TP53 mutation analysis in chronic lymphocytic leukemia: comparison of different detection methods

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Authors	KANTOROVÁ Barbara MALČÍKOVÁ Jitka ŠMARDOVÁ Jana PAVLOVÁ Šárka TRBUŠEK Martin TOM Nikola PLEVOVÁ Karla TICHÝ Boris TRUONG Sim DIVISKOVA Eva KOTAŠKOVÁ Jana OLTOVÁ Alexandra PATTEN Nancy BRYCHTOVÁ Yvona DOUBEK Michael MAYER Jiří POSPÍŠILOVÁ Šárka
Year of publication	2015
Type	Article in Periodical
Magazine / Source	Tumor Biology
MU Faculty or unit	Central European Institute of Technology
Citation
Web	http://download.springer.com/static/pdf/851/art%253A10.1007%252Fs13277-014-2971-0.pdf?auth66=1421241751_ae3f1991394a2e909c07bb253d606b7c&ext=.pdf
Doi	http://dx.doi.org/10.1007/s13277-014-2971-0
Field	Oncology and hematology
Keywords	TP53 gene; Mutationanalysis; Detection methods; Chronic lymphocytic leukemia
Attached files	ZVV_2014_142_1216519_TP53_mutation.pdf
Description	TP53 gene defects represent a strong adverse prog- nostic factor for patient survival and treatment resistance in chronic lymphocytic leukemia (CLL). Although various methods for TP53 mutation analysis have been reported, none of them allow the identification of all occurring sequence variants, and the most suitable methodology is still being discussed. The aim of this study was to determine the limita- tions of commonly used methods for TP53 mutation exami- nation in CLL and propose an optimal approach for their detection. We examined 182 CLL patients enriched for high- risk cases using denaturing high-performance liquid chroma- tography (DHPLC), functional analysis of separated alleles in yeast (FASAY), and the AmpliChip p53 Research Test in parallel. The presence of T53 gene mutations was also evalu- ated using ultra-deep next generation sequencing (NGS) in 69 patients. In total, 79 TP53 mutations in 57 (31 %) patients were found; among them, missense substitutions predominat- ed (68 % of detected mutations). Comparing the efficacy of the methods used, DHPLC and FASAY both combined with direct Sanger sequencing achieved the best results, identifying 95 % and 93 % of TP53 -mutated patients. Nevertheless, we showed that in CLL patients carrying low-proportion TP53 mutation, the more sensitive approach, e.g., ultra-deep NGS, might be more appropriate. TP53 gene analysis using DHPLC or FASAYis a suitable approach for mutation detection. Ultra- deep NGS has the potential to overcome shortcomings of methods currently used, allows the detection of minor propor- tion mutations, and represents thus a promising methodology for near future.
Related projects:	CEITEC - Central European Institute of Technology Molekulární charakterizace B buněčných receptorů a jejich vztah k evoluci genetických změn u chronické lymfocytární leukémie Časná identifikace CLL pacientů s dosud nevyselektovanými mutacemi v proteinu p53