EGFR/KRAS/BRAF Mutations in Primary Lung Adenocarcinomas and Corresponding Locoregional Lymph Node Metastases

Abstract

Purpose: The epidermal growth factor receptor (EGFR) and its downstream factors KRAS and BRAF are mutated with different frequencies in non^small cell lung cancer and mutations predict clinical response to EGFR inhibitors. The present study compared the mutational status of EGFR, KRAS, and BRAF in primary tumors with the one in corresponding lymph node metastases. Experimental Design: Direct bidirectional sequencing of EGFR gene exons 18 to 21, KRAS gene codons 12/13 and 61to 68, and BRAF exon 15 was done on 96 paired samples of primary lung adenocarcinomas and corresponding locoregional lymph node metastases. In addition, comparative genomic hybridization analyses in two pairs of corresponding primary and metastatic tumor samples with discordant EGFR mutation status were done. Results: Mutations in EGFR, KRAS, and BRAF were observed in 7 (7%), 36 (38%), and 2 (2%) patients, respectively. Interestingly, KRAS mutations were observed in two patients with an EGFR mutation. Mutations in primary tumors and lymph node metastases were identical in 1of 7 (14%) patients in case of EGFR and 11 of 36 (31%) patients in case of KRAS. One patient harbored different KRAS mutations in primary and corresponding metastatic tumors. Comparative genomic hybridization analysis revealed similar patterns of chromosomal changes, strongly supporting a common clonal origin of primary tumors and metastases. Conclusions:The possibilityofdifferences inthe mutationalstatus of EGFR, KRAS, BRAF between primary tumors and corresponding lymph node metastases should be considered whenever these mutations are used for the selection of patients for EGFR-directed tyrosine kinase inhibitor therapy.

Molecular pathology of lung cancer: key to personalized medicine

“…However, BRAF exon 15 mutations were tested on 96 paired samples of primary lung adenocarcinomas and corresponding locoregional lymph node metastases. 133 BRAF mutations were observed in two patients with KRAS mutations demonstrating the possibility of both mutations in BRAF and KRAS occurring in the same tumor.…”

“…It is expected that in the presence of additional molecular alterations, the effectiveness of TKI therapy for EGFR-mutant tumors will be reduced. The most frequently encountered alterations include KRAS mutations, [118][119][120][121] MET amplification, [122][123][124][125] ALK gene fusion, [126][127][128][129] PIK3CA mutations, 130 BRAF mutations, 118,[131][132][133] and IGF1R overexpression ( Figure 3). 134,135 Is EGFR mutation specific for lung adenocarcinoma?…”

“…132,133 Both KRAS and BRAF genes are part of the signaling cascade for the EGFR family proteins. 211 The BRAF protein is a serine/threonine protein kinase that is activated by KRAS in a GTP-dependent manner.…”

Standardization of Epidermal Growth Factor Receptor (EGFR) Measurement by Quantitative Immunofluorescence and Impact on Antibody-Based Mutation Detection in Non–Small Cell Lung Cancer

“…Five micrograms total protein for each lysate was resolved using SDS-PAGE on an 8% Bis-Tris gel (NuPAGE; Invitrogen Corp., Carlsbad, CA) using NuPAGE MOPS [3-(N-morpholino) propane sulfonic acid] SDS running buffer at 45 mA. On each gel, seven dilutions (10,25,50,125,250, 500, and 1000 ng) of recombinant intracellular domain of EGFR (recEGFR; Cell Signaling Technology, Inc., Beverly, MA) were also resolved for construction of the standard curve. Resolved protein was transferred using NuPAGE transfer buffer at 50V for 2 hours.…”

Targetable “Driver” Mutations in Non Small Cell Lung Cancer

“…In laboratory studies, cells with BRAF mutations were resistant to EGFR inhibitors. Previous studies had reported two novel alterations in BRAF namely an inframe deletion in exon 15 and K601L missense mutation in their study group [57].…”