In chronic lymphocytic leukemia (CLL), analysis of TP53 aberrations (deletion and/or mutation) is a crucial part of treatment decision-making algorithms. Technological and treatment advances have resulted in the need for an update of the last recommendations for TP53 analysis in CLL, published by ERIC, the European Research Initiative on CLL, in 2018. Based on the current knowledge of the relevance of low-burden TP53-mutated clones, a specific variant allele frequency (VAF) cut-off for reporting TP53 mutations is no longer recommended, but instead, the need for thorough method validation by the reporting laboratory is emphasized. The result of TP53 analyses should always be interpreted within the context of available laboratory and clinical information, treatment indication, and therapeutic options. Methodological aspects of introducing next-generation sequencing (NGS) in routine practice are discussed with a focus on reliable detection of low-burden clones. Furthermore, potential interpretation challenges are presented, and a simplified algorithm for the classification of TP53 variants in CLL is provided, representing a consensus based on previously published guidelines. Finally, the reporting requirements are highlighted, including a template for clinical reports of TP53 aberrations. These recommendations are intended to assist diagnosticians in the correct assessment of TP53 mutation status, but also physicians in the appropriate understanding of the lab reports, thus decreasing the risk of misinterpretation and incorrect management of patients in routine practice whilst also leading to improved stratification of patients with CLL in clinical trials.

• MeSH
• chronická lymfatická leukemie * genetika   diagnóza   MeSH
• lidé MeSH
• mutace * MeSH
• mutační analýza DNA metody   normy   MeSH
• nádorový supresorový protein p53 * genetika   MeSH
• vysoce účinné nukleotidové sekvenování * metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• nádorový supresorový protein p53 * MeSH
• TP53 protein, human MeSH Prohlížeč

Anaplastic large-cell lymphoma (ALCL) is a T-cell malignancy predominantly driven by the oncogenic anaplastic lymphoma kinase (ALK), accounting for approximately 15% of all paediatric non-Hodgkin lymphoma. Patients with central nervous system (CNS) relapse are particularly difficult to treat with a 3-year overall survival of 49% and a median survival of 23.5 months. The second-generation ALK inhibitor brigatinib shows superior penetration of the blood-brain barrier unlike the first-generation drug crizotinib and has shown promising results in ALK+ non-small-cell lung cancer. However, the benefits of brigatinib in treating aggressive paediatric ALK+ ALCL are largely unknown. We established a patient-derived xenograft (PDX) resource from ALK+ ALCL patients at or before CNS relapse serving as models to facilitate the development of future therapies. We show in vivo that brigatinib is effective in inducing the remission of PDX models of crizotinib-resistant (ALK C1156Y, TP53 loss) ALCL and furthermore that it is superior to crizotinib as a second-line approach to the treatment of a standard chemotherapy relapsed/refractory ALCL PDX pointing to brigatinib as a future therapeutic option.

• Klíčová slova
• ALCL, PDX, brigatinib, crizotinib, tyrosine kinase inhibitors,
• MeSH
• anaplastická lymfomová kináza MeSH
• anaplastický velkobuněčný lymfom * farmakoterapie   patologie   MeSH
• dítě MeSH
• heterografty MeSH
• inhibitory proteinkinas terapeutické užití   MeSH
• krizotinib farmakologie   terapeutické užití   MeSH
• lidé MeSH
• lokální recidiva nádoru farmakoterapie   MeSH
• nádory plic * farmakoterapie   MeSH
• nemalobuněčný karcinom plic * MeSH
• organofosforové sloučeniny farmakologie   terapeutické užití   MeSH
• tyrosinkinasové receptory terapeutické užití   MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• anaplastická lymfomová kináza MeSH
• brigatinib MeSH Prohlížeč
• inhibitory proteinkinas MeSH
• krizotinib MeSH
• organofosforové sloučeniny MeSH
• tyrosinkinasové receptory MeSH

BACKGROUND: Telomeres are protective structures at chromosome ends which shorten gradually with increasing age. In chronic lymphocytic leukemia (CLL), short telomeres have been associated with unfavorable disease outcome, but the link between clonal evolution and telomere shortening remains unresolved. METHODS: We investigated relative telomere length (RTL) in a well-characterized cohort of 198 CLL patients by qPCR and focused in detail on a subgroup 26 patients who underwent clonal evolution of TP53 mutations (evolTP53). In the evolTP53 subgroup we explored factors influencing clonal evolution and corresponding changes in telomere length through measurements of telomerase expression, lymphocyte doubling time, and BCR signaling activity. RESULTS: At baseline, RTL of the evolTP53 patients was scattered across the entire RTL spectrum observed in our CLL cohort. RTL changed in the follow-up samples of 16/26 (62%) evolTP53 cases, inclining to reach intermediate RTL values, i.e., longer telomeres shortened compared to baseline while shorter ones prolonged. For the first time we show that TP53 clonal shifts are linked to RTL change, including unexpected RTL prolongation. We further investigated parameters associated with RTL changes. Unstable telomeres were significantly more frequent among younger patients (P = 0.032). Shorter telomeres were associated with decreased activity of the B-cell receptor signaling components p-ERK1/2, p-ZAP-70/SYK, and p-NFκB (P = 0.04, P = 0.01, and P = 0.02, respectively). CONCLUSIONS: Our study revealed that changes of telomere length reflect evolution in leukemic subclone proportion, and are associated with specific clinico-biological features of the explored cohort.

• Klíčová slova
• BCR signaling, Chronic Lymphocytic Leukemia, Clonal evolution, TP53, Telomere,
• MeSH
• chronická lymfatická leukemie genetika   MeSH
• klonální evoluce genetika   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mutace MeSH
• nádorový supresorový protein p53 genetika   MeSH
• protoonkogenní proteiny c-bcr metabolismus   MeSH
• signální transdukce MeSH
• telomerasa genetika   MeSH
• telomery ultrastruktura   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• nádorový supresorový protein p53 MeSH
• protoonkogenní proteiny c-bcr MeSH
• telomerasa MeSH
• TP53 protein, human MeSH Prohlížeč

Patients with chronic lymphocytic leukemia (CLL) bearing TP53 mutations experience chemorefractory disease and are therefore candidates for targeted therapy. However, the significance of low-burden TP53 mutations with <10% variant allele frequency (VAF) remains a matter for debate. Herein, we describe clonal evolution scenarios of low-burden TP53 mutations, the clinical impact of which we analyzed in a "real-world" CLL cohort. TP53 status was assessed by targeted next-generation sequencing (NGS) in 511 patients entering first-line treatment with chemo- and/or immunotherapy and 159 patients in relapse before treatment with targeted agents. Within the pretherapy cohort, 16% of patients carried low-burden TP53 mutations (0.1% to 10% VAF). Although their presence did not significantly shorten event-free survival after first-line therapy, it affected overall survival (OS). In a subgroup with TP53 mutations of 1% to 10% VAF, the impact on OS was observed only in patients with unmutated IGHV who had not received targeted therapy, as patients benefited from switching to targeted agents, regardless of initial TP53 mutational status. Analysis of the clonal evolution of low-burden TP53 mutations showed that the highest expansion rates were associated with fludarabine, cyclophosphamide, and rituximab regimen in both first- and second-line treatments (median VAF increase, 14.8× and 11.8×, respectively) in contrast to treatment with less intense treatment regimens (1.6×) and no treatment (0.8×). In the relapse cohort, 33% of patients carried low-burden TP53 mutations, which did not expand significantly upon targeted treatment (median VAF change, 1×). Sporadic cases of TP53 mutations' clonal shifts were connected with the development of resistance-associated mutations. Altogether, our data support the incorporation of low-burden TP53 variants in clinical decision making.

• MeSH
• chronická lymfatická leukemie genetika   terapie   MeSH
• dospělí MeSH
• imunoterapie MeSH
• Kaplanův-Meierův odhad MeSH
• klonální evoluce * účinky léků   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mutace účinky léků   MeSH
• nádorové buňky kultivované MeSH
• nádorový supresorový protein p53 genetika   MeSH
• protokoly protinádorové kombinované chemoterapie terapeutické užití   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• nádorový supresorový protein p53 MeSH
• TP53 protein, human MeSH Prohlížeč

Chronic lymphocytic leukemia is associated with a highly heterogeneous disease course in terms of clinical outcomes and responses to chemoimmunotherapy. This heterogeneity is partly due to genetic aberrations identified in chronic lymphocytic leukemia cells such as mutations of TP53 and/or deletions in chromosome 17p [del(17p)], resulting in loss of one TP53 allele. These aberrations are associated with markedly decreased survival and predict impaired response to chemoimmunotherapy thus being among the strongest predictive markers guiding treatment decisions in chronic lymphocytic leukemia. Clinical trials demonstrate the importance of accurately testing for TP53 aberrations [both del(17p) and TP53 mutations] before each line of treatment to allow for appropriate treatment decisions that can optimize patients' outcomes. The current report reviews the diagnostic methods to detect TP53 disruption better, the role of TP53 aberrations in treatment decisions and current therapies available for patients with chronic lymphocytic leukemia carrying these abnormalities. The standardization in sequencing technologies for accurate identification of TP53 mutations and the importance of continued evaluation of TP53 aberrations throughout initial and subsequent lines of therapy remain unmet clinical needs as new therapeutic alternatives become available.

• MeSH
• chromozomální delece * MeSH
• chronická lymfatická leukemie diagnóza   genetika   terapie   MeSH
• lidé MeSH
• lidské chromozomy, pár 17 genetika   MeSH
• mutace * MeSH
• nádorový supresorový protein p53 genetika   MeSH
• přežití bez známek nemoci MeSH
• prognóza MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• nádorový supresorový protein p53 MeSH

In chronic lymphocytic leukemia (CLL), TP53 gene defects, due to deletion of the 17p13 locus and/or mutation(s) within the TP53 gene, are associated with resistance to chemoimmunotherapy and a particularly dismal clinical outcome. On these grounds, analysis of TP53 aberrations has been incorporated into routine clinical diagnostics to improve patient stratification and optimize therapeutic decisions. The predictive implications of TP53 aberrations have increasing significance in the era of novel targeted therapies, i.e., inhibitors of B-cell receptor (BcR) signaling and anti-apoptotic BCL2 family members, owing to their efficacy in patients with TP53 defects. In this report, the TP53 Network of the European Research Initiative on Chronic Lymphocytic Leukemia (ERIC) presents updated recommendations on the methodological approaches for TP53 mutation analysis. Moreover, it provides guidance to ensure that the analysis is performed in a timely manner for all patients requiring treatment and that the data is interpreted and reported in a consistent, standardized, and accurate way. Since next-generation sequencing technologies are gaining prominence within diagnostic laboratories, this report also offers advice and recommendations for the interpretation of TP53 mutation data generated by this methodology.

• MeSH
• chronická lymfatická leukemie genetika   MeSH
• geny p53 genetika   MeSH
• lidé MeSH
• mutační analýza DNA metody   MeSH
• vysoce účinné nukleotidové sekvenování metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• směrnice pro lékařskou praxi MeSH
• Geografické názvy
• Evropa MeSH

Chronic lymphocytic leukemia is a disease with up-regulated expression of the transmembrane tyrosine-protein kinase ROR1, a member of the Wnt/planar cell polarity pathway. In this study, we identified COBLL1 as a novel interaction partner of ROR1. COBLL1 shows clear bimodal expression with high levels in chronic lymphocytic leukemia patients with mutated IGHV and approximately 30% of chronic lymphocytic leukemia patients with unmutated IGHV. In the remaining 70% of chronic lymphocytic leukemia patients with unmutated IGHV, COBLL1 expression is low. Importantly, chronic lymphocytic leukemia patients with unmutated IGHV and high COBLL1 have an unfavorable disease course with short overall survival and time to second treatment. COBLL1 serves as an independent molecular marker for overall survival in chronic lymphocytic leukemia patients with unmutated IGHV. In addition, chronic lymphocytic leukemia patients with unmutated IGHV and high COBLL1 show impaired motility and chemotaxis towards CCL19 and CXCL12 as well as enhanced B-cell receptor signaling pathway activation demonstrated by increased PLCγ2 and SYK phosphorylation after IgM stimulation. COBLL1 expression also changes during B-cell maturation in non-malignant secondary lymphoid tissue with a higher expression in germinal center B cells than naïve and memory B cells. Our data thus suggest COBLL1 involvement not only in chronic lymphocytic leukemia but also in B-cell development. In summary, we show that expression of COBLL1, encoding novel ROR1-binding partner, defines chronic lymphocytic leukemia subgroups with a distinct response to microenvironmental stimuli, and independently predicts survival of chronic lymphocytic leukemia with unmutated IGHV.

• MeSH
• analýza přežití MeSH
• chronická lymfatická leukemie klasifikace   diagnóza   genetika   mortalita   MeSH
• lidé MeSH
• mutace MeSH
• pohyb buněk MeSH
• polarita buněk MeSH
• prognóza MeSH
• signální dráha Wnt MeSH
• sirotčí receptory podobné receptoru tyrosinkinasy metabolismus   MeSH
• těžké řetězce imunoglobulinů genetika   MeSH
• transkripční faktory metabolismus   MeSH
• variabilní oblast imunoglobulinu genetika   MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• COBLL1 protein, human MeSH Prohlížeč
• ROR1 protein, human MeSH Prohlížeč
• sirotčí receptory podobné receptoru tyrosinkinasy MeSH
• těžké řetězce imunoglobulinů MeSH
• transkripční faktory MeSH
• variabilní oblast imunoglobulinu MeSH

Treatment options for TP53-mutated lymphoid tumors are very limited. In experimental models, TP53-mutated lymphomas were sensitive to direct inhibition of checkpoint kinase 1 (Chk1), a pivotal regulator of replication. We initially tested the potential of the highly specific Chk1 inhibitor SCH900776 to synergize with nucleoside analogs (NAs) fludarabine, cytarabine and gemcitabine in cell lines derived from B-cell malignancies. In p53-proficient NALM-6 cells, SCH900776 added to NAs enhanced signaling towards Chk1 (pSer317/pSer345), effectively blocked Chk1 activation (Ser296 autophosphorylation), increased replication stress (p53 and γ-H2AX accumulation) and temporarily potentiated apoptosis. In p53-defective MEC-1 cell line representing adverse chronic lymphocytic leukemia (CLL), Chk1 inhibition together with NAs led to enhanced and sustained replication stress and significantly potentiated apoptosis. Altogether, among 17 tested cell lines SCH900776 sensitized four of them to all three NAs. Focusing further on MEC-1 and co-treatment of SCH900776 with fludarabine, we disclosed chromosome pulverization in cells undergoing aberrant mitoses. SCH900776 also increased the effect of fludarabine in a proportion of primary CLL samples treated with pro-proliferative stimuli, including those with TP53 disruption. Finally, we observed a fludarabine potentiation by SCH900776 in a T-cell leukemia 1 (TCL1)-driven mouse model of CLL. Collectively, we have substantiated the significant potential of Chk1 inhibition in B-lymphoid cells.

• Klíčová slova
• SCH900776, TP53, checkpoint kinase 1/Chk1, chronic lymphocytic leukemia, nucleoside analogs,
• MeSH
• apoptóza MeSH
• B-lymfocyty cytologie   MeSH
• buněčný cyklus MeSH
• checkpoint kinasa 1 antagonisté a inhibitory   MeSH
• chronická lymfatická leukemie farmakoterapie   genetika   metabolismus   MeSH
• cytarabin aplikace a dávkování   MeSH
• deoxycytidin aplikace a dávkování   analogy a deriváty   MeSH
• gemcitabin MeSH
• lidé MeSH
• mitóza MeSH
• mutace MeSH
• myši transgenní MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádorový supresorový protein p53 genetika   MeSH
• nukleosidy genetika   MeSH
• proliferace buněk MeSH
• pyrazoly farmakologie   MeSH
• pyrimidiny farmakologie   MeSH
• screeningové testy protinádorových léčiv MeSH
• signální transdukce MeSH
• viabilita buněk MeSH
• vidarabin aplikace a dávkování   analogy a deriváty   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• checkpoint kinasa 1 MeSH
• CHEK1 protein, human MeSH Prohlížeč
• cytarabin MeSH
• deoxycytidin MeSH
• fludarabine MeSH Prohlížeč
• gemcitabin MeSH
• MK-8776 MeSH Prohlížeč
• nádorový supresorový protein p53 MeSH
• nukleosidy MeSH
• pyrazoly MeSH
• pyrimidiny MeSH
• TP53 protein, human MeSH Prohlížeč
• vidarabin MeSH

• MeSH
• adenin analogy a deriváty   MeSH
• alely MeSH
• bicyklické sloučeniny heterocyklické terapeutické užití   MeSH
• chinazolinony terapeutické užití   MeSH
• chronická lymfatická leukemie diagnóza   farmakoterapie   genetika   patologie   MeSH
• cílená molekulární terapie MeSH
• exprese genu MeSH
• fosfatidylinositol-3-kinasy třídy I antagonisté a inhibitory   genetika   metabolismus   MeSH
• frekvence genu MeSH
• lidé MeSH
• nádorové biomarkery genetika   metabolismus   MeSH
• nádorový supresorový protein p53 nedostatek   genetika   MeSH
• piperidiny MeSH
• prognóza MeSH
• proteinkinasa BTK MeSH
• protinádorové látky terapeutické užití   MeSH
• puriny terapeutické užití   MeSH
• pyrazoly terapeutické užití   MeSH
• pyrimidiny terapeutické užití   MeSH
• sulfonamidy terapeutické užití   MeSH
• tyrosinkinasy antagonisté a inhibitory   genetika   metabolismus   MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH
• úvodníky MeSH
• Názvy látek
• adenin MeSH
• bicyklické sloučeniny heterocyklické MeSH
• chinazolinony MeSH
• fosfatidylinositol-3-kinasy třídy I MeSH
• ibrutinib MeSH Prohlížeč
• idelalisib MeSH Prohlížeč
• nádorové biomarkery MeSH
• nádorový supresorový protein p53 MeSH
• PIK3CD protein, human MeSH Prohlížeč
• piperidiny MeSH
• proteinkinasa BTK MeSH
• protinádorové látky MeSH
• puriny MeSH
• pyrazoly MeSH
• pyrimidiny MeSH
• sulfonamidy MeSH
• TP53 protein, human MeSH Prohlížeč
• tyrosinkinasy MeSH
• venetoclax MeSH Prohlížeč

TP53 gene defects represent a strong adverse prognostic 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 limitations of commonly used methods for TP53 mutation examination 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 chromatography (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 evaluated 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 predominated (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 FASAY is a suitable approach for mutation detection. Ultra-deep NGS has the potential to overcome shortcomings of methods currently used, allows the detection of minor proportion mutations, and represents thus a promising methodology for near future.

• MeSH
• chronická lymfatická leukemie genetika   MeSH
• dospělí MeSH
• geny p53 * MeSH
• hybridizace in situ fluorescenční MeSH
• jednonukleotidový polymorfismus MeSH
• lidé středního věku MeSH
• lidé MeSH
• mutace * MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• senioři MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• vysokoúčinná kapalinová chromatografie MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH