Rapid and Efficient Detection of EGFR Mutations in Problematic Cytologic Specimens by High-Resolution Melting Analysis

Hlinkova, Katarina; Babál, Pavel; Berzinec, Peter; Majer, I.; Ilenčíková, Denisa

doi:10.1007/bf03257190

Cited by 14 publications

(6 citation statements)

References 29 publications

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“…Small biopsies are often suitable for dissection and molecular analysis. Cytologic samples cannot always be used for mutational analysis because of paucity of neoplastic cells and/or large cellular heterogeneity, which may require highly sensitive mutation detection methods (12,13). In particular, bronchoalveolar lavage (BAL) and pleural fluid are less adequate for molecular testing than other biologic specimens achieved by transbronchial or transthoracic fine-needle aspiration (FNA) due to the lower rate of tumor cells (14).…”

Section: Introductionmentioning

confidence: 99%

Effective Assessment of egfr Mutation Status in Bronchoalveolar Lavage and Pleural Fluids by Next-Generation Sequencing

Buttitta

Felicioni

Grammastro

et al. 2013

Clinical Cancer Research

110

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Purpose: The therapeutic choice for patients with lung adenocarcinoma depends on the presence of EGF receptor (EGFR) mutations. In many cases, only cytologic samples are available for molecular diagnosis. Bronchoalveolar lavage (BAL) and pleural fluid, which represent a considerable proportion of cytologic specimens, cannot always be used for molecular testing because of low rate of tumor cells.Experimental Design: We tested the feasibility of EGFR mutation analysis on BAL and pleural fluid samples by next-generation sequencing (NGS), an innovative and extremely sensitive platform. The study was devised to extend the EGFR test to those patients who could not get it due to the paucity of biologic material. A series of 830 lung cytology specimens was used to select 48 samples (BAL and pleural fluid) from patients with EGFR mutations in resected tumors. These samples included 36 cases with 0.3% to 9% of neoplastic cells (series A) and 12 cases without evidence of tumor (series B). All samples were analyzed by Sanger sequencing and NGS on 454 Roche platform. A mean of 21,130 AE 2,370 sequences per sample were obtained by NGS.Results: In series A, EGFR mutations were detected in 16% of cases by Sanger sequencing and in 81% of cases by NGS. Seventy-seven percent of cases found to be negative by Sanger sequencing showed mutations by NGS. In series B, all samples were negative for EGFR mutation by Sanger sequencing whereas 42% of them were positive by NGS.Conclusions: The very sensitive EGFR-NGS assay may open up to the possibility of specific treatments for patients otherwise doomed to re-biopsies or nontargeted therapies.

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Section: Introductionmentioning

confidence: 99%

Effective Assessment of egfr Mutation Status in Bronchoalveolar Lavage and Pleural Fluids by Next-Generation Sequencing

Buttitta

Felicioni

Grammastro

et al. 2013

Clinical Cancer Research

110

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“…The current study has several strengths compared with the majority of previously published investigations . In particular, both cytological and histological samples from each included patient were used, thus allowing for the direct comparison of the suitability of these types of biological material for molecular testing.…”

Section: Discussionmentioning

confidence: 99%

“…Some institutions already use the molecular analysis of cytological material in a regular diagnostic setting . This practice is based on a significant number of reports demonstrating the suitability of cytological preparations for detection of the EGFR mutation; however, only a few of these studies used paired histological (control) samples and included tumor series with a considerable frequency of mutations . To our knowledge, the results of ALK analysis in archived cytological samples have not been presented to date.…”

Section: Introductionmentioning

confidence: 99%

Detection of EGFR mutations and EML4‐ALK rearrangements in lung adenocarcinomas using archived cytological slides

Mitiushkina

Iyevleva

Poltoratskiy

et al. 2013

Cancer Cytopathology

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BACKGROUND:Although the molecular analysis of epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase (ALK) in archived lung cancer tissues is relatively well established, the genetic testing of cytological material has not yet become a routine. METHODS: The current study used cell samples that were obtained by bronchial brushing, transthoracic needle aspiration, or biopsy imprint preparation between 1993 and 2008. Islets of malignant cells were visually located on the archived cytological slides, lysed in situ by a drop of sodium dodecyl sulfate-containing buffer, and subjected to the standard DNA and RNA extraction. Examination of paraffin-embedded tissue blocks (resection specimens or biopsy material) from the same patients was performed in parallel. RESULTS: A total of 75 cytological/histological lung adenocarcinoma sample pairs underwent polymerase chain reaction analysis for the EGFR mutation. Two cytological samples and 1 morphological sample failed to produce DNA. Concordance for the wild-type and mutation status was observed in 54 of 72 and 14 of 72 informative pairs, respectively; 3 pairs and 1 pair, respectively, had mutation only in the cytological or histological material. The discrepancies could be explained by the failure to ensure a high percentage of lung cancer cells in the analyzed samples or, alternatively, by the genuine intratumoral molecular heterogeneity of some neoplasms. RNA extraction followed by reverse transcriptase-polymerase chain reaction analysis for the EML4-ALK translocation was performed for 44 EGFR mutation-negative sample pairs; failures were observed for 2 cytological and 6 histological specimens. All informative pairs were concordant either for the norm (32 of 36 pairs) or for the presence of EML4-ALK gene fusion (4 of 36 pairs). CONCLUSIONS: Archived cytological slides appear to be well suited both for EGFR and ALK analysis.

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“…Therefore, the most exact predictive marker of EGFR-TKIs may be EGFR mutation. Unfortunately, the detection of an EGFR mutation is difficult due to a limited amount of available tissue 12. Thus, another biomarker that can improve the prediction of response to these targeted drugs is needed.…”

Section: Introductionmentioning

confidence: 99%

EGFRPolymorphism as a Predictor of Clinical Outcome in Advanced Lung Cancer Patients Treated withEGFR-TKI

et al. 2012

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PurposeMutations in the epidermal growth factor receptor (EGFR) have been confirmed as predictors of the efficacy of treatment with EGFR-tyrosine kinase inhibitors (TKIs). We investigated whether polymorphisms of the EGFR gene were associated with clinical outcomes in non-small cell lung cancer (NSCLC) patients treated with EGFR-TKI.Materials and MethodsA polymorphic dinucleotide repeat in intron 1 [CA simple sequence repeat in intron 1(CA-SSR1)] in intron 1 and single nucleotide polymorphisms (SNP-216) in the promoter region of the EGFR gene were evaluated in 71 NSCLC patients by restriction fragment length polymorphism and DNA sequencing. The relationship between genetic polymorphisms and clinical outcomes of treatment with EGFR-TKIs was evaluated.ResultsSNP-216G/T polymorphisms were associated with the efficacy of EGFR-TKI. The response rate for the SNP-216G/T tended to be higher than that for G/G (62.5% vs. 27.4%, p=0.057). The SNP-216G/T genotype was also associated with longer progression-free survival compared with the GG genotype (16.7 months vs. 5.1 months, p=0.005). However, the length of CA-SSR1 was not associated with the efficacy of EGFR-TKI.ConclusionSNP-216G/T polymorphism was a potential predictor of clinical outcomes in NSCLC patients treated with EGFR-TKI.

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Rapid and Efficient Detection of EGFR Mutations in Problematic Cytologic Specimens by High-Resolution Melting Analysis

Cited by 14 publications

References 29 publications

Effective Assessment of egfr Mutation Status in Bronchoalveolar Lavage and Pleural Fluids by Next-Generation Sequencing

Effective Assessment of egfr Mutation Status in Bronchoalveolar Lavage and Pleural Fluids by Next-Generation Sequencing

Detection of EGFR mutations and EML4‐ALK rearrangements in lung adenocarcinomas using archived cytological slides

EGFRPolymorphism as a Predictor of Clinical Outcome in Advanced Lung Cancer Patients Treated withEGFR-TKI

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