cell line with MLH1 homozygous deletion, indicated by a dramatically elevated rate of somatic substitutions, small insertions/deletions (indels), as well as indels in microsatellites. Notably, we found that MLH1 expression was decreased by nearly half in cell lines with an allelic loss of MLH1. While these cell lines were negative in conventional microsatellite instability assay, they showed a 10.5-fold increase in the rate of somatic indels, e.g., truncating indels in TP53 and TGFBR2, indicating MLH1 haploinsufficiency in the correction of DNA indel errors. We further analyzed the exomes of 15 renal cell carcinomas and confirmed MLH1 haploinsufficiency. We observed a much higher rate of indel mutations in the affected cases and identified recurrent truncating indels in several cancer genes such as VHL, PBRM1, and JARID1C. Together, our data suggest that MLH1 hemizygous deletion, through increasing the rate of indel mutations, could drive the development and progression of sporadic cancers.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Exoma , Instabilidade Genômica , Haploinsuficiência , Proteínas Nucleares/genética , Neoplasias Pancreáticas/genética , Alelos , Linhagem Celular Tumoral , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Perda de Heterozigosidade , Proteína 1 Homóloga a MutL , Mutação , Taxa de Mutação , Reprodutibilidade dos TestesRESUMO
Background: The nucleic acid quality from formalin-fixed paraffin-embedded (FFPE) tumor vary among samples, resulting in substantial variability in the quality of comprehensive cancer genomic profiling tests. The objective of the study is to investigate how nucleic acid quality affects sequencing quality. We also examined the variations in nucleic acid quality among different hospitals or cancer types. Methods: Three nucleic acid quality metrics (ddCq, Q-value, and DV200) and five sequencing quality metrics (on-target rate, mean depth, coverage uniformity, target exon coverage, and coverage of the housekeeping gene) were examined using 585 samples from the Todai OncoPanel, a dual DNA-RNA panel. Results: In the DNA panel, ddCq served as an indicator of sequencing depth and Q-value reflected the uniformity of sequencing across different regions. It was essential to have favorable values not only for ddCq but also for Q-value to obtain ideal sequencing results. For the RNA panel, DV200 proved to be a valuable metric for assessing the coverage of the housekeeping genes. Significant inter-hospital differences were observed for DNA quality (ddCq and Q-value), but not for RNA quality (DV200). Differences were also observed among cancer types, with Q-value being the lowest in lung and the highest in cervix, while DV200 was the highest in lung and the lowest in bowel. Conclusions: We demonstrated distinct characteristics and high predictive performances of ddCq, Q-value, and DV200. Variations were observed in the nucleic acid quality across hospitals and cancer types. Further study is warranted on preanalytical factors in comprehensive cancer genomic profiling tests.
RESUMO
BACKGROUND: Comprehensive cancer genomic profiling tests have recently been used clinically to guide optimal treatment. Currently approved tests use DNA from tissue or plasma samples to analyze a few hundred genes. RNA panels complement DNA panels to detect fusion and exon skipping. METHODS: Between April 2017 and March 2022, we analyzed non-small cell lung cancer samples using Todai OncoPanel, a matched tumor/normal pair panel targeting both DNA and RNA. Publicly available genomic data was downloaded from the Center for Cancer Genomics and Advanced Therapeutics database on 2022/11/3. RESULTS: Sixty non-small cell lung cancer (NSCLC) samples were analyzed. With the DNA panel, 32 samples (53%) had TP53 loss-of-function mutations. Among adenocarcinoma, 17 (33%) had EGFR activating mutations, and 6 (12%) had ERBB2 activating mutations. One BRCA1 and one BRCA2 pathogenic germline variant were also detected. With the RNA panel, 11 fusion genes were detected, all in adenocarcinoma. Specifically, EML4-ALK and KIF5B-RET were detected from one sample each, and 9 others were all novel fusions with unknown pathogenicity. In addition, 4 of 60 (7%) NSCLC samples harbored MET exon 14 skipping. Analysis of the Center for Cancer Genomics and Advanced Therapeutics database found 37 MET exon 14 splice site mutations in 1514 NSCLC samples (2%, p = 0.039). CONCLUSIONS: Analysis of NSCLC with Todai OncoPanel detected many druggable targets. Its RNA panel may detect MET exon 14 skipping with high sensitivity.
RESUMO
The cellular origin of clear cell ovarian carcinoma (CCOC), a major histological subtype of ovarian carcinoma remains elusive. Here, we explored the candidate cellular origin and identify molecular subtypes using integrated genomic/epigenomic analysis. We performed whole exome-sequencing, microarray, and DNA methylation array in 78 CCOC samples according to the original diagnosis. The findings revealed that ARID1A and/or PIK3CA mutations were mutually exclusive with DNA repair related genes, including TP53, BRCA1, and ATM. Clustering of CCOC and other ovarian carcinomas (n = 270) with normal tissues from the fallopian tube, ovarian surface epithelium, endometrial epithelium, and pelvic peritoneum mesothelium (PPM) in a methylation array showed that major CCOC subtypes (with ARID1A and/or PIK3CA mutations) were associated with the PPM-lile cluster (n = 64). This cluster was sub-divided into three clusters: (1) mismatch repair (MMR) deficient with tumor mutational burden-high (n = 2), (2) alteration of ARID1A (n = 51), and (3) ARID1A wild-type (n = 11). The remaining samples (n = 14) were subdivided into (4) ovarian surface epithelium-like (n = 11) and (5) fallopian tube-like (considered as high-grade serous histotype; n = 3). Among these, subtypes (1-3) and others (4 and 5) were found to be associated with immunoreactive signatures and epithelial-mesenchymal transition, respectively. These results contribute to the stratification of CCOC into biological subtypes.
Assuntos
Adenocarcinoma de Células Claras , Metilação de DNA , Proteínas de Ligação a DNA , Mutação , Neoplasias Ovarianas , Fatores de Transcrição , Humanos , Feminino , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Adenocarcinoma de Células Claras/genética , Adenocarcinoma de Células Claras/patologia , Genômica/métodos , Classe I de Fosfatidilinositol 3-Quinases/genética , Epigenômica/métodos , Sequenciamento do Exoma , Pessoa de Meia-IdadeRESUMO
The objective of this study was to clarify the impact of adverse reactions on immune dynamics. We investigated the pattern of systemic adverse reactions after the second and third coronavirus disease 2019 (COVID-19) vaccinations and their relationship with immunoglobulin G against severe acute respiratory syndrome coronavirus 2 spike 1 protein titers, neutralizing antibody levels, peak cellular responses, and the rate of decrease after the third vaccination in a large-scale community-based cohort in Japan. Participants who received a third vaccination with BNT162b2 (Pfizer/BioNTech) or mRNA-1273 (Moderna), had two blood samples, had not had COVID-19, and had information on adverse reactions after the second and third vaccinations (n = 2198) were enrolled. We collected data on sex, age, adverse reactions, comorbidities, and daily medicine using a questionnaire survey. Participants with many systemic adverse reactions after the second and third vaccinations had significantly higher humoral and cellular immunity in the peak phase. Participants with multiple systemic adverse reactions after the third vaccination had small changes in the geometric values of humoral immunity and had the largest geometric mean of cellar immunity in the decay phase. Systemic adverse reactions after the third vaccination helped achieve high peak values and maintain humoral and cellular immunity. This information may help promote uptake of a third vaccination, even among those who hesitate due to adverse reactions.
Assuntos
Vacina BNT162 , COVID-19 , Humanos , Anticorpos Antivirais , Vacina BNT162/efeitos adversos , Terapias Complementares , COVID-19/prevenção & controle , Imunidade Celular , Imunidade Humoral , Vacinação/efeitos adversosRESUMO
Background: Despite accumulating research on the molecular characteristics of meningiomas, no definitive molecularly targeted therapy for these tumors has been established to date. Molecular mechanisms underlying meningioma progression also remain unclear. Comprehensive genetic testing approaches can reveal actionable gene aberrations in meningiomas. However, there is still limited information on whether profiling the molecular status of subsequent recurrent meningiomas could influence the choice of molecular-targeted therapies. Case presentation: We report a case of meningioma with malignant progression and multiple recurrences. We performed matched tumor pair analysis using the Todai OncoPanel to investigate the possibility of additional standard treatments. The loss of several chromosomal regions, including NF2 and CDKN2A, which is associated with aggressive meningiomas, was considered a significant driver event for malignant progression. Using additional matched tumor pair analysis, mutations in TRAF7, ARID1A, and ERBB3 were identified as subclonal driver events at the time of recurrence. No genetic aberrations were found for which evidence-based targeted therapy was applicable. We also reviewed previous reports of molecular therapies in meningioma to discuss issues with the current molecular testing approach. Conclusion: Gene panel testing platforms such as the Todai OncoPanel represent a powerful approach to elucidate actionable genetic alterations in various types of tumors, although their use is still limited to the diagnosis and prediction of prognosis in meningiomas. To enable targeted molecular therapy informed by gene-panel testing, further studies including matched tumor pair analyses are required to understand the molecular characteristics of meningiomas and develop treatments based on genetic abnormalities.
RESUMO
Diabetes is known to increase the risk of nonalcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC). Here we treat male STAM (STelic Animal Model) mice, which develop diabetes, NASH and HCC associated with dysbiosis upon low-dose streptozotocin and high-fat diet (HFD), with insulin or phlorizin. Although both treatments ameliorate hyperglycemia and NASH, insulin treatment alone lead to suppression of HCC accompanied by improvement of dysbiosis and restoration of antimicrobial peptide production. There are some similarities in changes of microflora from insulin-treated patients comorbid with diabetes and NASH. Insulin treatment, however, fails to suppress HCC in the male STAM mice lacking insulin receptor specifically in intestinal epithelial cells (ieIRKO), which show dysbiosis and impaired gut barrier function. Furthermore, male ieIRKO mice are prone to develop HCC merely on HFD. These data suggest that impaired gut insulin signaling increases the risk of HCC, which can be countered by restoration of insulin action in diabetes.