title: Routine genetic testing of lung cancer specimens derived from surgery, bronchoscopy and fluid aspiration by next generation sequencing.
abstract: After the development of EGFR tyrosine kinase inhibitors (TKIs), genetic testing of EGFR became required for effective treatment of lung cancer. Initially, the testing was conducted separately for each mutated region. However, many EGFR mutations have since been identified that determine the efficacy of EGFR-TKIs. Therefore, genetic testing of EGFR by next generation sequencing (NGS) may be a suitable strategy for lung cancer. Here we examined the applicability of the NGS method in regard to sensitivity, time and cost. A total of 939 specimens were obtained from 686 lung cancer patients at our hospital. DNA and RNA were simultaneously extracted from specimens derived from surgery, bronchoscopy, and fluid aspiration. Specimens included cerebrospinal fluid, pleural effusion, abdominal fluid, and pericardial effusion. From RNA, target regions (EGFR, KRAS, ALK fusion and RET fusion) were enriched by RT-PCR and sequenced with MiSeq. From DNA, PCR or PCR-RFLP conventional methods were performed. NGS and conventional methods were carried out routinely per week. Among the total 939 specimens, 38 specimens could not be examined with NGS. Among these, 34 specimens were analyzed by conventional testing with simultaneously extracted DNA. The remaining four specimens could not be tested with either method. Compared with the conventional method, the concordance rate of mutations was 99% (892/901), excluding specimens with NGS failure. The time period required from processing of specimens to results was 4 days, and the cost per sample was sufficiently low. In conclusion, the genetic testing with NGS method was useful for lung cancer treatment. The cost, sensitivity and time were able to tolerate routine examinations.
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Gou
authors.initials: G
authors.lastname: Yamamoto
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Mari
authors.initials: M
authors.lastname: Kikuchi
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Shiho
authors.initials: S
authors.lastname: Kobayashi
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Arai
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Fujiyoshi
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Tomokazu
authors.initials: T
authors.lastname: Wakatsuki
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Miho
authors.initials: M
authors.lastname: Kakuta
authors.affiliation: Department of Thoracic Oncology, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Yamane
authors.affiliation: Department of Thoracic Surgery, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Yoshihito
authors.initials: Y
authors.lastname: Iijima
authors.affiliation: Department of Thoracic Oncology, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Hideaki
authors.initials: H
authors.lastname: Mizutani
authors.affiliation: Department of Thoracic Surgery, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Nakajima
authors.affiliation: Department of Thoracic Oncology, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Junko
authors.initials: J
authors.lastname: Sudo
authors.affiliation: Department of Thoracic Surgery, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Hiroyasu
authors.initials: H
authors.lastname: Kinoshita
authors.affiliation: Department of Thoracic Oncology, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Futoshi
authors.initials: F
authors.lastname: Kurimoto
authors.affiliation: Department of Thoracic Surgery, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Hirohiko
authors.initials: H
authors.lastname: Akiyama
authors.affiliation: Department of Thoracic Surgery, School of Medicine, Kanazawa Medical University, Uchinaka, Kahoku, Ishikawa 920-0293, Japan.
authors.forename: Hidetaka
authors.initials: H
authors.lastname: Uramoto
authors.affiliation: Department of Thoracic Oncology, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Sakai
authors.affiliation: Department of Surgery, Kurume University, Kurume, Fukuoka 830-0011, Japan.
authors.forename: Yoshito
authors.initials: Y
authors.lastname: Akagi
authors.affiliation: Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, Ina, Kitaadachi, Saitama 362-0806, Japan.
authors.forename: Kiwamu
authors.initials: K
authors.lastname: Akagi
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000077548:Anaplastic Lymphoma Kinase; D001999:Bronchoscopy; D066246:ErbB Receptors / Q000235:genetics; D005260:Female; D005820:Genetic Testing; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008175:Lung Neoplasms / Q000175:diagnosis* / Q000235:genetics* / Q000473:pathology / Q000601:surgery; D008297:Male; D008875:Middle Aged; D009154:Mutation; D015514:Oncogene Proteins, Fusion / Q000235:genetics* / Q000302:isolation & purification; D051096:Proto-Oncogene Proteins c-ret / Q000235:genetics; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics
chemical_list: C117307:KRAS protein, human; D015514:Oncogene Proteins, Fusion; C000626173:ALK protein, human; D000077548:Anaplastic Lymphoma Kinase; C512478:EGFR protein, human; D066246:ErbB Receptors; D051096:Proto-Oncogene Proteins c-ret; C099282:RET protein, human; D020794:Receptor Protein-Tyrosine Kinases; D016283:Proto-Oncogene Proteins p21(ras)