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Screening for EGFR and KRAS mutations in non-small cell lung carcinomas using DNA extraction by hydrothermal pressure
Yan Liu1, Bing-Quan Wu, Hao-Hao Zhong
1Department of Pathology, Peking University Health Science Center Beijing, China.
Abstract:
EGFR and KRAS mutations correlate with response to tyrosine kinase inhibitors in patients with non-small cell lung carcinoma (NSCLC). We reported a hydrothermal pressure method of simultaneous deparaffinization and lysis of formalin-fixed paraffin embedded (FFPE) tissue followed by conventional chaotropic salt column purification to obtain high quality DNA for mutation analysis using PCR-base direct sequencing. This study assessed the feasibility of using this method to screen for exons 18-21 of EGFR and exon 2 of KRAS gene mutations in surgical resection and core needle biopsy specimens from 251 NSCLC patients. EGFR mutations were identified in 140 (55.8%) NSCLC patients (118 in adenocarcinoma, 11 in squamous cell carcinoma, 7 in adenocarcinoma and 4 in NSCLC-not otherwise specified), including four novel substitutions (L718M, A743V, L815P, V819E). EGFR mutations were frequently present in female patients (72 of 113, 63.7%) and NSCLC with adenocarcinoma component (125/204, 61.3%) with statistical significance. Twenty-one patients had multiple mutations at different exons of EGFR, in which seventeen patients had deletions in exon 19. KRAS mutations were found in 18 (7.2%) patients (15 in adenocarcinoma, 2 in squamous cell carcinoma and one in NSCLC-not otherwise specified), including an uncommon substitution G13C. Deparaffinization and lysis by hydrothermal pressure, coupled with purification and PCR-based sequencing, provides a robust screening approach for EGFR and KRAS mutation analysis of FFPE tissues from either surgical resection or core needle biopsy in clinical personalized management of lung cancer.
Insights
A new hydrothermal pressure method efficiently extracts high-quality DNA from lung cancer tissues for EGFR and KRAS mutation analysis. This technique aids in personalized treatment strategies for non-small cell lung carcinoma (NSCLC) patients.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- EGFR and KRAS mutations are key biomarkers for tyrosine kinase inhibitor response in non-small cell lung carcinoma (NSCLC).
- Accurate DNA extraction from formalin-fixed paraffin-embedded (FFPE) tissues is crucial for reliable mutation detection.
Purpose of the Study:
- To evaluate a novel hydrothermal pressure method for simultaneous deparaffinization and lysis of FFPE tissues.
- To assess the feasibility of this method for screening EGFR and KRAS mutations in NSCLC patient samples.
Main Methods:
- A hydrothermal pressure technique was used for tissue deparaffinization and lysis.
- High-quality DNA was obtained using chaotropic salt column purification.
- PCR-based direct sequencing was employed for mutation analysis of EGFR (exons 18-21) and KRAS (exon 2).
Main Results:
- EGFR mutations were detected in 55.8% of 251 NSCLC patients, including four novel substitutions.
- KRAS mutations were identified in 7.2% of patients, including an uncommon G13C substitution.
- The method demonstrated high efficiency in analyzing both surgical resection and core needle biopsy specimens.
Conclusions:
- Hydrothermal pressure deparaffinization and lysis is a robust method for EGFR and KRAS mutation screening in FFPE NSCLC tissues.
- This approach supports personalized medicine by enabling efficient molecular profiling.
- The method is suitable for clinical settings utilizing both surgical and biopsy samples.
