title: A diagnostic algorithm using EGFR mutation-specific antibodies for rapid response EGFR-TKI treatment in patients with non-small cell lung cancer.
abstract: BACKGROUND
  We previously reported that EGFR mutation-specific antibodies performed well in immunohisto/cytochemical analysis. Assessment of EGFR mutation status for EGFR-TKIs (tyrosin kinase inhibitors) treatment of non-small cell lung cancer (NSCLC) patients can be performed by DNA-based assay. To establish a testing algorithm for EGFR mutation status in NSCLC patients, we utilized the peptide nucleic acid-locked nucleic acid (PNA-LNA) PCR clamp assay to determine the EGFR mutation, and immunostaining to detect the delE746-A750 and L858R mutation protein.
  
  
  METHODS
  One hundred thirty-three patients with NSCLC were examined by immunostaining with EGFR mutation-specific antibodies, and by the PNA-LNA PCR clamp assay, using histological or cytological samples. The expression levels of immunostaining were classified as positive (score 2+), negative (score 0) or equivocal (score 1+), indicating questionable, negative or weak expression. Of these patients, 42 NSCLC patients received EGFR-TKIs treatment and we analyzed whether expression of mutant EGFR proteins was correlated with progression-free survival in patients with NSCLC treated with EGFR-TKIs.
  
  
  RESULTS
  In the 133 samples, positive, negative and equivocal results for EGFR mutation-specific antibodies were observed in 37 patients (27.8%), 85 patients (63.9%) and 11 patients (8.3%), respectively. Thirty-seven patients showed positive EGFR expression by immunostaining, and 35 (94.6%) of these also tested positive in the DNA-based assay. Of 85 EGFR-negative patients by immunostaining, 77 (90.6%) also tested negative in the DNA-based assay. Of the 11 patients with equivocal immunostaining results, DNA-based assay detected EGFR mutation in 8 (72.7%) patients. Immunostaining for the EGFR mutation-specific antibodies showed a sensitivity of 81.4%, specificity of 97.5%, positive predictive value of 94.6% and negative predictive value of 90.6%, excluding the 11 patients with equivocal results. In NSCLC patients treated with EGFR-TKIs, the progression-free survival after the start of EGFR-TKIs treatment was significantly longer in patients with EGFR positive expression than in those with equivocal and negative expression (P=0.002).
  
  
  CONCLUSIONS
  Our results suggest that patients testing positive for EGFR mutations by immunostaining are good candidates for rapid response EGFR-TKI therapy. Immunostaining for EGFR mutation-specific antibodies is a new test for EGFR-TKI inhibition and could be a useful indicator with regard to patient management.
authors.affiliation: Department of Diagnostic Pathology, Kurume University Hospital, Kurume, Japan. akihiko4@med.kurume-u.ac.jp
authors.forename: Akihiko
authors.initials: A
authors.lastname: Kawahara
authors.forename: Tomoki
authors.initials: T
authors.lastname: Taira
authors.forename: Koichi
authors.initials: K
authors.lastname: Azuma
authors.forename: Masaki
authors.initials: M
authors.lastname: Tominaga
authors.forename: Satoshi
authors.initials: S
authors.lastname: Hattori
authors.forename: Mayuko
authors.initials: M
authors.lastname: Kawahara
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Abe
authors.forename: Tomohiko
authors.initials: T
authors.lastname: Yamaguchi
authors.forename: Jun
authors.initials: J
authors.lastname: Akiba
authors.forename: Shinzo
authors.initials: S
authors.lastname: Takamori
authors.forename: Akihiro
authors.initials: A
authors.lastname: Hayashi
authors.forename: Masayoshi
authors.initials: M
authors.lastname: Kage
mesh_terms: D000465:Algorithms; D000906:Antibodies; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D002289:Carcinoma, Non-Small-Cell Lung / Q000175:diagnosis / Q000188:drug therapy* / Q000235:genetics / Q000401:mortality; D018572:Disease-Free Survival; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000276:immunology; D005260:Female; D006801:Humans; D008175:Lung Neoplasms / Q000175:diagnosis / Q000188:drug therapy* / Q000235:genetics / Q000401:mortality; D008297:Male; D009154:Mutation; D009367:Neoplasm Staging; D047428:Protein Kinase Inhibitors / Q000627:therapeutic use*
chemical_list: D000906:Antibodies; D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D066246:ErbB Receptors