PMID- 28882180 OWN - NLM STAT- MEDLINE DCOM- 20180510 LR - 20181202 IS - 1756-9966 (Electronic) IS - 0392-9078 (Print) IS - 0392-9078 (Linking) VI - 36 IP - 1 DP - 2017 Sep 7 TI - High performance of targeted next generation sequencing on variance detection in clinical tumor specimens in comparison with current conventional methods. PG - 121 LID - 10.1186/s13046-017-0591-4 [doi] LID - 121 AB - BACKGROUND: Next generation sequencing (NGS) is being increasingly applied for assisting cancer molecular diagnosis. However, it is still needed to validate NGS accuracy on detection of DNA alternations based on a large number of clinical samples, especially for DNA rearrangements and copy number variations (CNVs). This study is to set up basic parameters of targeted NGS for clinical diagnosis and to understand advantage of targeted NGS in comparison with the conventional methods of molecular diagnosis. METHODS: Genomic DNA from 1000 Genomes Project and DNA from cancer cell lines have been used to establish the basic parameters for targeted NGS. The following confirmation was conducted by clinical samples. The multiple variants tested by amplification-refractory mutation system (ARMS), fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) were evaluated by targeted NGS to determine the sensitivity. Furthermore, the multiple variants detected by targeted NGS were confirmed by current conventional methods to elucidate the specificity. RESULTS: At sequencing depth of 500x, the maximal sensitivities on detecting single nucletic variances (SNVs) and small insertions/deletions (Indels) can reach 99% and 98.7% respectively, and in 20% of cancer cells, CNV detection can reach to the maximal level. The following confirmation of the sensitivity and specificity was conducted by a large cohort of clinical samples. For SNV and indel detection in clinical samples, targeted NGS can identify all hotspot mutations with 100% sensitivity and specificity. On ALK fusion detection, about 86% IHC-identified cases could be identified by targeted NGS and all ALK fusion detected by targeted NGS were confirmed by IHC. For HER2-amplification, 14 HER2-amplification cases identified by target NGS were all confirmed by FISH and about 93.3% of Her-2 IHC (3+) cases were identified by targeted NGS. Finally, the targeted NGS platform developed here has accurately detected EGFR hotspot mutations in 215 NSCLC patients. CONCLUSIONS: DNA from cancer cell lines is better than standard DNA as a reference to establish basic parameters for targeted NGS. Comparison of the conventional methods using a large cohort of patient samples confirmed the high preformance of targeted NGS on detecting DNA alterations. FAU - Su, Dan AU - Su D AD - Pathology Department, Zhejiang Cancer Hospital, Hangzhou, 310022, China. sudan@zjcc.org.cn. AD - Key Laboratory of Diagnosis and Treatment Technology on Thoracic Oncology of Zhejiang Province, Hangzhou, 310022, China. sudan@zjcc.org.cn. FAU - Zhang, Dadong AU - Zhang D AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. AD - Changhai Hospital, The Second Military Medical University, Shanghai, 200433, China. FAU - Chen, Kaiyan AU - Chen K AD - Pathology Department, Zhejiang Cancer Hospital, Hangzhou, 310022, China. AD - Key Laboratory of Diagnosis and Treatment Technology on Thoracic Oncology of Zhejiang Province, Hangzhou, 310022, China. FAU - Lu, Jing AU - Lu J AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. FAU - Wu, Junzhou AU - Wu J AD - Pathology Department, Zhejiang Cancer Hospital, Hangzhou, 310022, China. AD - Key Laboratory of Diagnosis and Treatment Technology on Thoracic Oncology of Zhejiang Province, Hangzhou, 310022, China. FAU - Cao, Xinkai AU - Cao X AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. FAU - Ying, Lisha AU - Ying L AD - Pathology Department, Zhejiang Cancer Hospital, Hangzhou, 310022, China. AD - Key Laboratory of Diagnosis and Treatment Technology on Thoracic Oncology of Zhejiang Province, Hangzhou, 310022, China. FAU - Jin, Qihuang AU - Jin Q AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. AD - Changhai Hospital, The Second Military Medical University, Shanghai, 200433, China. FAU - Ye, Yizhou AU - Ye Y AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. FAU - Xie, Zhenghua AU - Xie Z AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. FAU - Xiong, Lei AU - Xiong L AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. FAU - Mao, Weimin AU - Mao W AD - Pathology Department, Zhejiang Cancer Hospital, Hangzhou, 310022, China. Weimin.mao@zjcc.org.cn. AD - Key Laboratory of Diagnosis and Treatment Technology on Thoracic Oncology of Zhejiang Province, Hangzhou, 310022, China. Weimin.mao@zjcc.org.cn. FAU - Li, Fugen AU - Li F AD - The Research and Development Center of Precision Medicine, 3D Medicine Inc., Shanghai, 201114, China. fugen.li@3dmedcare.com. LA - eng PT - Journal Article DEP - 20170907 PL - England TA - J Exp Clin Cancer Res JT - Journal of experimental & clinical cancer research : CR JID - 8308647 RN - 0 (Oncogene Proteins, Fusion) RN - EC 2.7.10.1 (ALK protein, human) RN - EC 2.7.10.1 (Anaplastic Lymphoma Kinase) RN - EC 2.7.10.1 (ERBB2 protein, human) RN - EC 2.7.10.1 (Receptor Protein-Tyrosine Kinases) RN - EC 2.7.10.1 (Receptor, ErbB-2) SB - IM MH - Anaplastic Lymphoma Kinase MH - DNA Copy Number Variations/*genetics MH - Female MH - Gene Rearrangement/*genetics MH - High-Throughput Nucleotide Sequencing MH - Human Genome Project MH - Humans MH - INDEL Mutation/*genetics MH - Immunohistochemistry MH - In Situ Hybridization, Fluorescence MH - Male MH - Neoplasms/*genetics/pathology MH - Oncogene Proteins, Fusion/genetics MH - Receptor Protein-Tyrosine Kinases/genetics MH - Receptor, ErbB-2/genetics PMC - PMC5590190 OTO - NOTNLM OT - Amplification-refractory mutation system OT - Clinical tumor samples OT - Fluorescence in situ hybridization OT - Immunohistochemistry OT - Targeted next generation sequencing COIS- COMPETING INTERESTS: All authors with 3D medicine affiliation are current or former employees. PUBLISHER'S NOTE: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. EDAT- 2017/09/09 06:00 MHDA- 2018/05/11 06:00 PMCR- 2017/09/07 CRDT- 2017/09/09 06:00 PHST- 2017/06/28 00:00 [received] PHST- 2017/08/30 00:00 [accepted] PHST- 2017/09/09 06:00 [entrez] PHST- 2017/09/09 06:00 [pubmed] PHST- 2018/05/11 06:00 [medline] PHST- 2017/09/07 00:00 [pmc-release] AID - 10.1186/s13046-017-0591-4 [pii] AID - 591 [pii] AID - 10.1186/s13046-017-0591-4 [doi] PST - epublish SO - J Exp Clin Cancer Res. 2017 Sep 7;36(1):121. doi: 10.1186/s13046-017-0591-4.