Identification of driver genes contributes to the understanding of cancer etiology and is imperative for the development of individualized therapies. Gene amplification is a major event in ...oncogenesis. Driver genes with tumor-specific amplification-dependent overexpression can be therapeutic targets. In this study, we aimed to identify amplification-dependent driver genes in 1,454 solid tumors, across more than 15 cancer types, by integrative analysis of gene expression and copy number. Amplification-dependent overexpression of 64 known driver oncogenes were found in 587 tumors (40%); genes frequently observed were MYC (25%) and MET (18%) in colorectal cancer; SKP2 (21%) in lung squamous cell carcinoma; HIST1H3B (19%) and MYCN (13%) in liver cancer; KIT (57%) in gastrointestinal stromal tumors; and FOXL2 (12%) in squamous cell carcinoma across tissues. Genomic aberrations in 138 known cancer driver genes and 491 established fusion genes were found in 1,127 tumors (78%). Further analyses of 820 cancer-related genes revealed 16 as potential driver genes, with amplification-dependent overexpression restricted to the remaining 22% of samples (327 tumors) initially undetermined genetic drivers. Among them, AXL, which encodes a receptor tyrosine kinase, was recurrently overexpressed and amplified in sarcomas. Our studies of amplification-dependent overexpression identified potential drug targets in individual tumors.
Mutually exclusive KIT and PDGFRA mutations are considered to be the earliest events in gastrointestinal stromal tumors (GIST), but insufficient for their malignant progression. Herein, we aimed to ...identify driver genes and signaling pathways relevant to GIST progression. We investigated genetic profiles of 707 driver genes, including mutations, gene fusions, copy number gain or loss, and gene expression for 65 clinical specimens of surgically dissected GIST, consisting of six metastatic tumors and 59 primary tumors from stomach, small intestine, rectum, and esophagus. Genetic alterations included oncogenic mutations and amplification‐dependent expression enhancement for oncogenes (OG), and loss of heterozygosity (LOH) and expression reduction for tumor suppressor genes (TSG). We assigned activated OG and inactivated TSG to 27 signaling pathways, the activation of which was compared between malignant GIST (metastasis and high‐risk GIST) and less malignant GIST (low‐ and very low‐risk GIST). Integrative molecular profiling indicated that a greater incidence of genetic alterations of driver genes was detected in malignant GIST (96%, 22 of 23) than in less malignant GIST (73%, 24 of 33). Malignant GIST samples groups showed mutations, LOH, and aberrant expression dominantly in driver genes associated with signaling pathways of PI3K (PIK3CA, AKT1, and PTEN) and the cell cycle (RB1, CDK4, and CDKN1B). Additionally, we identified potential PI3K‐related genes, the expression of which was upregulated (SNAI1 and TPX2) or downregulated (BANK1) in malignant GIST. Based on our observations, we propose that inhibition of PI3K pathway signals might potentially be an effective therapeutic strategy against malignant progression of GIST.
Signaling pathways involved in 65 GIST samples were investigated. Pathways were curated by genetic alterations, including oncogenic mutations and amplification‐dependent expression enhancement for oncogenes, and loss of heterozygosity and expression reduction for tumor suppressor genes. Malignant GIST with metastasis and high‐risk showed mutations, LOH, and aberrant expression dominantly in driver genes associated with signaling pathways of PI3K and the cell cycle.
High-throughput sequencing has greatly contributed to precision medicine. However, challenges remain in reporting secondary findings (SFs) of germline pathogenic variants and managing the affected ...patients. The aim of this study was to examine the incidence of SFs in Japanese cancer patients using whole exome sequencing (WES) and to understand patient preferences regarding SF disclosure. WES was conducted for 2480 cancer patients. Genomic data were screened and classified for variants of 59 genes listed by the American College of Medical Genetics and Genomics SF v2.0 and for an additional 13 hereditary cancer-related genes. Majority of the participants (68.9%; 1709/2480) opted for disclosure of their SFs. Thirty-two pathogenic or likely pathogenic variants, including
BRCA1
(7 patients),
BRCA2
(4),
CHEK2
(4),
PTEN
(3),
MLH1
(3),
SDHB
(2),
MSH6
(1),
NF1
(1),
EXT2
(1),
NF1
(1),
NTRK1
(1),
MYH7
(3),
MYL2
(1),
TNNT2
(1),
LDLR
(2),
FBN1
(1), and
KCNH2
(1) were recognized in 36 patients (1.5%)
.
Twenty-eight (77.8%) patients underwent genetic counseling and received their SF results. Eighteen (64.3%) patients underwent clinical management for SFs. Genetic validation tests were administered significantly more frequently to patients with than without a SF-related personal history (
P
= 0.025). This was a first attempt at a large-scale systematic exome analysis in Japan; nevertheless, many cancer patients opted for disclosure of SFs and accepted or considered clinical management.
Despite the frequent detection of
KRAS
driver mutations in patients with colorectal cancer (CRC), no effective treatments that target mutant KRAS proteins have been introduced into clinical practice. ...In this study, we identified potential effector molecules, based on differences in gene expression between CRC patients carrying wild-type
KRAS
(
n
= 390) and those carrying
KRAS
mutations in codon 12 (
n
= 240). CRC patients with wild-type
KRAS
harboring mutations in
HRAS
,
NRAS
,
PIK3CA
,
PIK3CD
,
PIK3CG
,
RALGDS
,
BRAF,
or
ARAF
were excluded from the analysis. At least 11 promising candidate molecules showed greater than two-fold change between the
KRAS
G12 mutant and wild-type and had a Benjamini-Hochberg-adjusted
P
value of less than 1E-08, evidence of significantly differential expression between these two groups. Among these 11 genes examined in cell lines transfected with
KRAS
G12 mutants,
BMP4
,
PHLDA1,
and
GJB5
showed significantly higher expression level in
KRAS
G12A, G12D, and G12V transfected cells than in the wild-type transfected cells. We expect that this study will lead to the development of novel treatments that target signaling molecules functioning with
KRAS
G12-driven CRC.
This study aimed to establish the Japanese Cancer Genome Atlas (JCGA) using data from fresh frozen tumor tissues obtained from 5143 Japanese cancer patients, including those with colorectal cancer ...(31.6%), lung cancer (16.5%), gastric cancer (10.8%) and other cancers (41.1%). The results are part of a single‐center study called “High‐tech Omics‐based Patient Evaluation” or “Project HOPE” conducted at the Shizuoka Cancer Center, Japan. All DNA samples and most RNA samples were analyzed using whole‐exome sequencing, cancer gene panel sequencing, fusion gene panel sequencing and microarray gene expression profiling, and the results were annotated using an analysis pipeline termed “Shizuoka Multi‐omics Analysis Protocol” developed in‐house. Somatic driver alterations were identified in 72.2% of samples in 362 genes (average, 2.3 driver events per sample). Actionable information on drugs that is applicable in the current clinical setting was associated with 11.3% of samples. When including those drugs that are used for investigative purposes, actionable information was assigned to 55.0% of samples. Germline analysis revealed pathogenic mutations in hereditary cancer genes in 9.2% of samples, among which 12.2% were confirmed as pathogenic mutations by confirmatory test. Pathogenic mutations associated with non–cancerous hereditary diseases were detected in 0.4% of samples. Tumor mutation burden (TMB) analysis revealed 5.4% of samples as having the hypermutator phenotype (TMB ≥ 20). Clonal hematopoiesis was observed in 8.4% of samples. Thus, the JCGA dataset and the analytical procedures constitute a fundamental resource for genomic medicine for Japanese cancer patients.
The present study aims to establish the Japanese Cancer Genome Atlas (JCGA) by analyzing fresh frozen tumor tissues obtained from 5143 Japanese cancer patients. Somatic driver and druggable alterations were detected in 72.2% and 11.3% of samples, respectively, and germline pathogenic mutations in hereditary cancer genes were identified in 9.2% of samples. The JCGA dataset and analytical procedures constitute a fundamental resource for genomic medicine for Japanese cancer patients.
Tumor mutational burden (TMB) and mutational signatures reflect the process of mutation accumulation in cancer. However, the significance of these emerging characteristics remains unclear. In the ...present study, we used whole‐exome sequencing to analyze the TMB and mutational signature in solid tumors of 4046 Japanese patients. Eight predominant signatures—microsatellite instability, smoking, POLE, APOBEC, UV, mismatch repair, double‐strand break repair, and Signature 16—were observed in tumors with TMB higher than 1.0 mutation/Mb, whereas POLE and UV signatures only showed moderate correlation with TMB, suggesting the extensive accumulation of mutations due to defective POLE and UV exposure. The contribution ratio of Signature 16, which is associated with hepatocellular carcinoma in drinkers, was increased in hypopharynx cancer. Tumors with predominant microsatellite instability signature were potential candidates for treatment with immune checkpoint inhibitors such as pembrolizumab and were found in 2.8% of cases. Furthermore, based on microarray analysis, tumors with predominant signatures were classified into 2 subgroups depending on the expression of immune‐related genes reflecting differences in the immune context of the tumor microenvironment. Tumor subpopulations differing in the content of infiltrating immune cells might respond differently to immunotherapeutics. An understanding of cancer characteristics based on TMB and mutational signatures could provide new insights into mutation‐driven tumorigenesis.
By applying whole‐exome sequencing and microarray technologies for high‐throughput genomic and transcriptional analysis, this study investigates the mutational burden and signatures in a large cohort of tumor samples of different histology from Japanese patients. The contribution ratio of Signature 16, which is associated with hepatocellular carcinoma in drinkers, was increased in hypopharynx cancer. Furthermore, based on microarray analysis, tumors with predominant signatures were classified into 2 subgroups depending on the expression of immune‐related genes, reflecting differences in the immune context of the tumor microenvironment.
The detection of copy number variations (CNVs) and somatic mutations in cancer is important for the selection of specific drugs for patients with cancer. In cancers with sporadic tumor cells, low ...tumor content prevents the accurate detection of somatic alterations using targeted sequencing. To efficiently identify CNVs, we performed tumor cell enrichment using tissue suspensions of formalin-fixed paraffin-embedded (FFPE) tissue sections with low tumor cell content. Tumor-enriched and residual fractions were separated from FFPE tissue suspensions of intestinal and diffuse-type gastric cancers containing sporadic tumor cells, and targeted sequencing was performed on 225 cancer-related genes. Sequencing of a targeted panel of cancer-related genes using tumor-enriched fractions increased the number of detectable CNVs and the copy number of amplified genes. Furthermore, CNV analysis using the normal cell-enriched residual fraction as a reference for CNV scoring allowed targeted sequencing to detect CNV characteristics of diffuse-type gastric cancer with low tumor content. Our approach improves the CNV detection rate in targeted sequencing with tumor enrichment and the accuracy of CNV detection in archival samples without paired blood.
Tumor mutational burden analysis using whole‐exome sequencing highlights features of tumors with various mutations or known driver alterations. Cancers with few changes in the exon regions have ...unclear characteristics, even though low‐mutated tumors are often detected in pan‐cancer analysis. In the present study, we analyzed tumors with low tumor mutational burden listed in the Japanese version of The Cancer Genome Atlas, a data set of 5020 primary solid tumors. Our analysis revealed that detection rates of known driver mutations and copy number variation were decreased in samples with tumor mutational burden below 1.0 (ultralow tumor), compared with those in samples with low tumor mutational burden (≤5 mutations/Mb). This trend was also observed in The Cancer Genome Atlas data set. In the ultralow tumor mutational burden tumors, expression analysis showed decreased TP53 inactivation and chromosomal instability. TP53 inactivation frequently correlated with PI3K/mTOR‐related gene expression, implying suppression of the PI3K/mTOR pathway in ultralow tumor mutational burden tumors. In common with mutational burden, the T cell‐inflamed gene expression profiling signature was a potential marker for prediction of an immune checkpoint inhibitor response, and some ultralow tumor mutational burden tumor populations highly expressed this signature. Our analysis focused on how these tumors could provide insight into tumors with low somatic alteration that are difficult to detect solely using whole‐exome sequencing.
This study describes our analysis of tumors with a low tumor burden level in the Japanese version of The Cancer Genome Atlas (JCGA), a data set of 5020 primary solid tumors. In the ultralow tumor mutational burden tumors, expression analysis showed decreased TP53 inactivation and chromosomal instability. Our analysis focuses on how these tumors can provide insight into tumors with low somatic alteration that are difficult to detect solely by whole exome sequencing.
Lung adenocarcinoma is the most common histological type of lung cancer, and its incidence is increasing worldwide. To identify genetic factors influencing risk of lung adenocarcinoma, we conducted a ...genome-wide association study and two validation studies in the Japanese population comprising a total of 6,029 individuals with lung adenocarcinoma (cases) and 13,535 controls. We confirmed two previously reported risk loci, 5p15.33 (rs2853677, P(combined) = 2.8 × 10(-40), odds ratio (OR) = 1.41) and 3q28 (rs10937405, P(combined) = 6.9 × 10(-17), OR = 1.25), and identified two new susceptibility loci, 17q24.3 (rs7216064, P(combined) = 7.4 × 10(-11), OR = 1.20) and 6p21.3 (rs3817963, P(combined) = 2.7 × 10(-10), OR = 1.18). These data provide further evidence supporting a role for genetic susceptibility in the development of lung adenocarcinoma.
Tumor mutational burden (TMB) is an emerging characteristic in cancer and has been associated with microsatellite instability, defective DNA replication/repair, and response to PD-1 and PD-L1 ...blockade immunotherapy. When estimating TMB, targeted panel sequencing is performed using a few hundred genes; however, a comparison of TMB results obtained with this platform and with whole exome sequencing (WES) has not been performed for various cancer types. In the present study, we compared TMB results using the above two platforms in 2,908 solid tumors that were obtained from Japanese patients. For next-generation sequencing, we used fresh-frozen tissue specimens. The Ion Proton System was employed to detect somatic mutations in the coding genome and to sequence an available cancer panel that targeted 409 genes. We then selected 2,040 samples with sufficient tumor cellularity for TMB analysis. In tumors with TMB-high (TMB ≥ 20 mutations/Mb), TMB derived from WES correlated well with the estimated TMB (eTMB) based on panel sequencing, whereas TMB in the remaining tumors showed a weak correlation. In particular, eTMB was overestimated in tumors with low-frequency mutations, resulting in the accumulation of EGFR mutations not being discriminated as a feature of lung cancer with low-frequency mutations. The eTMB in tumors harboring POLE mutations and microsatellite instability was not overestimated, suggesting that panel sequencing could accurately estimate TMB in tumors with high-frequency mutations such as hypermutator tumors. These results may provide helpful information for interpreting TMB results based on clinical sequencing using a targeted gene panel.
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