title: Detection of mutations in EGFR in circulating lung-cancer cells.
abstract: BACKGROUND
  The use of tyrosine kinase inhibitors to target the epidermal growth factor receptor gene (EGFR) in patients with non-small-cell lung cancer is effective but limited by the emergence of drug-resistance mutations. Molecular characterization of circulating tumor cells may provide a strategy for noninvasive serial monitoring of tumor genotypes during treatment.
  
  
  METHODS
  We captured highly purified circulating tumor cells from the blood of patients with non-small-cell lung cancer using a microfluidic device containing microposts coated with antibodies against epithelial cells. We performed EGFR mutational analysis on DNA recovered from circulating tumor cells using allele-specific polymerase-chain-reaction amplification and compared the results with those from concurrently isolated free plasma DNA and from the original tumor-biopsy specimens.
  
  
  RESULTS
  We isolated circulating tumor cells from 27 patients with metastatic non-small-cell lung cancer (median number, 74 cells per milliliter). We identified the expected EGFR activating mutation in circulating tumor cells from 11 of 12 patients (92%) and in matched free plasma DNA from 4 of 12 patients (33%) (P=0.009). We detected the T790M mutation, which confers drug resistance, in circulating tumor cells collected from patients with EGFR mutations who had received tyrosine kinase inhibitors. When T790M was detectable in pretreatment tumor-biopsy specimens, the presence of the mutation correlated with reduced progression-free survival (7.7 months vs. 16.5 months, P<0.001). Serial analysis of circulating tumor cells showed that a reduction in the number of captured cells was associated with a radiographic tumor response; an increase in the number of cells was associated with tumor progression, with the emergence of additional EGFR mutations in some cases.
  
  
  CONCLUSIONS
  Molecular analysis of circulating tumor cells from the blood of patients with lung cancer offers the possibility of monitoring changes in epithelial tumor genotypes during the course of treatment.
authors.affiliation: Massachusetts General Hospital Cancer Center, Boston 02129, USA.
authors.forename: Shyamala
authors.initials: S
authors.lastname: Maheswaran
authors.forename: Lecia V
authors.initials: LV
authors.lastname: Sequist
authors.forename: Sunitha
authors.initials: S
authors.lastname: Nagrath
authors.forename: Lindsey
authors.initials: L
authors.lastname: Ulkus
authors.forename: Brian
authors.initials: B
authors.lastname: Brannigan
authors.forename: Chey V
authors.initials: CV
authors.lastname: Collura
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Inserra
authors.forename: Sven
authors.initials: S
authors.lastname: Diederichs
authors.forename: A John
authors.initials: AJ
authors.lastname: Iafrate
authors.forename: Daphne W
authors.initials: DW
authors.lastname: Bell
authors.forename: Subba
authors.initials: S
authors.lastname: Digumarthy
authors.forename: Alona
authors.initials: A
authors.lastname: Muzikansky
authors.forename: Daniel
authors.initials: D
authors.lastname: Irimia
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Settleman
authors.forename: Ronald G
authors.initials: RG
authors.lastname: Tompkins
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Lynch
authors.forename: Mehmet
authors.initials: M
authors.lastname: Toner
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Haber
mesh_terms: D000328:Adult; D000368:Aged; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D004252:DNA Mutational Analysis / Q000379:methods; D004273:DNA, Neoplasm / Q000032:analysis*; D018450:Disease Progression; D016903:Drug Monitoring / Q000379:methods*; D005260:Female; D018773:Genes, erbB-1; D005819:Genetic Markers; D005838:Genotype; D006801:Humans; D053208:Kaplan-Meier Estimate; D008175:Lung Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D008297:Male; D046210:Microfluidic Analytical Techniques; D008875:Middle Aged; D009154:Mutation; D009360:Neoplastic Cells, Circulating; D021141:Nucleic Acid Amplification Techniques; D016016:Proportional Hazards Models; D047428:Protein Kinase Inhibitors / Q000627:therapeutic use
chemical_list: D004273:DNA, Neoplasm; D005819:Genetic Markers; D047428:Protein Kinase Inhibitors