Abstract
BackgroundThe recent development of antibodies specific for the major hotspot mutations in the epidermal growth factor receptor (EGFR), L858R and E746_A750del, may provide an opportunity to use immunohistochemistry (IHC) as a screening test for EGFR gene mutations. This study was designed to optimize the IHC protocol and the criteria for interpretation of the results using DNA sequencing as the gold-standard.MethodsTumor sections from fifty lung adenocarcinoma specimens from Chinese patients were immunostained using L858R and E746_A750del-specific antibodies using three different antigen retrieval solutions, and the results were evaluated using three different sets of criteria. The same specimens were used for DNA purification and analysis of EGFR gene mutations.ResultsIn this study the optimal buffer for antigen retrieval was EDTA (pH 8.0), and the optimal scoring method was to call positive results when there was moderate to strong staining of membrane and/or cytoplasm in >10% of the tumor cells. Using the optimized protocol, L858R-specific IHC showed a sensitivity of 81% and a specificity of 97%, and E746_A750del-specific IHC showed a sensitivity of 59% and a specificity of 100%, both compared with direct DNA analysis. Additionally, the mutant proteins as assessed by IHC showed a more homogeneous than heterogeneous pattern of expression.ConclusionsOur data demonstrate that mutation-specific IHC, using optimized procedures, is a reliable prescreening test for detecting EGFR mutations in lung adenocarcinoma.Virtual SlidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2059012601872392
Highlights
The recent development of antibodies specific for the major hotspot mutations in the epidermal growth factor receptor (EGFR), L858R and E746_A750del, may provide an opportunity to use immunohistochemistry (IHC) as a screening test for EGFR gene mutations
L858R-specific IHC showed a sensitivity of 81%, a specificity of 97%, a positive predictive value (PPV) of 93%, and a negative predictive value (NPV) of 92% on Score A; a sensitivity of 44%, a specificity of 97%, a PPV of 88%, and a NPV of 79% on Score B; and a sensitivity of 69%, a specificity of 74%, a PPV of 55%, and a NPV of 83% on Score C
Immunohistochemistry using mutation-specific mAbs is demonstrated to be a reliable test for detecting EGFR mutations in adenocarcinoma of the lung in our study
Summary
The recent development of antibodies specific for the major hotspot mutations in the epidermal growth factor receptor (EGFR), L858R and E746_A750del, may provide an opportunity to use immunohistochemistry (IHC) as a screening test for EGFR gene mutations. Most of them confirmed a high degree of specificity, but the reported sensitivities were quite variable ranging from 24% to 100% (Table 1) [8,9,10,11,12,13,14] This inconsistency may be related to differences in methodology and interpretation [10,13,15], as well as population specific differences in gene mutations and differences in the level of protein expression [16]. This inconsistency suggests that further study is needed in diverse populations before EGFR mutation-specific IHC can be implemented as a clinical tool
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