Rapid detection of epidermal growth factor receptor mutations in lung cancer by the SMart-Amplification Process.
OBJECTIVE
A positive response to gefitinib in non-small cell lung cancer (NSCLC) has been correlated to mutations in epidermal growth factor receptor (EGFR) gene. Previous reports have been based mainly on diagnostic screening by sequencing. However, sequencing is a time-consuming and complicated procedure, not suitable for routine clinical use.


METHODS
We have developed rapid, simple, and sensitive mutation detection assays based on the SMart Amplification Process (SMAP) and applied it for analyzing EGFR gene mutations in clinical samples. By using SMAP, we can detect mutations within 30 min including sample preparation. To validate the assay system for potential use in clinical diagnostics, we examined 45 NSCLC patients for EGFR mutations using sequencing and SMAP.


RESULTS
The outcomes of the SMAP assay perfectly matched the sequencing results, except in one case where SMAP was able to identify a mutation that was not detected by sequencing. We also evaluated the sensitivity and specificity of SMAP in mutation detection for EGFR. In a serial dilution study, SMAP was able to find a mutation in a sample containing only 0.1% of the mutant allele in a mixture of wild-type genomic DNA. We also could show amplification of mutated DNA with only 30 copies per reaction.


CONCLUSIONS
The SMAP method offers higher sensitivity and specificity than alternative technologies, while eliminating the need for sequencing to identify mutations in the EGFR gene of NSCLC. It provides a robust and point-of-care accessible approach for a rapid identification of most patients likely to respond to gefitinib.
Hoshi
Kanako
K
Genome Exploration Research Group (Genome Network Project Core Group), RIKEN Genomic Sciences Center, RIKEN Yokohama Institute, 1-7-22 Suehiro-cho, Japan. Canaty@aol.com
Takakura
Hideki
H
Mitani
Yasumasa
Y
Tatsumi
Kenji
K
Momiyama
Nobuyoshi
N
Ichikawa
Yasushi
Y
Togo
Shinji
S
Miyagi
Toru
T
Kawai
Yuki
Y
Kogo
Yasushi
Y
Kikuchi
Takeshi
T
Kato
Chiaki
C
Arakawa
Takahiro
T
Uno
Syuji
S
Cizdziel
Paul E
PE
Lezhava
Alexander
A
Ogawa
Noburou
N
Hayashizaki
Yoshihide
Y
Shimada
Hiroshi
H
D000368:Aged; D000369:Aged, 80 and over; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics*; D045744:Cell Line, Tumor; D066246:ErbB Receptors / Q000235:genetics*; D005091:Exons; D005260:Female; D017353:Gene Deletion; D005838:Genotype; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D009154:Mutation; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012680:Sensitivity and Specificity