Correlation of EGFR Immunohistochemistry and Molecular Analysis in Lung Adenocarcinoma
Eisenstein Jeffrey, Guillory Tesha, Cohen Cynthia. Emory University, Atlanta, GA
Background: Lung adenocarcinomas with epidermal growth factor receptor (EGFR) mutations portend increased survival and response to targeted therapy. 90% of mutations that respond to gefitnib and erlotinib are delE746-A750 in exon 19 (del 19) and L858R in exon 21 (L858R). Accepted method of analysis is PCR amplification and sequencing of exons 18 through 21, performed by reference laboratories. Monoclonal antibodies are available against the two common mutations; studies show that immunohistochemistry (IHC) correlates with mutational analysis on cytology specimens. The aims of our study are: 1) to augment the number of cases evaluated in the literature; 2) to validate stains for our laboratory; and 3) to evaluate use of IHC as screening for mutational analysis referral.
Design: Of 444 specimens with EGFR mutational analysis from 2010-2012, we identified 54 mutated cases with paraffin blocks for staining. 58 known wild-types were selected as negatives. Two rabbit monoclonal Abs with specificities for 15 bp, E746-A750 deletion in exon 19, and exon 21 L858R (Cell Signaling Technology, Inc., Danvers, MA) were used at 1:50 dilution with high pH antigen retrieval. IHC slides were blindly reviewed and staining was scored: 0 (none); 1+ (faint membrane/cytoplasm); 2+ (moderate membrane); 3+ (intense/crisp membrane). Cutoff for positivity was >10% of tumor cells staining 1+ or any amount of 2+ or 3+.
Results: Fifty-four known EGFR mutations by molecular analysis included: twenty-four (del 19); twenty-one (L858R); nine (various mutations including insertion 20 and T790M). By IHC, four false negatives were identified in the del 19 group; zero in the L858R group and three in the various mutation group. Results of 58 known wild type (no mutations by molecular analysis) included: one false positive (del 19); two false positives (L858R).
|PCR - EGFR Mutated (54)||PCR- EGFR WT (58)|
|+IHC # (%)||47 (87)||3 (5)|
|-IHC # (%)||7 (13)||55 (95)|