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@article{1212604, author = {Robešová, Blanka and Bajerová, Monika and Lišková, Květoslava and Skřičková, Jana and Tomíšková, Marcela and Pospíšilová, Šárka and Mayer, Jiří and Dvořáková, Dana}, article_location = {EAST PARK SHANNON (Ireland)}, article_number = {1}, doi = {http://dx.doi.org/10.1016/j.lungcan.2014.04.002}, keywords = {Lung cancer; Targeted therapy; NSCLC; EML4-ALK; Real time PCR; FISH}, language = {eng}, issn = {0169-5002}, journal = {Lung Cancer}, title = {TaqMan based real time PCR assay targeting EML4-ALK fusion transcripts in NSCLC}, url = {http://www.sciencedirect.com/science/article/pii/S016950021400169X}, volume = {85}, year = {2014} }
TY - JOUR ID - 1212604 AU - Robešová, Blanka - Bajerová, Monika - Lišková, Květoslava - Skřičková, Jana - Tomíšková, Marcela - Pospíšilová, Šárka - Mayer, Jiří - Dvořáková, Dana PY - 2014 TI - TaqMan based real time PCR assay targeting EML4-ALK fusion transcripts in NSCLC JF - Lung Cancer VL - 85 IS - 1 SP - 25-30 EP - 25-30 PB - ELSEVIER IRELAND LTD SN - 01695002 KW - Lung cancer KW - Targeted therapy KW - NSCLC KW - EML4-ALK KW - Real time PCR KW - FISH UR - http://www.sciencedirect.com/science/article/pii/S016950021400169X L2 - http://www.sciencedirect.com/science/article/pii/S016950021400169X N2 - 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. ER -
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. \textit{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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