Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testing
Abstract Comprehensive genomic profiling (CGP) is increasingly used as a clinical laboratory test and being applied to cancer treatment; however, standardization and external quality assessments (EQA) have not been fully developed. This study performed cost-effective EQA and proficiency tests (PT) f...
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| Language: | English |
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Nature Portfolio
2025-01-01
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| Series: | Scientific Reports |
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| Online Access: | https://doi.org/10.1038/s41598-024-84714-4 |
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| author | Kazuyuki Matsushita Takayuki Ishige Kousuke Watanabe Toshiaki Akahane Akihide Tanimoto Michiko Yoshimoto Munekazu Yamakuchi Teruto Hashiguchi Yoshinaga Okugawa Makoto Ikejiri Toshikazu Yamaguchi Tadashi Yamasaki Mayu Takeda Masaaki Hibi Naoki Akiyama Kaho Shimizu Naonori Hashimoto Hiroko Sato Yoshinori Tanaka Fumie Amari the EQA working group of Japan Association for Clinical Laboratory Science(JACLS) |
| author_facet | Kazuyuki Matsushita Takayuki Ishige Kousuke Watanabe Toshiaki Akahane Akihide Tanimoto Michiko Yoshimoto Munekazu Yamakuchi Teruto Hashiguchi Yoshinaga Okugawa Makoto Ikejiri Toshikazu Yamaguchi Tadashi Yamasaki Mayu Takeda Masaaki Hibi Naoki Akiyama Kaho Shimizu Naonori Hashimoto Hiroko Sato Yoshinori Tanaka Fumie Amari the EQA working group of Japan Association for Clinical Laboratory Science(JACLS) |
| author_sort | Kazuyuki Matsushita |
| collection | DOAJ |
| description | Abstract Comprehensive genomic profiling (CGP) is increasingly used as a clinical laboratory test and being applied to cancer treatment; however, standardization and external quality assessments (EQA) have not been fully developed. This study performed cost-effective EQA and proficiency tests (PT) for CGP testing among multiple institutions those belong to the EQA working group of Japan Association for Clinical Laboratory Science (JACLS). This study revealed that preanalytical processes, such as derived nucleic acids (NA) extraction from formalin fixed paraffine embedded (FFPE) samples, are critical. First, EQA with extracted DNA from cell lines showed a detection rate of 100% (9 out of 9) in KRAS (c.38G > A; p.G13D), PIK3CA (p.H1047R), and B-Raf proto-oncogene, serine/threonine kinase (BRAF) (c.1799 T > A; p.V600E) in cases of > 10% variant allele frequency (VAF). However, BRAF (c.1799 T > A; p.V600E) detection decreased to 67% (6 out of 9) for a VAF of 4.9%. Second, when DNA was extracted from FFPE samples, pathogenic variants and variants with companion diagnostic indications were detected in all 10 participating laboratories. Each variant had < 20% VAFs on average (8.1–19.1%) and wide variability among laboratories was observed (relative standard deviation, 13–60%). Nonetheless, BRAF (c.1798_1799delinsAA; p.V600K) of 8.1% VAF, EGFR (c.2235_2249del; p.E746_A750del) of 9.7% VAF, and EGFR (c.2254_2277del; p.S752_I759del) of 9.8% VAF were detected with 70% (7/10), 70% (7/10), and 60% (6/10) frequency, respectively. Therefore, 10% VAF in pre-analytic processing for DNA extraction from FFPE was critical for variant detection in CGP analysis. Further, incorrect results were reported in case independent variant calling of BRAF; c.1798_1799delinsAA (p.V600K) was mistakenly interpreted as c.1798G > A, and c.1799 T > A was on the other strand. In conclusion, the EQA/PT among 10 institutes with common samples revealed the importance of VAF in pre-analysis and helped us understand the significance of the pipeline and common pitfalls usually ignored by the internal quality control in a single institute. |
| format | Article |
| id | doaj-art-6b97909e764a4765ac2e7962d8ce36c4 |
| institution | Kabale University |
| issn | 2045-2322 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj-art-6b97909e764a4765ac2e7962d8ce36c42025-01-12T12:23:20ZengNature PortfolioScientific Reports2045-23222025-01-0115111210.1038/s41598-024-84714-4Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testingKazuyuki Matsushita0Takayuki Ishige1Kousuke Watanabe2Toshiaki Akahane3Akihide Tanimoto4Michiko Yoshimoto5Munekazu Yamakuchi6Teruto Hashiguchi7Yoshinaga Okugawa8Makoto Ikejiri9Toshikazu Yamaguchi10Tadashi Yamasaki11Mayu Takeda12Masaaki Hibi13Naoki Akiyama14Kaho Shimizu15Naonori Hashimoto16Hiroko Sato17Yoshinori Tanaka18Fumie Amari19the EQA working group of Japan Association for Clinical Laboratory Science(JACLS)Department of Laboratory Medicine, Chiba University HospitalDepartment of Laboratory Medicine, Chiba University HospitalDepartment of Clinical Laboratory, The University of TokyoDepartment of Pathology, Graduate School of Medical and Dental Sciences, Kagoshima UniversityDepartment of Pathology, Graduate School of Medical and Dental Sciences, Kagoshima UniversityApplication support development, Global Management Division, Sysmex CorporationDepartment of Laboratory and Vascular Medicine, Graduate School of Medical and Dental Sciences, Kagoshima UniversityDepartment of Laboratory and Vascular Medicine, Graduate School of Medical and Dental Sciences, Kagoshima UniversityDepartment of Genomic Medicine, Mie University HospitalDepartment of Clinical Laboratory, Mie University HospitalBML, Inc.BML, Inc.Department of Medical Research for Intractable Diseases, Fujita Health UniversityAdvanced Medical Care Testing Department, SRL Genomics Division, SRL, Inc.Lab Assay Business, LS Business, Sysmex CorporationLab Assay Business, LS Business, Sysmex CorporationLab Assay Business, LS Business, Sysmex CorporationGenetic Analysis Department, Kawasaki Registered Clinical Laboratory, Riken Genesis Co., Ltd.Bioinformatics Department, Kawasaki Registered Clinical Laboratory, Riken Genesis Co., Ltd.Genetic Analysis Department, Tsukiji Registered Clinical Laboratory, Riken Genesis Co., Ltd.Abstract Comprehensive genomic profiling (CGP) is increasingly used as a clinical laboratory test and being applied to cancer treatment; however, standardization and external quality assessments (EQA) have not been fully developed. This study performed cost-effective EQA and proficiency tests (PT) for CGP testing among multiple institutions those belong to the EQA working group of Japan Association for Clinical Laboratory Science (JACLS). This study revealed that preanalytical processes, such as derived nucleic acids (NA) extraction from formalin fixed paraffine embedded (FFPE) samples, are critical. First, EQA with extracted DNA from cell lines showed a detection rate of 100% (9 out of 9) in KRAS (c.38G > A; p.G13D), PIK3CA (p.H1047R), and B-Raf proto-oncogene, serine/threonine kinase (BRAF) (c.1799 T > A; p.V600E) in cases of > 10% variant allele frequency (VAF). However, BRAF (c.1799 T > A; p.V600E) detection decreased to 67% (6 out of 9) for a VAF of 4.9%. Second, when DNA was extracted from FFPE samples, pathogenic variants and variants with companion diagnostic indications were detected in all 10 participating laboratories. Each variant had < 20% VAFs on average (8.1–19.1%) and wide variability among laboratories was observed (relative standard deviation, 13–60%). Nonetheless, BRAF (c.1798_1799delinsAA; p.V600K) of 8.1% VAF, EGFR (c.2235_2249del; p.E746_A750del) of 9.7% VAF, and EGFR (c.2254_2277del; p.S752_I759del) of 9.8% VAF were detected with 70% (7/10), 70% (7/10), and 60% (6/10) frequency, respectively. Therefore, 10% VAF in pre-analytic processing for DNA extraction from FFPE was critical for variant detection in CGP analysis. Further, incorrect results were reported in case independent variant calling of BRAF; c.1798_1799delinsAA (p.V600K) was mistakenly interpreted as c.1798G > A, and c.1799 T > A was on the other strand. In conclusion, the EQA/PT among 10 institutes with common samples revealed the importance of VAF in pre-analysis and helped us understand the significance of the pipeline and common pitfalls usually ignored by the internal quality control in a single institute.https://doi.org/10.1038/s41598-024-84714-4External quality assessment (EQA)Proficiency testing (PT)Genetic variantsComprehensive cancer genome testingNGS |
| spellingShingle | Kazuyuki Matsushita Takayuki Ishige Kousuke Watanabe Toshiaki Akahane Akihide Tanimoto Michiko Yoshimoto Munekazu Yamakuchi Teruto Hashiguchi Yoshinaga Okugawa Makoto Ikejiri Toshikazu Yamaguchi Tadashi Yamasaki Mayu Takeda Masaaki Hibi Naoki Akiyama Kaho Shimizu Naonori Hashimoto Hiroko Sato Yoshinori Tanaka Fumie Amari the EQA working group of Japan Association for Clinical Laboratory Science(JACLS) Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testing Scientific Reports External quality assessment (EQA) Proficiency testing (PT) Genetic variants Comprehensive cancer genome testing NGS |
| title | Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testing |
| title_full | Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testing |
| title_fullStr | Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testing |
| title_full_unstemmed | Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testing |
| title_short | Importance of EQA/PT for the detection of genetic variants in comprehensive cancer genome testing |
| title_sort | importance of eqa pt for the detection of genetic variants in comprehensive cancer genome testing |
| topic | External quality assessment (EQA) Proficiency testing (PT) Genetic variants Comprehensive cancer genome testing NGS |
| url | https://doi.org/10.1038/s41598-024-84714-4 |
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