Besides many other mutations in known cancer driver genes, mantle cell lymphoma (MCL) is characterized by recurrent genetic alterations of important regulators of the phosphoinositol-3-kinase (PI3K) cascade including PIK3CA gains and PTEN losses. To evaluate the biological and functional consequences of these aberrations in MCL, we have introduced transgenic expression of PIK3CA (PIK3CA UP) and performed knockout/knockdown of PTEN gene (PTEN KO/KD) in 5 MCL cell lines. The modified cell lines were tested for associated phenotypes including dependence on upstream B-cell receptor (BCR) signaling (by an additional BCR knockout). PIK3CA overexpression decreased the dependence of the tested MCL on prosurvival signaling from BCR, decreased levels of oxidative phosphorylation, and increased resistance to 2-deoxy-glucose, a glycolysis inhibitor. Unchanged protein kinase B (AKT) phosphorylation status and unchanged sensitivity to a battery of PI3K inhibitors suggested that PIK3CA gain might affect MCL cells in AKT-independent manner. PTEN KO was associated with a more distinct phenotype: AKT hyperphosphorylation and overactivation, increased resistance to multiple inhibitors (most of the tested PI3K inhibitors, Bruton tyrosine kinase inhibitor ibrutinib, and BCL2 inhibitor venetoclax), increased glycolytic rates with resistance to 2-deoxy-glucose, and significantly decreased dependence on prosurvival BCR signaling. Our results suggest that the frequent aberrations of the PI3K pathway may rewire associated signaling with lower dependence on BCR signaling, better metabolic and hypoxic adaptation, and targeted therapy resistance in MCL.
- MeSH
- chemorezistence genetika MeSH
- cílená molekulární terapie MeSH
- fosfatidylinositol-3-kinasy třídy I * genetika metabolismus MeSH
- fosfatidylinositol-3-kinasy metabolismus MeSH
- fosfohydroláza PTEN * metabolismus genetika MeSH
- lidé MeSH
- lymfom z plášťových buněk * farmakoterapie genetika metabolismus MeSH
- nádorové buněčné linie MeSH
- protoonkogenní proteiny c-akt metabolismus MeSH
- receptory antigenů B-buněk metabolismus MeSH
- signální transdukce MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Cell surface expression of PD-1, PD-L1 and PD-L2 immune checkpoints on B and T cells obtained from patients with mantle cell lymphoma shows ambiguous results across many studies and creates obstacles for the implementation of immune checkpoint inhibitors into the therapy of mantle cell lymphoma. Using multiparameter flow cytometry we analysed surface expression of PD-1, PD-L1 and PD-L2 molecules on B and T cells of 31 newly diagnosed mantle cell lymphomas and compared it with the results of 26 newly diagnosed chronic lymphocytic leukaemias and 20 healthy volunteers. To gain insight into the age-dependent changes of surface expression of these immune checkpoints, flow cytometric subanalysis of 30 healthy volunteers of 25-93 years of age was conducted. Overall, we demonstrated weak surface expression of PD-1, PD-L1 and PD-L2 on B and T cells of mantle cell lymphoma patients (< 10 % when compared to healthy individuals). A significant age-dependent increase in the expression of PD-1 and its ligand PD-L2 was observed in healthy volunteers. Our results suggest that neither PD-1 nor its ligands represent relevant druggable targets for the therapy of mantle cell lymphoma. The observed age-dependent changes in healthy population could impact efficiency of immune checkpoint inhibitors and could be at least partly connected with increased incidence of cancer with age.
- MeSH
- antigeny CD273 metabolismus MeSH
- antigeny CD274 metabolismus MeSH
- antigeny CD279 metabolismus MeSH
- B-lymfocyty MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- lymfom z plášťových buněk metabolismus MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- studie případů a kontrol MeSH
- T-lymfocyty MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- Publikační typ
- časopisecké články MeSH
Mantle cell lymphoma (MCL) is a heterogeneous malignancy with a broad spectrum of clinical behavior from indolent to highly aggressive cases. Despite the fact that MCL remains in most cases incurable by currently applied immunochemotherapy, our increasing knowledge on the biology of MCL in the last two decades has led to the design, testing, and approval of several innovative agents that dramatically changed the treatment landscape for MCL patients. Most importantly, the implementation of new drugs and novel treatment algorithms into clinical practice has successfully translated into improved outcomes of MCL patients not only in the clinical trials, but also in real life. This review focuses on recent advances in our understanding of the pathogenesis of MCL, and provides a brief survey of currently used treatment options with special focus on mode of action of selected innovative anti-lymphoma molecules. Finally, it outlines future perspectives of patient management with progressive shift from generally applied immunotherapy toward risk-stratified, patient-tailored protocols that would implement innovative agents and/or procedures with the ultimate goal to eradicate the lymphoma and cure the patient.
- MeSH
- antitumorózní látky farmakologie terapeutické užití MeSH
- buněčný cyklus účinky léků genetika MeSH
- chemorezistence MeSH
- cílená molekulární terapie * metody MeSH
- genetická variace MeSH
- klonální evoluce účinky léků genetika MeSH
- kombinovaná terapie MeSH
- lidé MeSH
- lymfom z plášťových buněk farmakoterapie etiologie metabolismus mortalita MeSH
- náchylnost k nemoci MeSH
- nádorové biomarkery antagonisté a inhibitory MeSH
- prognóza MeSH
- receptory antigenů B-buněk metabolismus MeSH
- recidiva MeSH
- regulace genové exprese u nádorů účinky léků MeSH
- signální transdukce účinky léků MeSH
- výsledek terapie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Integral membrane proteins are under-represented in standard proteomic analyses, mostly because of their low expression and absence of trypsin-cleavage sites in their hydrophobic transmembrane segments. Novel and effective strategies for membrane proteomic analysis aim at soluble N-glycosylated segments of integral membrane proteins (CSC, SPEG, N-glyco-FASP) or selectively target the hydrophobic transmembrane alpha-helical segments employing chemical peptide cleavage by CNBr (hpTC). We combined a solid phase enrichment of glycopeptides (SPEG) with a transmembrane segment-oriented hpTC method and a standard "detergent and trypsin" approach into a three-pronged "Pitchfork" strategy to maximize the membrane proteome coverage in human lymphoma cells. This strategy enabled the identification of >1200 integral membrane proteins from all cellular compartments, including 105 CD antigens, 24 G protein-coupled receptors, and 141 solute carrier transporters. The advantage of the combination lies in the complementarity of the methods. SPEG and hpTC target different sets of membrane proteins. HpTC provided identifications of proteins and peptides with significantly higher hydrophobicity compared to SPEG and detergent-trypsin approaches. Among all identified proteins, we observed 32 so-called "missing proteins". The Pitchfork strategy presented here is universally applicable and enables deep and fast description of membrane proteomes in only 3 LC-MS/MS runs per replicate. SIGNIFICANCE: Integral membrane proteins (IMPs) are encoded by roughly a quarter of human coding genes. Their functions and their specific localization makes IMPs highly attractive drug targets. In fact, roughly half of the currently approved drugs in medicine target IMPs. Our knowledge of membrane proteomes is, however, limited. We present a new strategy for the membrane proteome analysis that combines three complementary methods targeting different features of IMPs. Using the combined strategy, we identified over 1200 IMPs in human lymphoma tissue from all sub-cellular compartments in only 3 LC-MS/MS runs per replicate. The three-pronged "Pitchfork" strategy is universally applicable, and offers a fast way toward a reasonably concise description of membrane proteomes in multiple samples.
- MeSH
- chromatografie kapalinová MeSH
- heterografty MeSH
- lidé MeSH
- lymfom z plášťových buněk metabolismus MeSH
- membránové proteiny metabolismus MeSH
- myši MeSH
- nádorové proteiny metabolismus MeSH
- proteom metabolismus MeSH
- tandemová hmotnostní spektrometrie MeSH
- transplantace nádorů MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
PURPOSE: Mantle cell lymphoma (MCL) is an aggressive subtype of B-cell non-Hodgkin lymphomas characterized by (over)expression of BCL2. A BCL2-targeting drug, venetoclax, has promising anticancer activity in MCL. We analyzed molecular mechanisms of venetoclax resistance in MCL cells and tested strategies to overcome it. EXPERIMENTAL DESIGN: We confirmed key roles of proapoptotic proteins BIM and NOXA in mediating venetoclax-induced cell death in MCL. Both BIM and NOXA are, however, differentially expressed in cell lines compared with primary cells. First, NOXA protein is significantly overexpressed in most MCL cell lines. Second, deletions of BIM gene harbored by three commonly used MCL cell lines (JEKO-1, MINO, and Z138) were not found by array comparative genomic hybridization using a validation set of 24 primary MCL samples. RESULTS: We demonstrated that MCL1 and NOXA play important roles in mediating resistance to venetoclax. Consequently, we tested an experimental treatment strategy based on cotargeting BCL2 with venetoclax and MCL1 with a highly specific small-molecule MCL1 inhibitor S63845. The combination of venetoclax and S63845 demonstrated synthetic lethality in vivo on a panel of five patient-derived xenografts established from patients with relapsed MCL with adverse cytogenetics. CONCLUSIONS: Our data strongly support investigation of venetoclax in combination with S63845 as an innovative treatment strategy for chemoresistant MCL patients with adverse cytogenetics in the clinical grounds.
- MeSH
- antitumorózní látky farmakologie MeSH
- bicyklické sloučeniny heterocyklické farmakologie MeSH
- chemorezistence MeSH
- lidé MeSH
- lokální recidiva nádoru farmakoterapie metabolismus patologie MeSH
- lymfom z plášťových buněk farmakoterapie metabolismus patologie MeSH
- myši inbrední NOD MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- protein MCL-1 antagonisté a inhibitory metabolismus MeSH
- protoonkogenní proteiny c-bcl-2 antagonisté a inhibitory metabolismus MeSH
- pyrimidiny farmakologie MeSH
- sulfonamidy farmakologie MeSH
- synergismus léků * MeSH
- thiofeny farmakologie MeSH
- xenogenní modely - testy antitumorózní aktivity MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Mantle cell lymphoma (MCL) is a mature B-cell lymphoma characterized by poor clinical outcome. Recent studies revealed the importance of B-cell receptor (BCR) signaling in maintaining MCL survival. However, it remains unclear which role MALT1, an essential component of the CARD11-BCL10-MALT1 complex that links BCR signaling to the NF-κB pathway, plays in the biology of MCL. Here we show that a subset of MCLs is addicted to MALT1, as its inhibition by either RNA or pharmacologic interference induced cytotoxicity both in vitro and in vivo. Gene expression profiling following MALT1 inhibition demonstrated that MALT1 controls an MYC-driven gene expression network predominantly through increasing MYC protein stability. Thus, our analyses identify a previously unappreciated regulatory mechanism of MYC expression. Investigating primary mouse splenocytes, we could demonstrate that MALT1-induced MYC regulation is not restricted to MCL, but represents a common mechanism. MYC itself is pivotal for MCL survival because its downregulation and pharmacologic inhibition induced cytotoxicity in all MCL models. Collectively, these results provide a strong mechanistic rationale to investigate the therapeutic efficacy of targeting the MALT1-MYC axis in MCL patients.
- MeSH
- buněčná smrt MeSH
- heterografty MeSH
- kaspasy metabolismus fyziologie MeSH
- lidé MeSH
- lymfom z plášťových buněk metabolismus MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- nádorové proteiny metabolismus fyziologie MeSH
- NF-kappa B metabolismus MeSH
- protein MALT1 MeSH
- protoonkogenní proteiny c-myc metabolismus MeSH
- receptory antigenů B-buněk fyziologie MeSH
- regulace genové exprese u nádorů MeSH
- signální transdukce MeSH
- viabilita buněk MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Richter syndrome represents the transformation of the chronic lymphocytic leukemia (CLL) into an aggressive lymphoma, most frequently the diffuse large B-cell lymphoma (DLBCL). In this report we describe a patient with CLL, who developed a clonally-related pleomorphic highly-aggressive mantle cell lymphoma (MCL) after five cycles of a fludarabine-based second-line therapy for the first relapse of CLL. Molecular cytogenetic methods together with whole-exome sequencing revealed numerous gene alterations restricted to the MCL clone (apart from the canonical t(11;14)(q13;q32) translocation) including gain of one copy of ATM gene or emergence of TP53, CREBBP, NUP214, FUBP1 and SF3B1 gene mutations. Similarly, gene expression analysis revealed vast differences between the MCL and CLL transcriptome, including overexpression of cyclin D1, downregulation of cyclins D2 and D3, or downregulation of IL4R in the MCL clone. Backtracking analysis using quantitative PCR specifically detecting an MCL-restricted focal deletion of TP53 revealed that the pre-MCL clone appeared in the bone marrow and peripheral blood of the patient approximately 4 years before the clinical manifestation of MCL. Both molecular cytogenetic and sequencing data support the hypothesis of a slow development of the pre-MCL clone in parallel to CLL over several years, and thereby exclude the possibility that the transformation event occurred at the stage of the CLL relapse clone by mere t(11;14)(q13;q32) acquisition.
- MeSH
- chronická lymfatická leukemie genetika metabolismus patologie MeSH
- difúzní velkobuněčný B-lymfom genetika metabolismus patologie MeSH
- hybridizace in situ fluorescenční MeSH
- imunohistochemie MeSH
- lidé středního věku MeSH
- lidé MeSH
- lidské chromozomy, pár 11 MeSH
- lidské chromozomy, pár 14 MeSH
- lymfom z plášťových buněk genetika metabolismus patologie MeSH
- nádorové biomarkery genetika metabolismus MeSH
- nádorový supresorový protein p53 genetika metabolismus MeSH
- translokace genetická MeSH
- ztráta heterozygozity MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
D-type cyclins are involved in cell cycle regulation and play an important role in the pathogenesis of lymphomas. Aberrant expression of cyclin D1 is associated with mantle cell lymphoma (MCL) and serves as a diagnostic marker of MCL. Analysis of cyclin D expression in tumor tissues of patients with diffuse large B-cell lymphoma (DLBCL) which comprises a heterogeneous group of tumors may contribute to their stratification. We analyzed expression of cyclin D1, D2, and D3 mRNAs in 30 MCL and 104 DLBCL patients using qRT-PCR and addressed their significance for disease outcome. We confirmed a high level of cyclin D1 mRNA in 29 MCL cases (97%). One case (3%) was identified as positive for cyclin D2. Expression of cyclin D1 was limited to MCL and did not occur in DLBCL. Overexpression of cyclin D2, which is rare in MCL, occurred more frequently in DLBCL (11 cases, 10.6%). We showed that high expression of cyclin D2 in DLBCL cases de novo decreased the overall survival rate (P=0.016) and progression-free survival (P=0.009). The expression pattern of cyclin D3 was similar in both types of studied lymphomas and it did not affect the disease outcome.
- MeSH
- cyklin D genetika metabolismus MeSH
- difúzní velkobuněčný B-lymfom metabolismus mortalita MeSH
- exprese genu MeSH
- Kaplanův-Meierův odhad MeSH
- lidé středního věku MeSH
- lidé MeSH
- lymfom z plášťových buněk metabolismus mortalita MeSH
- nádorové biomarkery genetika metabolismus MeSH
- senioři MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Mantle cell lymphoma (MCL) is a chronically relapsing aggressive type of B-cell non-Hodgkin lymphoma considered incurable by currently used treatment approaches. Fludarabine is a purine analog clinically still widely used in the therapy of relapsed MCL. Molecular mechanisms of fludarabine resistance have not, however, been studied in the setting of MCL so far. We therefore derived fludarabine-resistant MCL cells (Mino/FR) and performed their detailed functional and proteomic characterization compared to the original fludarabine sensitive cells (Mino). We demonstrated that Mino/FR were highly cross-resistant to other antinucleosides (cytarabine, cladribine, gemcitabine) and to an inhibitor of Bruton tyrosine kinase (BTK) ibrutinib. Sensitivity to other types of anti-lymphoma agents was altered only mildly (methotrexate, doxorubicin, bortezomib) or remained unaffacted (cisplatin, bendamustine). The detailed proteomic analysis of Mino/FR compared to Mino cells unveiled over 300 differentially expressed proteins. Mino/FR were characterized by the marked downregulation of deoxycytidine kinase (dCK) and BTK (thus explaining the observed crossresistance to antinucleosides and ibrutinib), but also by the upregulation of several enzymes of de novo nucleotide synthesis, as well as the up-regulation of the numerous proteins of DNA repair and replication. The significant upregulation of the key antiapoptotic protein Bcl-2 in Mino/FR cells was associated with the markedly increased sensitivity of the fludarabine-resistant MCL cells to Bcl-2-specific inhibitor ABT199 compared to fludarabine-sensitive cells. Our data thus demonstrate that a detailed molecular analysis of drug-resistant tumor cells can indeed open a way to personalized therapy of resistant malignancies.
- MeSH
- antitumorózní látky farmakologie MeSH
- chemorezistence * MeSH
- chromatografie kapalinová metody MeSH
- izotopové značení metody MeSH
- lidé MeSH
- lymfom z plášťových buněk farmakoterapie metabolismus MeSH
- nádorové biomarkery metabolismus MeSH
- nádorové buňky kultivované MeSH
- proteomika metody MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- vidarabin analogy a deriváty farmakologie MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
