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MOTIVATION: Spaln is the earliest practical tool for self-sufficient genome mapping and spliced alignment of protein query sequences onto a mammalian-sized eukaryotic genomic sequence. However, its computational speed has become inadequate for the analysis of rapidly growing genomic and transcript sequence data. RESULTS: The dynamic programming calculation of Spaln has been sped up in two ways: 1) the introduction of the multi-intermediate unidirectional Hirschberg method and 2) SIMD-based vectorization. The new version, Spaln3, is approximately seven times faster than the latest Spaln version 2, and its gene prediction accuracy is consistently higher than that of Miniprot. AVAILABILITY AND IMPLEMENTATION: Https://github.com/ogotoh/spaln. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
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BACKGROUND: Tailored, preventive cancer care requires the identification of pathogenic germline variants (PGVs) among potentially at-risk blood relatives (BRs). Cascade testing is carried out for BRs of probands who are positive for PGVs of an inherited cancer but not for negative probands. This study was conducted to examine the prevalence of PGVs for BRs of PGV-negative probands. METHODS: PGV prevalence was assessed for 682 BRs of 281 probands with BRCA1/BRCA2 wild-type hereditary breast and ovarian cancer (HBOC) syndrome. RESULTS: PGVs were discovered in 22 (45.8%) of the 48 BRs of the PGV-positive probands and in 14 (2.2%) of 634 BRs of the PGV-negative probands. Eleven PGVs on high-risk BRCA1, BRCA2, and TP53 genes were present only in BRs and not in the probands (probands vs BRs in Fisher exact test; p = 0.0104; odds ratio [OR] = 0.000 [0.000-0.5489 of 95% confidence interval]), partly due to the nature of the selection criteria. The enrichment of high-risk PGVs among BRs was also significant as compared with a non-cancer East Asian population (p = 0.0016; OR = 3.0791 [1.5521-5.6694]). PGV prevalence, risk class of gene, and genotype concordance were unaffected by the cancer history among BRs. CONCLUSION: These findings imply the necessity to construct a novel testing scheme to complement cascade testing.
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The hyperplasia-carcinoma sequence is a stepwise tumourigenic programme towards endometrial cancer in which normal endometrial epithelium becomes neoplastic through non-atypical endometrial hyperplasia (NAEH) and atypical endometrial hyperplasia (AEH), under the influence of unopposed oestrogen. NAEH and AEH are known to exhibit polyclonal and monoclonal cell growth, respectively; yet, aside from focal PTEN protein loss, the genetic and epigenetic alterations that occur during the cellular transition remain largely unknown. We sought to explore the potential molecular mechanisms that promote the NAEH-AEH transition and identify molecular markers that could help to differentiate between these two states. We conducted target-panel sequencing on the coding exons of 596 genes, including 96 endometrial cancer driver genes, and DNA methylome microarrays for 48 NAEH and 44 AEH lesions that were separately collected via macro- or micro-dissection from the endometrial tissues of 30 cases. Sequencing analyses revealed acquisition of the PTEN mutation and the clonal expansion of tumour cells in AEH samples. Further, across the transition, alterations to the DNA methylome were characterised by hypermethylation of promoter/enhancer regions and CpG islands, as well as hypo- and hyper-methylation of DNA-binding regions for transcription factors relevant to endometrial cell differentiation and/or tumourigenesis, including FOXA2, SOX17, and HAND2. The identified DNA methylation signature distinguishing NAEH and AEH lesions was reproducible in a validation cohort with modest discriminative capability. These findings not only support the concept that the transition from NAEH to AEH is an essential step within neoplastic cell transformation of endometrial epithelium but also provide deep insight into the molecular mechanism of the tumourigenic programme. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
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Carcinoma Endometrioide , Metilação de DNA , Hiperplasia Endometrial , Neoplasias do Endométrio , Epigênese Genética , PTEN Fosfo-Hidrolase , Feminino , Humanos , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Carcinoma Endometrioide/genética , Carcinoma Endometrioide/patologia , PTEN Fosfo-Hidrolase/genética , Hiperplasia Endometrial/genética , Hiperplasia Endometrial/patologia , Hiperplasia Endometrial/metabolismo , Lesões Pré-Cancerosas/genética , Lesões Pré-Cancerosas/patologia , Mutação , Regulação Neoplásica da Expressão Gênica , Pessoa de Meia-Idade , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Ilhas de CpG/genética , IdosoRESUMO
Importance: Neoadjuvant chemoradiotherapy (CRT) is the standard of care for advanced rectal cancer. Yet, estimating response to CRT remains an unmet clinical challenge. Objective: To investigate and better understand the transcriptomic factors associated with response to neoadjuvant CRT and survival in patients with advanced rectal cancer. Design, Setting, and Participants: A single-center, retrospective, case series was conducted at a comprehensive cancer center. Pretreatment biopsies from 298 patients with rectal cancer who were later treated with neoadjuvant CRT between April 1, 2004, and September 30, 2020, were analyzed by RNA sequencing. Data analysis was performed from July 1, 2021, to May 31, 2022. Exposures: Chemoradiotherapy followed by total mesorectal excision or watch-and-wait management. Main Outcomes and Measures: Transcriptional subtyping was performed by consensus molecular subtype (CMS) classification. Immune cell infiltration was assessed using microenvironment cell populations-counter (MCP-counter) scores and single-sample gene set enrichment analysis (ssGSEA). Patients with surgical specimens of tumor regression grade 3 to 4 or whose care was managed by the watch-and-wait approach for more than 3 years were defined as good responders. Results: Of the 298 patients in the study, 205 patients (68.8%) were men, and the median age was 61 (IQR, 52-67) years. Patients classified as CMS1 (6.4%) had a significantly higher rate of good response, albeit survival was comparable among the 4 subtypes. Good responders exhibited an enrichment in various immune-related pathways, as determined by ssGSEA. Microenvironment cell populations-counter scores for cytotoxic lymphocytes were significantly higher for good responders than nonresponders (median, 0.76 [IQR, 0.53-1.01] vs 0.58 [IQR, 0.43-0.83]; P < .001). Cytotoxic lymphocyte MCP-counter score was independently associated with response to CRT, as determined in the multivariable analysis (odds ratio, 3.81; 95% CI, 1.82-7.97; P < .001). Multivariable Cox proportional hazards regression analysis, including postoperative pathologic factors, revealed the cytotoxic lymphocyte MCP-counter score to be independently associated with recurrence-free survival (hazard ratio [HR], 0.38; 95% CI, 0.16-0.92; P = .03) and overall survival (HR, 0.16; 95% CI, 0.03-0.83; P = .03). Conclusions and Relevance: In this case series of patients with rectal cancer treated with neoadjuvant CRT, the cytotoxic lymphocyte score in pretreatment biopsy samples, as computed by RNA sequencing, was associated with response to CRT and survival. This finding suggests that the cytotoxic lymphocyte score might serve as a biomarker in personalized multimodal rectal cancer treatment.
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Antineoplásicos , Neoplasias Retais , Masculino , Humanos , Pessoa de Meia-Idade , Feminino , Terapia Neoadjuvante , Resultado do Tratamento , Estudos Retrospectivos , Transcriptoma , Neoplasias Retais/genética , Neoplasias Retais/terapia , Neoplasias Retais/patologia , Biópsia , Microambiente Tumoral/genéticaRESUMO
Helicobacter pylori (HP) is a major etiologic driver of diffuse-type gastric cancer (DGC). However, improvements in hygiene have led to an increase in the prevalence of HP-naïve DGC; that is, DGC that occurs independent of HP. Although multiple genomic cohort studies for gastric cancer have been conducted, including studies for DGC, distinctive genomic differences between HP-exposed and HP-naïve DGC remain largely unknown. Here, we employed exome and RNA sequencing with immunohistochemical analyses to perform binary comparisons between 36 HP-exposed and 27 HP-naïve DGCs from sporadic, early-stage, and intramucosal or submucosal tumor samples. Among the samples, 33 HP-exposed and 17 HP-naïve samples had been preserved as fresh-frozen samples. HP infection status was determined using stringent criteria. HP-exposed DGCs exhibited an increased single nucleotide variant burden (HP-exposed DGCs; 1.97 [0.48-7.19] and HP-naïve DGCs; 1.09 [0.38-3.68] per megabase; p = 0.0003) and a higher prevalence of chromosome arm-level aneuploidies (p < 0.0001). CDH1 was mutated at similar frequencies in both groups, whereas the RHOA-ARHGAP pathway misregulation was exclusive to HP-exposed DGCs (p = 0.0167). HP-exposed DGCs showed gains in chromosome arms 8p/8q (p < 0.0001), 7p (p = 0.0035), and 7q (p = 0.0354), and losses in 16q (p = 0.0167). Immunohistochemical analyses revealed a higher expression of intestinal markers such as CD10 (p < 0.0001) and CDX2 (p = 0.0002) and a lower expression of the gastric marker, MUC5AC (p = 0.0305) among HP-exposed DGCs. HP-naïve DGCs, on the other hand, had a purely gastric marker phenotype. This work reveals that HP-naïve and HP-exposed DGCs develop along different molecular pathways, which provide a basis for early detection strategies in high incidence settings. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
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Infecções por Helicobacter , Helicobacter pylori , Neoplasias Gástricas , Mucosa Gástrica/patologia , Genômica , Infecções por Helicobacter/complicações , Helicobacter pylori/genética , Humanos , Nucleotídeos/metabolismo , Neoplasias Gástricas/patologiaRESUMO
BACKGROUND: In cancer therapy, higher-resolution tumor-agnostic biomarkers that predict response to immune checkpoint inhibitor (ICI) therapy are needed. Mutation signatures reflect underlying oncogenic processes that can affect tumor immunogenicity, and thus potentially delineate ICI treatment response among tumor types. METHODS: Based on mutational signature analysis, we developed a stratification for all solid tumors in The Cancer Genome Atlas (TCGA). Subsequently, we developed a new software (Genomic Subtyping and Predictive Response Analysis for Cancer Tumor ICi Efficacy, GS-PRACTICE) to classify new tumors submitted to whole-exome sequencing. Using existing data from 973 pan-cancer ICI-treated cases with outcomes, we evaluated the subtype-response predictive performance. RESULTS: Systematic analysis on TCGA samples identified eight tumor genomic subtypes, which were characterized by features represented by smoking exposure, ultraviolet light exposure, APOBEC enzyme activity, POLE mutation, mismatch repair deficiency, homologous recombination deficiency, genomic stability, and aging. The former five subtypes were presumed to form an immune-responsive group acting as candidates for ICI therapy because of their high expression of immune-related genes and enrichment in cancer types with FDA approval for ICI monotherapy. In the validation cohort, the samples assigned by GS-PRACTICE to the immune-reactive subtypes were significantly associated with ICI response independent of cancer type and TMB high or low status. CONCLUSIONS: The new tumor subtyping method can serve as a tumor-agnostic biomarker for ICI response prediction and will improve decision making in cancer treatment.
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Neoplasias , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Genômica , Humanos , Mutação , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/patologia , Sequenciamento do Exoma/métodosRESUMO
In order to identify genomic biomarkers for the outcome of mogamulizumab-containing treatment, an integrated molecular analysis of adult T-cell leukemia/lymphoma (ATL) was conducted on 64 mogamulizumab-naïve patients. Among driver genes, CCR4 and CCR7 alterations were observed in 22% and 11% of the patients, respectively, both consisting of single nucleotide variants (SNV)/insertion-deletions (indels) in the C-terminus. Patients with CCR4 alterations or without CCR7 alterations exhibited a more favorable clinical response (complete response [CR] rate 93%, 13/14; P=0.024, and CR rate 71%, 40/56; P=0.036, respectively). Additionally, TP53, CD28, and CD274 alterations were identified in 35%, 16%, and 10% of the patients, respectively. TP53 alterations included SNV/indels or copy number variations (CNV) such as homozygous deletion; CD28 alterations included SNV, CNV such as amplification, or fusion; CD274 alterations included CNV such as amplification, or structural variants. Univariate analysis revealed that TP53, CD28 or CD274 alterations were associated with worse overall survival (OS) (hazard ratio [HR]: 2.330, 95% confidence interval [CI]: 1.183-4.589; HR: 3.191, 95% CI: 1.287- 7.911; HR: 3.301, 95% CI: 1.130-9.641, respectively) but that CCR4 alterations were associated with better OS (HR: 0.286, 95% CI: 0.087-0.933). Multivariate analysis indicated that in addition to performance status, TP53, CCR4 or CD274 alterations (HR: 2.467, 95% CI: 1.197-5.085; HR: 0.155, 95% CI: 0.031-0.778; HR: 14.393, 95% CI: 2.437-85.005, respectively) were independently and significantly associated with OS. The present study contributes to the establishment of precision medicine using mogamulizumab in ATL patients.
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Leucemia-Linfoma de Células T do Adulto , Linfoma , Adulto , Anticorpos Monoclonais Humanizados , Antígenos CD28 , Variações do Número de Cópias de DNA , Genômica , Homozigoto , Humanos , Leucemia-Linfoma de Células T do Adulto/tratamento farmacológico , Leucemia-Linfoma de Células T do Adulto/genética , Leucemia-Linfoma de Células T do Adulto/patologia , Nucleotídeos , Receptores CCR7 , Deleção de Sequência , Resultado do TratamentoRESUMO
Aim: Gene set analysis has commonly been used to interpret DNA methylome data. However, summarizing the DNA methylation level of a gene is challenging due to variability in the number, density and methylation levels of CpG sites, and the numerous intergenic CpGs. Instead, we propose to use region sets to annotate the DNA methylome. Methods: We developed single sample region-set enrichment analysis for DNA methylome (methyl-ssRSEA) to conduct sample-wise, region-set enrichment analysis. Results: Methyl-ssRSEA can handle both microarray- and sequencing-based platforms and reproducibly recover the known biology from the methylation profiles of peripheral blood cells and breast cancers. The performance was superior to existing tools for region-set analysis in discriminating blood cell types. Conclusion: Methyl-ssRSEA offers a novel way to functionally interpret the DNA methylome in the cell.
Lay abstract Gene set analysis has been a common way to understand the meaning of DNA methylome data. However, organizing the DNA methylation level of a gene is challenging due to variation in the number, density and extent of methylation, of methylation sites, and the substantial number of methylation sites between genes. Instead, we propose to use region sets for the organization. We developed single sample region-set enrichment analysis for DNA methylome (methyl-ssRSEA) to conduct region-set analysis for every sample. Methyl-ssRSEA can handle both microarray- and sequencing-based methods and repeatedly find the known characters from the methylation patterns of peripheral blood cells and breast cancers. The performance was better than existing tools for region-set analysis in differentiating blood cell types. Methyl-ssRSEA offers a novel way to find the features of DNA methylome in the cell.
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Metilação de DNA , Epigênese Genética , Epigenômica/métodos , Transcriptoma , Algoritmos , Biomarcadores , Células Sanguíneas/metabolismo , Biologia Computacional/métodos , Ilhas de CpG , Perfilação da Expressão Gênica/métodos , Humanos , Anotação de Sequência MolecularRESUMO
There are numerous histological subtypes (histotypes) of gynecological malignancies, with each histotype considered to largely reflect a feature of the "cell of origin," and to be tightly linked with the clinical behavior and biological phenotype of the tumor. The recent advances in massive parallel sequencing technologies have provided a more complete picture of the range of the genomic alterations that can persist within individual tumors, and have highlighted the types and frequencies of driver-gene mutations and molecular subtypes often associated with these histotypes. Several large-scale genomic cohorts, including the Cancer Genome Atlas (TCGA), have been used to characterize the genomic features of a range of gynecological malignancies, including high-grade serous ovarian carcinoma, uterine corpus endometrial carcinoma, uterine cervical carcinoma, and uterine carcinosarcoma. These datasets have also been pivotal in identifying clinically relevant molecular targets and biomarkers, and in the construction of molecular subtyping schemes. In addition, the recent widespread use of clinical sequencing for the more ubiquitous types of gynecological cancer has manifested in a series of large genomic datasets that have allowed the characterization of the genomes, driver mutations, and histotypes of even rare cancer types, with sufficient statistical power. Here, we review the field of gynecological cancer, and seek to describe the genomic features by histotype. We also will demonstrate how these are linked with clinicopathological attributes and highlight the potential tumorigenic mechanisms.
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Neoplasias dos Genitais Femininos/genética , Genômica , Mutação , Carcinogênese/genética , Feminino , HumanosRESUMO
Genes involved in the homologous recombination repair pathway-as exemplified by BRCA1, BRCA2, PALB2, ATM, and CHEK2-are frequently associated with hereditary breast and ovarian cancer syndrome. Germline mutations in the loci of these genes with loss of heterozygosity or additional somatic truncation at the WT allele lead to the development of breast cancers with characteristic clinicopathological features and prominent genomic features of homologous recombination deficiency, otherwise referred to as "BRCAness." Although clinical genetic testing for these and other genes has increased the chances of identifying pathogenic variants, there has also been an increase in the prevalence of variants of uncertain significance, which poses a challenge to patient care because of the difficulties associated with making further clinical decisions. To overcome this challenge, we sought to develop a methodology to reclassify the pathogenicity of these unknown variants using statistical modeling of BRCAness. The model was developed with Lasso logistic regression by comparing 116 genomic attributes derived from 37 BRCA1/2 biallelic mutant and 32 homologous recombination-quiescent breast cancer exomes. The model showed 95.8% and 86.7% accuracies in the training cohort and The Cancer Genome Atlas validation cohort, respectively. Through application of the model for variant reclassification of homologous recombination-associated hereditary breast and ovarian cancer causal genes and further assessment with clinicopathological features, we finally identified one likely pathogenic and five likely benign variants. As such, the BRCAness model developed from the tumor exome was robust and provided a reasonable basis for variant reclassification.
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Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Predisposição Genética para Doença , Recombinação Homóloga , Modelos Genéticos , Adulto , Idoso , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteína BRCA1/genética , Proteína BRCA2/genética , Mama/patologia , Mama/cirurgia , Neoplasias da Mama/patologia , Neoplasias da Mama/cirurgia , Quinase do Ponto de Checagem 2/genética , Análise Mutacional de DNA , Conjuntos de Dados como Assunto , Exoma/genética , Proteína do Grupo de Complementação N da Anemia de Fanconi/genética , Feminino , Testes Genéticos/métodos , Mutação em Linhagem Germinativa , Humanos , Mastectomia , Pessoa de Meia-Idade , Sequenciamento do ExomaRESUMO
Emerging evidence suggests that an increased density of pre-treatment CD8+ tumor-infiltrating lymphocytes (TILs) is associated with good response to chemoradiotherapy (CRT) in patients with locally advanced rectal cancer. However, the significance of T-cell complexity in the clinical setting remains unknown. High-throughput T-cell receptor (TCR) ß sequencing was applied to quantify the TCR repertoire of pre-treatment biopsies from 67 patients with advanced rectal cancer receiving preoperative CRT. Diversity index was used to represent the complexity of the TCR repertoire in a tumor. Pre-treatment CD8+ TIL densities were assessed by immunohistochemistry. Changes in TCR repertoire before and after CRT were also analysed in 23 patients. Diversity indices were significantly higher for good responders than for non-responders (P = 0.031). The multivariate analysis revealed that both CD8+ TIL density and TCR diversity index were independently associated with good response to CRT (P < 0.001 and P = 0.049, respectively). Patients who were high for both CD8+ TIL density and TCR diversity (double-high) had markedly better responses to CRT than double-low patients (84.2% vs 16.7%, P < 0.0001). Larger changes in TCR repertoires before and after CRT were correlated with better recurrence-free survival (P = 0.027). The complexity and dynamic change in the TCR repertoire might serve as a useful indicator of response to CRT in combination with CD8+ TIL density in patients with rectal cancer.
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Quimiorradioterapia/métodos , Terapia Neoadjuvante/métodos , Neoplasias Retais/tratamento farmacológico , Neoplasias Retais/radioterapia , Linfócitos T/metabolismo , Feminino , Humanos , MasculinoRESUMO
Microsatellite instability (MSI) is categorized by mutation frequency: high MSI (MSI-H), low MSI (MSI-L) and microsatellite stable (MSS). MSI-H tumors have a distinct immunogenic phenotype, with immunotherapies using checkpoint inhibitors already approved for the treatment of MSI-H gastroesophageal adenocarcinoma (GEA); this is not observed for MSI-L or MSS. Here, we tested the hypothesis that MSI-L tumors are also a distinct phenotype and potentially immunogenic. MSI-PCR assays (BAT25, BAT26, BAT40, D2S123, D5S346 and D17S250) were performed on 363 Epstein-Barr virus-negative, surgically resected esophagogastric junction (EGJ) adenocarcinoma samples. Tumors were characterized as MSI-H (≥2 markers), MSI-L (1 marker) or MSS (0 markers). CD8+ cell counts, PD-L1 and HER2 expression levels, TP53 mutations, epigenetic alterations and prognostic significance were also examined. All pathological and molecular experiments were conducted using serial, whole-tumor sections of chemo-naïve surgical specimens. MSI-H and MSI-L were assigned to 28 (7.7%) and 24 (6.6%) cases, respectively. Compared to MSS cases, MSI-L cases had significantly higher intratumoral CD8+ cell infiltration (P = .048) and favorable EGJ cancer-specific survival (multivariate hazard ratio = 0.35, 95% CI, 0.12-0.82; P = .012). MSI-L tumors were also significantly associated with TP53-truncating mutations as compared to MSI-H (P = .009) and MSS (P = .012) cases, and this trend was also observed in GEA data from The Cancer Genome Atlas (TCGA). Indel mutational burden among TCGA MSI-L tumors was significantly higher than that of MSS tumors (P = .016). These results suggest that MSI-L tumors may have a distinct tumor phenotype and be potentially immunogenic in EGJ adenocarcinoma.
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Adenocarcinoma/imunologia , Neoplasias Esofágicas/imunologia , Junção Esofagogástrica , Instabilidade de Microssatélites , Adenocarcinoma/genética , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Idoso , Idoso de 80 Anos ou mais , Infecções por Vírus Epstein-Barr/complicações , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/mortalidade , Neoplasias Esofágicas/patologia , Feminino , Genes p53 , Humanos , Masculino , Pessoa de Meia-Idade , MutaçãoRESUMO
Gene-structure-aware multiple sequence alignment (GSA-MSA) is conventionally used as a tool for analyzing evolutionary changes in gene structure, i.e., gain and loss of introns during the course of evolution of homologous eukaryotic genes. Recently, however, it has become apparent that GSA-MSA is a powerful tool for detecting and remedying gene-prediction errors prevalent in genome annotations produced by various genome projects. Unfortunately, the construction of GSA-MSAs has so far required tedious procedures, thereby preventing researchers from enjoying the potential benefits of GSA-MSAs. In this chapter, we introduce a straightforward way for constructing GSA-MSAs when one or more genomic sequences and a set of transcript sequences (protein or full-length cDNAs/CDSs) are given. Our method requires no external tool or extra data, such as annotation files, although a supplementary script can generate a gene-structure-informed (GSI) transcript sequence file from annotation files.
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Mapeamento Cromossômico/métodos , Genômica/métodos , Splicing de RNA/genética , Alinhamento de Sequência/métodos , Software , Algoritmos , Sequência de Aminoácidos , DNA Complementar/genética , Bases de Dados Genéticas , Íntrons , FilogeniaRESUMO
OBJECTIVE: Carcinosarcoma (CS) of the uterus or ovary is a rare, biphasic tumor comprising epithelial and mesenchymal elements, and exhibits more aggressive clinical features than its carcinoma counterpart. Four molecular subtypes of CS were recently established based on genomic aberration profiles (POLE, MSI, CNH, and CNL) and shown to be associated with multiple clinicopathological parameters, including patient outcomes. However, the role of the immune microenvironment in CS remains unclear. Here, we investigated the influence of the immune cells that infiltrate CS to better understand the immunological status of gynecological CS. METHODS: Tumor immune microenvironmental analyses on CS samples were performed using immune cell profiling with RNA-seq, transcriptomic subtyping with microenvironmental genes, and T-cell receptor repertoire assay. Carcinoma and sarcoma elements from CS samples were also assessed separately. RESULTS: Relying on estimations of tumor-infiltrating cell types from RNA-seq data, POLE and MSI (hypermutator) tumors showed an enrichment of M1 macrophages, plasma cells and CD8+ T cells, whereas CNH and CNL (non-hypermutator) tumors had high levels of M2 macrophages. Further subclassification by immune-related, non-cancer genes identified a fraction of tumors with distinct patient outcomes, particularly those with the CNH genomic aberration subtype. T-cell heterogeneity was independently correlated with prolonged progression-free survival. Differential analysis of carcinoma and sarcoma elements identified many shared mutations but there was little overlap in the T-cell receptor repertoire between the two elements. CONCLUSIONS: Tumor immune microenvironmental analyses could offer potential clinical utility in the stratification of gynecological CS above classification by genomic aberration subtype alone.
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Carcinossarcoma/genética , Neoplasias Ovarianas/genética , Receptores de Antígenos de Linfócitos T/genética , Microambiente Tumoral/imunologia , Neoplasias Uterinas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinossarcoma/imunologia , Carcinossarcoma/patologia , Estudos de Coortes , Biologia Computacional , Feminino , Heterogeneidade Genética , Humanos , Linfócitos do Interstício Tumoral , Pessoa de Meia-Idade , Mutação , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Ovário/imunologia , Ovário/patologia , Prognóstico , RNA-Seq , Microambiente Tumoral/genética , Neoplasias Uterinas/imunologia , Neoplasias Uterinas/patologia , Útero/imunologia , Útero/patologia , Sequenciamento do ExomaRESUMO
Panel sequencing of susceptibility genes for hereditary breast and ovarian cancer (HBOC) syndrome has uncovered numerous germline variants; however, their pathogenic relevance and ethnic diversity remain unclear. Here, we examined the prevalence of germline variants among 568 Japanese patients with BRCA1/2-wildtype HBOC syndrome and a strong family history. Pathogenic or likely pathogenic variants were identified on 12 causal genes for 37 cases (6.5%), with recurrence for 4 SNVs/indels and 1 CNV. Comparisons with non-cancer east-Asian populations and European familial breast cancer cohorts revealed significant enrichment of PALB2, BARD1, and BLM mutations. Younger onset was associated with but not predictive of these mutations. Significant somatic loss-of-function alterations were confirmed on the wildtype alleles of genes with germline mutations, including PALB2 additional somatic truncations. This study highlights Japanese-associated germline mutations among patients with BRCA1/2 wildtype HBOC syndrome and a strong family history, and provides evidence for the medical care of this high-risk population.
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DNA fragmentation is a fundamental step during library preparation in hybridization capture-based, short-read sequencing. Ultra-sonication has been used thus far to prepare DNA of an appropriate size, but this method is associated with a considerable loss of DNA sample. More recently, studies have employed library preparation methods that rely on enzymatic fragmentation with DNA endonucleases to minimize DNA loss, particularly in nano-quantity samples. Yet, despite their wide use, the effect of enzymatic fragmentation on the resultant sequences has not been carefully assessed. Here, we used pairwise comparisons of somatic variants of the same tumor DNA samples prepared using ultrasonic and enzymatic fragmentation methods. Our analysis revealed a substantially larger number of recurrent artifactual SNVs/indels in endonuclease-treated libraries as compared with those created through ultrasonication. These artifacts were marked by palindromic structure in the genomic context, positional bias in sequenced reads, and multi-nucleotide substitutions. Taking advantage of these distinctive features, we developed a filtering algorithm to distinguish genuine somatic mutations from artifactual noise with high specificity and sensitivity. Noise cancelling recovered the composition of the mutational signatures in the tumor samples. Thus, we provide an informatics algorithm as a solution to the sequencing errors produced as a consequence of endonuclease-mediated fragmentation, highlighted for the first time in this study.
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Artefatos , Fragmentação do DNA , Biblioteca Gênica , Análise de Sequência de DNARESUMO
Endometrial endometrioid adenocarcinoma (EEA) is conventionally considered to be a single pathologic entity that develops through a hyperplasia-carcinoma sequence under the influence of estrogen. Previously, another EEA subtype was described and proposed to arise directly from normal endometrium. These conventional and de novo subtypes are designated groups 1 and 2, respectively. To identify the molecular mechanisms of these distinct tumorigenic processes, we conducted comprehensive integrated analyses of genomic data with hormonal status for group 1 paired carcinoma and hyperplasia and group 2 carcinoma samples. Although group 1 carcinomas mostly exhibited genomically stable characteristics and the activation of estrogen signaling, group 2 EEAs showed enriched hypermutator and CpG island methylator phenotypes. Pairwise comparisons of hyperplasia and carcinoma, along with time-course analyses of the hyperplasia-carcinoma sequence, revealed the acquisition of driver mutations in the evolutionary process of hyperplasia but not in the transition from hyperplasia to carcinoma. The current study confirms the existence of two different histopathologic programs during EEA development that harbor distinct molecular bases and demonstrates the biological relevance of these differential tumorigenic processes.
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Adenocarcinoma/patologia , Biomarcadores Tumorais/genética , Carcinogênese/patologia , Carcinoma Endometrioide/patologia , Neoplasias do Endométrio/classificação , Neoplasias do Endométrio/patologia , Regulação Neoplásica da Expressão Gênica , Adenocarcinoma/genética , Carcinogênese/genética , Carcinoma Endometrioide/genética , Estudos de Casos e Controles , Hiperplasia Endometrial/genética , Hiperplasia Endometrial/patologia , Neoplasias do Endométrio/genética , Endométrio/metabolismo , Endométrio/patologia , Epigênese Genética , Feminino , Seguimentos , Perfilação da Expressão Gênica , Genômica , Humanos , Mutação , Prognóstico , Receptores de Estrogênio/metabolismo , TranscriptomaRESUMO
Carcinosarcoma (CS) of the uterus or ovary is a rare, aggressive and biphasic neoplasm composed of carcinoma and sarcoma elements. Previous genomic studies have identified the driver genes and genomic properties associated with CS. However, there is still no molecular subtyping scheme with clinical relevance for this disease. Here, we sequence 109 CS samples, focusing on 596 genes. We identify four molecular subtypes that resemble those observed in endometrial carcinoma: POLE-mutated, microsatellite instability, copy number high, and copy number low subtypes. These molecular subtypes are linked with DNA repair deficiencies, potential therapeutic strategies, and multiple clinicopathological features, including patient outcomes. Multi-regional comparative sequencing reveals genomic alteration-independent CS cell differentiation. Transcriptome and DNA methylome analyses confirm epithelial-mesenchymal transition as a mechanism of sarcoma differentiation. The current study thus provides therapeutic possibilities for CS as well as clues to understanding the molecular histogenic mechanism of its development.
Assuntos
Carcinossarcoma/genética , Neoplasias Ovarianas/genética , Neoplasias Uterinas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Endometrioide/genética , Carcinoma Endometrioide/patologia , Carcinossarcoma/classificação , Carcinossarcoma/patologia , Análise por Conglomerados , Variações do Número de Cópias de DNA/genética , Metilação de DNA , DNA Polimerase II/genética , Distúrbios no Reparo do DNA/genética , Árvores de Decisões , Transição Epitelial-Mesenquimal/genética , Feminino , Neoplasias dos Genitais Femininos/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mutação INDEL , Instabilidade de Microssatélites , Pessoa de Meia-Idade , Mutação , Neoplasias Císticas, Mucinosas e Serosas/genética , Neoplasias Císticas, Mucinosas e Serosas/patologia , Neoplasias Ovarianas/classificação , Neoplasias Ovarianas/patologia , Neoplasias Peritoneais/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , Polimorfismo de Nucleotídeo Único , RNA Mensageiro/metabolismo , Análise de Sequência de DNA , Transcriptoma , Neoplasias Uterinas/classificação , Neoplasias Uterinas/patologia , Adulto JovemRESUMO
Motivation: Intron length distribution (ILD) is a specific feature of a genome that exhibits extensive species-specific variation. Whereas ILD contributes to up to 30% of the total information content for intron recognition in some species, rendering it an important component of computational gene prediction, very few studies have been conducted to quantitatively characterize ILDs of various species. Results: We developed a set of computer programs (fitild, compild, etc.) to build statistical models of ILDs and compare them with one another. Each ILD of more than 1000 genomes was fitted with fitild to a statistical model consisting of one, two, or three components of Frechet distributions. Several measures of distances between ILDs were calculated by compild. A theoretical model was presented to better understand the origin of the observed shape of an ILD. Availability and implementation: The C++ source codes are available at https://github.com/ogotoh/fitild.git/. Supplementary information: Supplementary data are available at Bioinformatics online.