Pavlova, SarkaFernández Rodríguez, M. ConcepciónBellosillo Paricio, BeatrizPospisilova, Sarka2025-11-252025-11-252025Pavlova S, Malcikova J, Radova L, Bonfiglio S, Cowland JB, Brieghel C, Andersen MK, Karypidou M, Biderman B, Doubek M, Lazarian G, Rapado I, Vynck M, Porret NA, Andres M, Rosenberg D, Sahar D, Martinez-Laperche C, Buño I, Hindley A, Donaldson D, Sanchez JB, Garcia-Marco JA, Serrano-Alcala A, Ferrer-Lores B, Fernandez-Rodriguez C, Bellosillo B, Stilgenbauer S, Tausch E, Nikdin H, Quinn F, Atkinson E, Van de Corput L, Yildiz C, Bilbao-Sieyro C, Florido Y, Thiede C, Schuster C, Stoj A, Czekalska S, Chatzidimitriou A, Laidou S, Bidet A, Dussiau C, Nollet F, Piras G, Monne M, Smirnova S, Nikitin E, Sloma I, Claudel A, Largeaud L, Ysebaert L, Valk PJM, Christian A, Walewska R, Oscier D, Sebastião M, Da Silva MG, Galieni P, Angelini M, Rossi D, Spina V, Matos S, Martins V, Stok¿osa T, Pepek M, Baliakas P, Andreu R, Luna I, Kahre T, Murumets U, Pikousova T, Kurucova T, Laird S, Ward D, Alcoceba M, Balanzategui A, Scarfo L, Gandini F, Zapparoli E, Blanco A, Abrisqueta P, Rodriguez-Vicente AE, Benito R, Bravetti C, Davi F, Gameiro P, Martinez-Lopez J, Tazon-Vega B, Baran-Marszak F, Davis Z, Catherwood M, Sudarikov A, Rosenquist R, Niemann CU, Stamatopoulos K, Ghia P, Pospisilova S. Detection of clinically relevant variants in the TP53 gene below 10% allelic frequency: A multicenter study by ERIC, the European Research Initiative on CLL. HemaSphere. 2025;9(1):e70065. DOI: 10.1002/hem3.700652572-9241http://hdl.handle.net/10230/72004In chronic lymphocytic leukemia, the reliability of next-generation sequencing (NGS) to detect TP53 variants ≤10% allelic frequency (low-VAF) is debated. We tested the ability to detect 23 such variants in 41 different laboratories using their NGS method of choice. The sensitivity was 85.6%, 94.5%, and 94.8% at 1%, 2%, and 3% VAF cut-off, respectively. While only one false positive (FP) result was reported at >2% VAF, it was more challenging to distinguish true variants <2% VAF from background noise (37 FPs reported by 9 laboratories). The impact of low-VAF variants on time-to-second-treatment (TTST) and overall survival (OS) was investigated in a series of 1092 patients. Among patients not treated with targeted agents, patients with low-VAF TP53 variants had shorter TTST and OS versus wt-TP53 patients, and the relative risk of second-line treatment or death increased continuously with increasing VAF. Targeted therapy in ≥2 line diminished the difference in OS between patients with low-VAF TP53 variants and wt-TP53 patients, while patients with high-VAF TP53 variants had inferior OS compared to wild type-TP53 cases. Altogether, NGS-based approaches are technically capable of detecting low-VAF variants. No strict threshold can be suggested from a technical standpoint, laboratories reporting TP53 mutations should participate in a standardized validation set-up. Finally, whereas low-VAF variants affected outcomes in patients receiving chemoimmunotherapy, their impact on those treated with novel therapies remains undetermined. Our results pave the way for the harmonized and accurate TP53 assessment, which is indispensable for elucidating the role of TP53 mutations in targeted treatment.application/pdfeng© 2025 The Author(s). HemaSphere published by John Wiley & Sons Ltd on behalf of European Hematology Association. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License (http://creativecommons.org/licenses/by-nc-nd/4.0/), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.Leucèmia limfocítica crònicainfo:eu-repo/semantics/article2025-11-25http://dx.doi.org/10.1002/hem3.70065info:eu-repo/semantics/openAccess