title: Droplet Digital PCR for Absolute Quantification of EML4-ALK Gene Rearrangement in Lung Adenocarcinoma.
abstract: Crizotinib treatment significantly prolongs progression-free survival, increases response rates, and improves the quality of life in patients with ALK-positive non-small-cell lung cancer. Droplet Digital PCR (ddPCR), a recently developed technique with high sensitivity and specificity, was used in this study to evaluate the association between the abundance of ALK rearrangements and crizotinib effectiveness. FFPE tissues were obtained from 103 consecutive patients with lung adenocarcinoma. Fluorescent in situ hybridization (FISH) and ddPCR were performed. The results revealed that 14 (13.6%) of the 103 patients were positive by dual-color, break-apart FISH. Three variants (1, 2, and 3) of the EML4-ALK gene rearrangements were detected. Thirteen of 14 ALK-positive cases identified by FISH were confirmed by ddPCR (four with variant 1, two with variant 2, and seven with variant 3). The case missed by ddPCR was identified as KIF5B-ALK gene rearrangement by PCR-based direct sequencing. Sixteen patients were detected with low copy numbers of EML4-ALK gene rearrangement, which failed to meet the positive cutoff point of FISH. Two of them responded well to crizotinib after unsuccessful chemotherapy. Our study indicates that ddPCR can be used as a molecular analytical tool to accurately measure the EML4-ALK rearrangement copy numbers in FFPE samples of lung adenocarcinoma patients.
authors.affiliation: Department of Pathology, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, China.
authors.forename: Qiushi
authors.initials: Q
authors.lastname: Wang
authors.affiliation: Department of Pathology, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Yang
authors.affiliation: Department of Respiration, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: He
authors.affiliation: Department of Pathology, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Ma
authors.affiliation: Department of Pathology, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, China.
authors.forename: Li
authors.initials: L
authors.lastname: Lin
authors.affiliation: Department of Pathology, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, China.
authors.forename: Ping
authors.initials: P
authors.lastname: Fu
authors.affiliation: Department of Pathology, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, China. Electronic address: dpbl_xhl@126.com.
authors.forename: Hualiang
authors.initials: H
authors.lastname: Xiao
mesh_terms: D000230:Adenocarcinoma / Q000175:diagnosis / Q000188:drug therapy / Q000235:genetics*; D000077192:Adenocarcinoma of Lung; D000328:Adult; D000368:Aged; D000970:Antineoplastic Agents / Q000627:therapeutic use; D000077547:Crizotinib; D005260:Female; D018628:Gene Dosage; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008175:Lung Neoplasms / Q000175:diagnosis / Q000188:drug therapy / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D015514:Oncogene Proteins, Fusion / Q000032:analysis* / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011379:Prognosis; D011720:Pyrazoles / Q000627:therapeutic use; D011725:Pyridines / Q000627:therapeutic use; D014178:Translocation, Genetic; D016896:Treatment Outcome
chemical_list: D000970:Antineoplastic Agents; C522504:EML4-ALK fusion protein, human; D015514:Oncogene Proteins, Fusion; D011720:Pyrazoles; D011725:Pyridines; D000077547:Crizotinib