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TP53 mutation analysis in chronic lymphocytic leukemia: comparison of different detection methods
Barbara Kantorova1, Jitka Malcikova, Jana Smardova
1Central European Institute of Technology (CEITEC), Masaryk University Brno, Kamenice 5, CZ-625 00, Brno, Czech Republic.
TP53 gene mutations impact chronic lymphocytic leukemia (CLL) outcomes. Denaturing high-performance liquid chromatography (DHPLC) and functional analysis of separated alleles in yeast (FASAY) effectively detect most TP53 mutations, while ultra-deep next-generation sequencing (NGS) is better for low-proportion variants.
Area of Science:
- Oncology
- Genetics
- Molecular Biology
Background:
- TP53 gene mutations are critical prognostic markers in chronic lymphocytic leukemia (CLL), correlating with poor survival and treatment resistance.
- Current TP53 mutation detection methods have limitations, necessitating a discussion on optimal methodologies for comprehensive analysis.
Purpose of the Study:
- To evaluate the limitations of standard TP53 mutation detection techniques in CLL.
- To propose an optimized strategy for identifying TP53 gene alterations in CLL patients.
Main Methods:
- Comparative analysis of denaturing high-performance liquid chromatography (DHPLC), functional analysis of separated alleles in yeast (FASAY), AmpliChip p53, and ultra-deep next-generation sequencing (NGS).
- Examination of 182 CLL patients, with 69 undergoing ultra-deep NGS analysis.
- Inclusion of high-risk CLL cases to enrich for TP53 mutations.
Main Results:
- A total of 79 TP53 mutations were identified in 57 patients (31%), with missense substitutions being the most common (68%).
- DHPLC and FASAY, when combined with Sanger sequencing, demonstrated high efficacy, detecting 95% and 93% of mutated patients, respectively.
- Ultra-deep NGS proved more suitable for detecting low-proportion TP53 mutations, highlighting its sensitivity.
Conclusions:
- DHPLC and FASAY are reliable methods for TP53 mutation detection in CLL.
- Ultra-deep NGS offers a promising approach to overcome current method limitations, enabling the detection of low-frequency mutations.
- An integrated approach combining established methods with ultra-deep NGS may provide the most comprehensive TP53 mutation analysis in CLL.
