Molecular Analysis of Liquid-Based Cytological Specimen Using Virtually Positive Sputum with Adenocarcinoma Cells.

• Material Type: 博士論文
• Title: Molecular Analysis of Liquid-Based Cytological Specimen Using Virtually Positive Sputum with Adenocarcinoma Cells.
• Author/Editor: Nishikawa, Takeshi; Fujii, Tomomi; Tatsumi, Shigenobu; Sugimoto, Aya; Sekita-Hatakeyama, Yoko; Shimada, Keiji; Yamazaki, Masaharu; Hatakeyama, Kinta; Ohbayashi, Chiho
• Author Heading: Nishikawa, Takeshi; Fujii, Tomomi; Tatsumi, Shigenobu; Sugimoto, Aya; Sekita-Hatakeyama, Yoko; Shimada, Keiji; Yamazaki, Masaharu; Hatakeyama, Kinta; Ohbayashi, Chiho
• Publication, Distribution, etc.: MDPI AG.
• Publication Date: 2020-02-05
• Publication Date (W3CDTF): 2020-02-05
• Alternative Title: 液状化細胞診材料を用いた遺伝子解析による腫瘍特異的遺伝子検出感度の検討
• Periodical title: Diagnostics Vol. No.
• No. or year of volume/issue: 10 2
• Volume: 10
• Issue: 2
• Pages: Article No.84-
• ISSN (Periodical Title): ISSN : 2075-4418
• Degree grantor/type: 奈良県立医科大学
• Date Granted: 2020-06-30
• Dissertation Number: 甲第750号
• Degree Type: 博士（医学）
• Text Language Code: eng
• Subject Heading: lung adenocarcinoma; EGFR mutation; liquid-based cytology; quantitative PCR
• Target Audience: 一般
• Note (General): type:Thesis; Liquid-based cytology (LBC) analysis of sputum is a useful diagnostic and prognostic tool for detecting lung cancer. DNA and RNA derived from lung cancer cells can be used for this diagnosis. However, the quality of cytological material is not always adequate for molecular analysis due to the effect of formalin in the commercially available fixation kits. In this study, we examined DNA and RNA extraction methods for LBC analysis with formalin fixation, using lung carcinoma cell lines and sputum. The human non-small cell lung cancer cell lines were fixed with LBC fixation reagents, such as CytoRich red preservative. Quantification of thyroid transcription factor-1 (TTF-1) and actin mRNA, epidermal growth factor receptor (EGFR) DNA in HCC827, H1975, and H1299 cells, and mutation analysis of EGFR in HCC827 and H1975 cells were performed by quantitative PCR (qPCR) and fluorescence resonance energy transfer (FRET)-based preferential homoduplex formation assay (F-PHFA) method, respectively. mRNA and DNA extracted from cell lines using RNA and/or DNA extraction kits for formalin-fixed paraffin-embedded (FFPE) fixed with various LBC solutions were efficiently detected by qPCR. The detection limit of EGFR mutations was at a rate of 5% mutated positive cells in LBC. The detection limit of the EGFR exon 19 deletion in HCC827 was detected in more than 1.5% of the positive cells in sputum. In contrast, the detection limit of the T790M/L858R mutation in H1975 was detected in more than 13% of the positive cells. We also detected EGFR mutations using next generation sequencing (NGS). The detection limit of NGS for EGFR mutation was lower than that of the F-PHFA method. Furthermore, more than 0.1% of positive cells could be cytomorphologically detected. Our results demonstrate that LBC systems are powerful tools for cytopathological and genetic analyses. However, careful attention should be paid to the incidence of false negative results in the genetic analysis of EGFR mutations detected by LBC.; 博士（医学）・甲第750号・令和2年6月30日; © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).; identifier:Diagnostics Vol.10 No.2 Article No.84 (2020 Feb); identifier:20754418; identifier:http://ginmu.naramed-u.ac.jp/dspace/handle/10564/3757; identifier:Diagnostics Vol. No., 10(2): Article No.84
• Access Restrictions: インターネット公開
• Related Material (DOI): 10.3390/diagnostics10020084
• Data Provider (Database): 国立情報学研究所 : 学術機関リポジトリデータベース（IRDB）（機関リポジトリ）; https://irdb.nii.ac.jp
• Original Data Provider (Database): 奈良県立医科大学 : 奈良県立医科大学機関リポジトリ GINMU

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http://ginmu.naramed-u.ac.jp/dspace/bitstream/10564/3757/1/01%e7%94%b2750%e6%9c%ac%e6%96%87%e3%81%ae%e8%a6%81%e6%97%a8.pdf fulltext

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