Rapid KRAS, EGFR, BRAF and PIK3CA Mutation Analysis of Fine Needle Aspirates from Non-Small-Cell Lung Cancer Using Allele-Specific qPCR

Eijk, Ronald van; Licht, Jappe; Schrumpf, Melanie; Yazdi, Mehrdad Talebian; Ruano, Dina; Forte, Giusi Irma; Nederlof, Petra M.; Veselic, Maud; Rabe, Klaus F.; Annema, Jouke T.; Smit, Vincent T.H.B.M.; Morreau, Hans; Wezel, Tom van

doi:10.1371/journal.pone.0017791

Cited by 173 publications

(107 citation statements)

References 35 publications

(34 reference statements)

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“…Allele‐specific qPCR was performed to confirm the status of KRAS, BRAF and PIK3CA mutation hotspot loci 29. TP53 (exons 5–8) and APC (mutation cluster region; exon 15) were analyzed via Sanger DNA sequencing.…”

Section: Methodsmentioning

confidence: 99%

Somatic mutation profiles in primary colorectal cancers and matching ovarian metastases: Identification of driver and passenger mutations

Crobach

Ruano

Eijk

et al. 2016

The Journal of Pathology CR

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The mutational profiles of primary colorectal cancers (CRCs) and corresponding ovarian metastases were compared. Using a custom‐made next generation sequencing panel, 115 cancer‐driving genes were analyzed in a cohort of 26 primary CRCs and 30 matching ovarian metastases (four with bilateral metastases). To obtain a complete overview of the mutational profile, low thresholds were used in bioinformatics analysis to prevent low frequency passenger mutations from being filtered out. A subset of variants was validated using Sanger and/or hydrolysis probe assays. The mutational landscape of CRC that metastasized to the ovary was not strikingly different from CRC in consecutive series. When comparing primary CRCs and their matching ovarian metastases, there was considerable overlap in the mutations of early affected genes. A subset of mutations demonstrated less overlap, presumably being passenger mutations. In particular, primary CRCs showed a substantially high number of passenger mutations. We also compared the primary CRCs and matching metastases for stratifying variants of six genes (KRAS, NRAS, BRAF, FBXW7, PTEN and PIK3CA) that select for established (EGFR directed) or future targeted therapies. In a total of 31 variants 12 were not found in either of the two locations. Tumours thus differed in the number of discordant variants between the primary tumours and matching metastases. Half of these discordant variants were definitive class 4/5 pathogenic variants. However, in terms of temporal heterogeneity, no clear relationship was observed between the number of discordant variants and the time interval between primary CRCs and the detection of ovarian metastases. This suggests that dormant metastases may be present from the early days of the primary tumours.

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Section: Methodsmentioning

confidence: 99%

Somatic mutation profiles in primary colorectal cancers and matching ovarian metastases: Identification of driver and passenger mutations

Crobach

Ruano

Eijk

et al. 2016

The Journal of Pathology CR

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“…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?…”

Section: Other Alterations That Affect the Egfr Tki Responsementioning

confidence: 99%

Molecular pathology of lung cancer: key to personalized medicine

et al. 2012

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The majority of lung adenocarcinoma patients with epidermal growth factor receptor-(EGFR) mutated or EML4-ALK rearrangement-positive tumors are sensitive to tyrosine kinase inhibitors. Both primary and acquired resistance in a significant number of those patients to these therapies remains a major clinical problem. The specific molecular mechanisms associated with tyrosine kinase inhibitor resistance are not fully understood. Clinicopathological observations suggest that molecular alterations involving so-called 'driver mutations' could be used as markers that aid in the selection of patients most likely to benefit from targeted therapies. In this review, we summarize recent developments involving the specific molecular mechanisms and markers that have been associated with primary and acquired resistance to EGFR-targeted therapy in lung adenocarcinomas. Understanding these mechanisms may provide new treatment avenues and improve current treatment algorithms.

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“…After DNA isolation, EGFR, BRAF, KRAS and PIK3CA somatic mutation analysis was performed using an allele-specific quantitative real-time PCR (qPCR) with hydrolysis probes (16) AKT, phosphorylated AKT, ERK, pERK, b-catenin and PTEN immunohistochemistry A tissue microarray was constructed containing three representative tissue cores (of 0.6 cm in diameter) from each tumour (17). Four-micrometer tissue sections were cut and transferred to glass slides using a tapetransfer system (Instrumedics, Hackensack, NJ, USA).…”

Section: Somatic Mutation Analysismentioning

confidence: 99%

Mutational analyses of epidermal growth factor receptor and downstream pathways in adrenocortical carcinoma

Hermsen

Haak

Krijger

et al. 2013

European Journal of Endocrinology

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Background: Adrenocortical carcinoma (ACC) is a rare disease with a poor prognosis and limited therapeutic options. Mitotane is considered the standard first-line therapy with only 30% of the patients showing objective tumour response. Defining predictive factors for response is therefore of clinical importance.The epidermal growth factor receptor (EGFR) has been implicated in the development of one-third of all malignancies. EGFR pathway members in ACC have been investigated, however, without available clinical data and relation to survival. Methods: In this study, mutation status of EGFR and downstream signalling pathways was evaluated in 47 ACC patients on mitotane using direct sequencing, a TaqMan allele-specific assay and immunohistochemistry. Archival formalin-fixed paraffin-embedded tumour tissue was used for all analyses. Patient data were obtained anonymously, after coupling with the collected tumour tissue. Results: One BRAF, two EGFR TK domain (c.2590GOA, p.864AOT) and 11 TP53, but no PIK3CA or KRAS, mutations were found. No relationship was found between mutation status, immunostaining and mitotane response or survival. Conclusion: In conclusion, our data suggest that the role of EGFR tyrosine kinase inhibitors in ACC is limited. Treatment with EGFR monoclonal antibodies on the other hand might be beneficial for a larger group of patients. The possible efficacy of this therapy in ACC should be evaluated in future trials.

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Rapid KRAS, EGFR, BRAF and PIK3CA Mutation Analysis of Fine Needle Aspirates from Non-Small-Cell Lung Cancer Using Allele-Specific qPCR

Cited by 173 publications

References 35 publications

Somatic mutation profiles in primary colorectal cancers and matching ovarian metastases: Identification of driver and passenger mutations

Somatic mutation profiles in primary colorectal cancers and matching ovarian metastases: Identification of driver and passenger mutations

Molecular pathology of lung cancer: key to personalized medicine

Mutational analyses of epidermal growth factor receptor and downstream pathways in adrenocortical carcinoma

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