title: Single-molecule detection of epidermal growth factor receptor mutations in plasma by microfluidics digital PCR in non-small cell lung cancer patients.
abstract: OBJECTIVE
  We aim to develop a digital PCR-based method for the quantitative detection of the two common epidermal growth factor receptor (EGFR) mutations (in-frame deletion at exon 19 and L858R at exon 21) in the plasma and tumor tissues of patients suffering from non-small cell lung cancers. These two mutations account for >85% of clinically important EGFR mutations associated with responsiveness to tyrosine kinase inhibitors.
  
  
  METHODS
  DNA samples were analyzed using a microfluidics system that simultaneously performed 9,180 PCRs at nanoliter scale. A single-mutant DNA molecule in a clinical specimen could be detected and the quantities of mutant and wild-type sequences were precisely determined.
  
  
  RESULTS
  Exon 19 deletion and L858R mutation were detectable in 6 (17%) and 9 (26%) of 35 pretreatment plasma samples, respectively. When compared with the sequencing results of the tumor samples, the sensitivity and specificity of plasma EGFR mutation analysis were 92% and 100%, respectively. The plasma concentration of the mutant sequences correlated well with the clinical response. Decreased concentration was observed in all patients with partial or complete clinical remission, whereas persistence of mutation was observed in a patient with cancer progression. In one patient, tyrosine kinase inhibitor was stopped after an initial response and the tumor-associated EGFR mutation reemerged 4 weeks after stopping treatment.
  
  
  CONCLUSIONS
  The sensitive detection and accurate quantification of low abundance EGFR mutations in tumor tissues and plasma by microfluidics digital PCR would be useful for predicting treatment response, monitoring disease progression and early detection of treatment failure associated with acquired drug resistance.
authors.affiliation: State Key Laboratory in Oncology in South China, Sir Y.K. Pao Centre for Cancer, Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong Special Administrative Region, People's Republic of China.
authors.forename: Tony K F
authors.initials: TK
authors.lastname: Yung
authors.forename: K C Allen
authors.initials: KC
authors.lastname: Chan
authors.forename: Tony S K
authors.initials: TS
authors.lastname: Mok
authors.forename: Joanna
authors.initials: J
authors.lastname: Tong
authors.forename: Ka-Fai
authors.initials: KF
authors.lastname: To
authors.forename: Y M Dennis
authors.initials: YM
authors.lastname: Lo
mesh_terms: D002289:Carcinoma, Non-Small-Cell Lung / Q000097:blood / Q000235:genetics*; D066246:ErbB Receptors / Q000235:genetics*; D006801:Humans; D008175:Lung Neoplasms / Q000097:blood / Q000235:genetics*; D046210:Microfluidic Analytical Techniques; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology
chemical_list: D047428:Protein Kinase Inhibitors; D066246:ErbB Receptors