Molecular diagnosis of activating EGFR mutations in non-small cell lung cancer using mutation-specific antibodies for immunohistochemical analysis.
OBJECTIVE
Therapeutic responses of non-small cell lung carcinoma (NSCLC) to epidermal growth factor receptor (EGFR)-targeted drugs, such as gefitinib and erlotinib, are closely associated with activating EGFR mutations. The most common mutations are delE746-A750 in exon 19 and L858R in exon 21, accounting for approximately 90% of all EGFR mutations. Recently, EGFR mutation-specific antibodies were developed and did well in immunohistochemical analysis, giving a sensitivity of approximately 90%. We have investigated whether this method detects activating EGFR mutations with sensitivity comparable with direct DNA sequencing, which is used to detect these mutations in NSCLC.


METHODS
We used antibodies specific for the E746-A750 deletion mutation in exon 19 and the L858R point mutation in exon 21 in Western blot analysis and immunohistochemistry to determine the presence of these mutations in NSCLC cell lines. We also examined these EGFR mutations in NSCLC tumor samples from 60 patients by immunohistochemically and direct DNA sequencing.


RESULTS
We were able to identify EGFR mutations in NSCLC tumor samples immunohistochemically with a sensitivity of 79% using the anti-delE746-A750 antibody and 83% using the anti-L858R antibody. Additional DNA sequencing markedly improved the sensitivity obtained by immunohistochemistry.


CONCLUSIONS
This simple and rapid assay for detecting EGFR mutations, even in the small bronchial biopsies obtained in stage IV NSCLC patients, will be useful for diagnosing responsiveness to EGFR-targeted drugs in patients with NSCLC. Combining this with DNA sequencing is recommended for the development of improved personalized EGFR-targeted therapeutics.
Kawahara
Akihiko
A
Department of Diagnostic Pathology, Kurume University Hospital, Kurume, Japan.
Yamamoto
Chizuko
C
Nakashima
Kazutaka
K
Azuma
Koichi
K
Hattori
Satoshi
S
Kashihara
Masaki
M
Aizawa
Hisamichi
H
Basaki
Yuji
Y
Kuwano
Michihiko
M
Kage
Masayoshi
M
Mitsudomi
Tetsuya
T
Ono
Mayumi
M
D000906:Antibodies; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics*; D045744:Cell Line, Tumor; D004252:DNA Mutational Analysis / Q000379:methods*; D017353:Gene Deletion; D018773:Genes, erbB-1 / Q000276:immunology*; D006801:Humans; D007150:Immunohistochemistry; D007158:Immunologic Techniques; D008175:Lung Neoplasms / Q000235:genetics*; D009154:Mutation; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
D000906:Antibodies