Comprehensive assay for the molecular profiling of cancer by target enrichment from formalin‐fixed paraffin‐embedded specimens

Kohsaka, Shinji; Tatsuno, Kenji; Ueno, Toshihide; Nagano, Masaaki; Shinozaki‐Ushiku, Aya; Ushiku, Tetsuo; Takai, Daiya; Ikegami, Masachika; Kobayashi, Hiroshi; Kage, Hidenori; Ando, Mizuo; Hata, Katsuya; Ueda, Hiroki; Yamamoto, Shozo; Kojima, Shinya; Oseto, Kumiko; Akaike, Keisuke; Suehara, Yoshiyuki; Hayashi, Takuo; Saito, Tsuyoshi; Takahashi, Fumiyuki; Takahashi, Kazuhisa; Takamochi, Kazuya; Suzuki, Kenji; Nagayama, Satoshi; Oda, Yoshinao; Mimori, Koshi; Ishihara, Soichiro; Yatomi, Yutaka; Nagase, Takahide; Nakajima, Jun; Tanaka, Sakae; Fukayama, Masashi; Oda, Katsutoshi; Nangaku, Masaomi; Miyazono, Kohei; Miyagawa, Kiyoshi; Aburatani, Hiroyuki; Mano, Hiroyuki

doi:10.1111/cas.13968

Cited by 55 publications

(68 citation statements)

References 47 publications

(71 reference statements)

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“…Although current standard clinical practice does not support the routine use of genomic/transcriptomic testing in early-stage, reliable prognostic biomarkers may be helpful to select patients who are at high-risk of recurrence and who might benefit from additional systemic therapies. We have indeed included the three genes in the clinical RNA sequencing panel that we designed 49 and plan to investigate their utility to guide decisions on adjuvant systemic therapy in a prospective LUAD cohort.…”

Section: Discussionmentioning

confidence: 99%

Identification of Novel CD74-NRG2α Fusion From Comprehensive Profiling of Lung Adenocarcinoma in Japanese Never or Light Smokers

Hayashi

Nagano

Ueno³

et al. 2020

Journal of Thoracic Oncology

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Introduction: Studies are yet to characterize the differences in molecular profiles of lung adenocarcinoma (LUAD) among divergent ethnic groups. Herein, we conducted comprehensive molecular profiling of LUAD in never or light smokers from Asia to discover novel targetable mutations and prognostic biomarkers of this distinct disease entity. Methods: We analyzed 996 cases of Japanese LUAD and performed whole-exome sequencing and RNA-seq in 125 cases of Japanese LUAD negative for the driver oncogenes defined by conventional laboratory testing. We also investigated the clinical and pathologic characteristics among the 996 cases. Results: Driver oncogenes were identified in 88 cases (70.4%) with specific hotspot mutations differing from those in The Cancer Genome Atlas study. Two actionable novel fusions of FGFR2 and NRG2a were also identified. Clustering on the basis of mRNA expression profiles, but not genetic mutational ones, could predict patient prognosis. The risk score generated by the expression of a three-gene set was a strong prognostic marker for overall survival and progression-free survival in our cohort, and was further validated using The Cancer Genome Atlas cohort. Among the 996 cases, each driver alteration is distributed across all histologic subtypes. Adenocarcinoma in situ was identified to harbor driver mutations, suggesting that these alterations are early events in the pathogenesis of LUAD. ERBB2 mutations were over-represented in young adults. Conclusions: This study indicates the value of applying gene expression profiling for predicting the prognosis after a surgical operation, and that the identification of actionable mutations is important for optimizing targeted drugs in Japanese LUAD.

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Section: Discussionmentioning

confidence: 99%

Identification of Novel CD74-NRG2α Fusion From Comprehensive Profiling of Lung Adenocarcinoma in Japanese Never or Light Smokers

Hayashi

Nagano

Ueno³

et al. 2020

Journal of Thoracic Oncology

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“…In addition to NGS analysis using DNA, RNA analysis allows robust detection of fusion transcripts and exon skipping . We have shown that small biopsy samples are feasible for targeted NGS analysis using RNA extracted from FFPE samples.…”

Section: Discussionmentioning

confidence: 99%

“…Targeted sequencing is considered superior to whole genome or whole exome sequencing in the clinical setting because of higher accuracy and lower costs . From the perspective of the sequencing laboratory, samples are ideally obtained through surgical resection to analyze sufficient amounts of tumor cells and correctly call mutations.…”

Section: Introductionmentioning

confidence: 99%

Small lung tumor biopsy samples are feasible for high quality targeted next generation sequencing

et al. 2019

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Next‐generation sequencing (NGS) has been implemented in clinical oncology to analyze multiple genes and to guide therapy. In patients with advanced lung cancer, small biopsies such as computed tomography‐guided needle biopsy (CTNB), endobronchial ultrasound‐guided transbronchial needle aspiration (EBUS‐TBNA) and transbronchial biopsy (TBB) are less invasive and are preferable to resection to make a pathological diagnosis. However, the quality of DNA/RNA and NGS from small lung tumor biopsy samples is unknown. Between April 2017 and March 2018, 107 consecutive samples were obtained from thoracic tumors or metastatic sites for targeted NGS analysis. Fifteen samples were obtained through CTNB, 11 through EBUS‐TBNA, 11 through TBB and 70 through surgical resection. All samples were formalin‐fixed and paraffin‐embedded. DNA and RNA quality was measured using the ddCq method and the percentage of RNA fragments above 200 nucleotides (DV200), respectively. Our custommade probes were designed to capture exon sequences of 464 cancer‐related genes and transcripts of 463 genes. DNA and RNA yield from the 3 biopsy methods were similar, and less than the yield obtained from resected samples. The quality of DNA and RNA was similar across all methods. Overall, 12 of 15 CTNB samples (80%), all 11 EBUS‐TBNA samples, and 9 of 11 TBB samples (82%) underwent successful NGS assays from DNA. NGS analysis from RNA was successful in all 12 CTNB samples, 9 of 11 EBUS‐TBNA samples (82%), and 8 of 11 TBB samples (73%). CTNB, EBUS‐TBNA and TBB mostly resulted in adequate DNA and RNA quality and enabled high‐quality targeted NGS analysis.

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“…It has been reported that CNVs could directly in uence gene expression and have drastic phenotypic consequences due to altering gene dosage or disrupting coding sequences. [23,24]. This work discovered that ampli cation was the main type of POLR2K change and POLR2K copy number was augmented, which was related to poor survival including overall survival (OS) and disease-free survival (DFS) ( Supplementary Figure 1).…”

Section: Discussionmentioning

confidence: 99%

Gene Expression and Regulatory Webwork of POLR2K in Bladder Carcinogenesis by Integrated Bioinformatics Approaches

Yang

Wang

Guo

et al. 2020

Preprint

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Background：RNA polymerase II subunit K (POLR2K) belongs to one of the multiple subunits of RNA polymerase II (Pol II), whose biological function is to synthesize mRNA. Aberrant POLR2K expression is related to carcinogenesis. However, POLR2K’s underlying role in bladder cancer has not been explored. In the current study, we intend to analyze the function of POLR2K and its regulatory network within bladder cancer.Methods: Public sequencing data was obtained from GEO and TCGA to investigate POLR2K expression and regulatory network within bladder cancer (BLCA) by using GEPIA and Oncomine as well as cBioPortal online tool. LinkedOmics was employed to identify genes displaying significantly differential expression patterns and to perform GO and KEGG analyses. After differential genes was assigned and ranked, GSEA analyses was performed to obtain target networks for transcription factors, miRNAs, and kinases that could regulate POLR2K–associated gene network. Subsequent functional webwork analyses were used to identify cancer-relevant pathways Moreover, POLR2K gene is verified, by ChIP-seq in MCF-7 cell line , with transcription factor binding evidence in the ENCODE Transcription Factor Binding Site Profiles dataset.Conclusions: The current study implies that POLR2K gene is overexpressed and often amplified in BLCA, providing the first evidence that POLR2K deregulation, in particular increased transcription, may promote BLCA. These findings uncover a unique expression patterns of POLR2K and its potential regulatory networks in BLCA, contributing greatly to study of the role of POLR2K in cancer development.

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Comprehensive assay for the molecular profiling of cancer by target enrichment from formalin‐fixed paraffin‐embedded specimens

Cited by 55 publications

References 47 publications

Identification of Novel CD74-NRG2α Fusion From Comprehensive Profiling of Lung Adenocarcinoma in Japanese Never or Light Smokers

Identification of Novel CD74-NRG2α Fusion From Comprehensive Profiling of Lung Adenocarcinoma in Japanese Never or Light Smokers

Small lung tumor biopsy samples are feasible for high quality targeted next generation sequencing

Gene Expression and Regulatory Webwork of POLR2K in Bladder Carcinogenesis by Integrated Bioinformatics Approaches

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