ROBEŠOVÁ, Blanka, Monika BAJEROVÁ, Květoslava LIŠKOVÁ, Jana SKŘIČKOVÁ, Marcela TOMÍŠKOVÁ, Šárka POSPÍŠILOVÁ, Jiří MAYER and Dana DVOŘÁKOVÁ. TaqMan based real time PCR assay targeting EML4-ALK fusion transcripts in NSCLC. Lung Cancer. EAST PARK SHANNON (Ireland): ELSEVIER IRELAND LTD, vol. 85, No 1, p. 25-30. ISSN 0169-5002. doi:10.1016/j.lungcan.2014.04.002. 2014.
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Basic information
Original name TaqMan based real time PCR assay targeting EML4-ALK fusion transcripts in NSCLC
Authors ROBEŠOVÁ, Blanka (203 Czech Republic, belonging to the institution), Monika BAJEROVÁ (203 Czech Republic, belonging to the institution), Květoslava LIŠKOVÁ (203 Czech Republic, belonging to the institution), Jana SKŘIČKOVÁ (203 Czech Republic, belonging to the institution), Marcela TOMÍŠKOVÁ (203 Czech Republic, belonging to the institution), Šárka POSPÍŠILOVÁ (203 Czech Republic, guarantor, belonging to the institution), Jiří MAYER (203 Czech Republic, belonging to the institution) and Dana DVOŘÁKOVÁ (203 Czech Republic, belonging to the institution).
Edition Lung Cancer, EAST PARK SHANNON (Ireland), ELSEVIER IRELAND LTD, 2014, 0169-5002.
Other information
Original language English
Type of outcome Article in a journal
Field of Study 30200 3.2 Clinical medicine
Country of publisher Ireland
Confidentiality degree is not subject to a state or trade secret
WWW URL
Impact factor Impact factor: 3.958
RIV identification code RIV/00216224:14740/14:00077719
Organization unit Central European Institute of Technology
Doi http://dx.doi.org/10.1016/j.lungcan.2014.04.002
UT WoS 000338617100005
Keywords in English Lung cancer; Targeted therapy; NSCLC; EML4-ALK; Real time PCR; FISH
Tags EL OK, kontrola MP, podil, rivok
Tags International impact, Reviewed
Changed by Changed by: Martina Prášilová, učo 342282. Changed: 28/1/2015 13:48.
Abstract
OBJECTIVES: Lung cancer with the ALK rearrangement constitutes only a small fraction of patients with non-small cell lung cancer (NSCLC). However, in the era of molecular-targeted therapy, efficient patient selection is crucial for successful treatment. In this context, an effective method for EML4-ALK detection is necessary. We developed a new highly sensitive variant specific TaqMan based real time PCR assay applicable to RNA from formalin-fixed paraffin-embedded tissue (FFPE). MATERIALS AND METHODS: This assay was used to analyze the EML4-ALK gene in 96 non-selected NSCLC specimens and compared with two other methods (end-point PCR and break-apart FISH). RESULTS: EML4-ALK was detected in 33/96 (34%) specimens using variant specific real time PCR, whereas in only 23/96 (24%) using end-point PCR. All real time PCR positive samples were confirmed with direct sequencing. A total of 46 specimens were subsequently analyzed by all three detection methods. Using variant specific real time PCR we identified EML4-ALK transcript in 17/46 (37%) specimens, using end-point PCR in 13/46 (28%) specimens and positive ALK rearrangement by FISH was detected in 8/46 (17.4%) specimens. Moreover, using variant specific real time PCR, 5 specimens showed more than one EML4-ALK variant simultaneously (in 2 cases the variants 1+3a+3b, in 2 specimens the variants 1+3a and in 1 specimen the variant 1+3b). In one case of 96 EML4-ALK fusion gene and EGFR mutation were detected. All simultaneous genetic variants were confirmed using end-point PCR and direct sequencing. CONCLUSION: Our variant specific real time PCR assay is highly sensitive, fast, financially acceptable, applicable to FFPE and seems to be a valuable tool for the rapid prescreening of NSCLC patients in clinical practice, so, that most patients able to benefit from targeted therapy could be identified.
Links
ED1.1.00/02.0068, research and development projectName: CEITEC - central european institute of technology
EE2.3.20.0045, research and development projectName: Podpora profesního růstu a mezinárodní integrace výzkumných týmů v oblasti molekulární medicíny
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