Prognostic Significance of the Cell Cycle Inhibitor p27Kip1 in Chronic B-Cell Lymphocytic Leukemia

Blood ◽  
1998 ◽  
Vol 91 (12) ◽  
pp. 4694-4700 ◽  
Author(s):  
Radovan Vrhovac ◽  
Alain Delmer ◽  
Ruoping Tang ◽  
Jean-Pierre Marie ◽  
Robert Zittoun ◽  
...  
Keyword(s):  
Cell Cycle ◽  
B Cell ◽  
Doubling Time ◽  
Kinase Inhibitor ◽  
Prognostic Significance ◽  
Absolute Number ◽  
Lymphocytic Leukemia ◽  
Interleukin 4 ◽  
Resting Lymphocytes

Abstract B-cell chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of resting lymphocytes. The identification of p27kip1, a cyclin-dependent kinase inhibitor that contributes to cell cycle arrest and represents a link between extracellular signals and cell cycle, prompted us to study p27 protein in the lymphocytes from 88 patients with B-CLL and 32 patients with other chronic B-lymphoproliferative disorders. The expression of p27 protein was higher in B-CLL samples with variations among them. In B-CLL, p27 levels were independent of absolute number of circulating lymphocytes, but strongly correlated with both lymphocyte and total tumor mass (TTM) doubling time. High p27 expression was associated with a poorer overall prognosis. In vitro, there was an increased spontaneous survival of B-CLL cells expressing high p27 levels. Interleukin-4 (IL-4) upregulated p27 levels in B-CLL cells, while fludarabine decreased p27 levels. Thus, our results indicate that p27 may be a valuable kinetic marker in B-CLL by providing instantaneous estimation of the disease doubling time. In addition, these results suggest that there is a link between p27 expression and the ability of CLL cells to undergo apoptosis.

Prognostic Significance of the Cell Cycle Inhibitor p27Kip1 in Chronic B-Cell Lymphocytic Leukemia

Blood ◽  
1998 ◽  
Vol 91 (12) ◽  
pp. 4694-4700 ◽  
Author(s):  
Radovan Vrhovac ◽  
Alain Delmer ◽  
Ruoping Tang ◽  
Jean-Pierre Marie ◽  
Robert Zittoun ◽  
...  
Keyword(s):  
Cell Cycle ◽  
B Cell ◽  
Doubling Time ◽  
Kinase Inhibitor ◽  
Prognostic Significance ◽  
Absolute Number ◽  
Lymphocytic Leukemia ◽  
Interleukin 4 ◽  
Resting Lymphocytes

B-cell chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of resting lymphocytes. The identification of p27kip1, a cyclin-dependent kinase inhibitor that contributes to cell cycle arrest and represents a link between extracellular signals and cell cycle, prompted us to study p27 protein in the lymphocytes from 88 patients with B-CLL and 32 patients with other chronic B-lymphoproliferative disorders. The expression of p27 protein was higher in B-CLL samples with variations among them. In B-CLL, p27 levels were independent of absolute number of circulating lymphocytes, but strongly correlated with both lymphocyte and total tumor mass (TTM) doubling time. High p27 expression was associated with a poorer overall prognosis. In vitro, there was an increased spontaneous survival of B-CLL cells expressing high p27 levels. Interleukin-4 (IL-4) upregulated p27 levels in B-CLL cells, while fludarabine decreased p27 levels. Thus, our results indicate that p27 may be a valuable kinetic marker in B-CLL by providing instantaneous estimation of the disease doubling time. In addition, these results suggest that there is a link between p27 expression and the ability of CLL cells to undergo apoptosis.

scholarly journals Emerging role of kinase-targeted strategies in chronic lymphocytic leukemia

Hematology ◽  
2012 ◽  
Vol 2012 (1) ◽  
pp. 88-96 ◽  
Author(s):  
Adrian Wiestner
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Absolute Lymphocyte Count ◽  
Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Cell Trafficking

Abstract Chronic lymphocytic leukemia (CLL) is a malignancy of mature B cells that depend on host factors in the tissue microenvironment for survival and proliferation. In vitro, CLL cells rapidly undergo apoptosis unless microenvironmental factors are provided that support their survival. Signaling pathways activated in the microenvironment in vivo include the B-cell receptor (BCR) and NF-κB pathways. Thus, CLL is a disease “addicted to the host” and is dependent on pathways that promote normal B-cell development, expansion, and survival; this is particularly true in the case of the BCR signaling cascade. Small-molecule inhibitors of kinases that are essential for BCR signal transduction abrogate the stimulating effects of the microenvironment on CLL cells. The orally administered tyrosine kinase inhibitors fostamatinib and ibrutinib and the phosphatidylinositol 3-kinase inhibitor GS-1101 have induced impressive responses in relapsed and refractory CLL patients, mostly with moderate side effects. Reductions in lymphadenopathy and splenomegaly are seen within weeks and are frequently accompanied by a transient rise in absolute lymphocyte count that is asymptomatic and probably the result of changes in CLL cell trafficking. This review discusses the biologic basis for kinase inhibitors as targeted therapy of CLL and summarizes the exciting early clinical experience with these agents.

scholarly journals Emerging role of kinase-targeted strategies in chronic lymphocytic leukemia

Blood ◽  
2012 ◽  
Vol 120 (24) ◽  
pp. 4684-4691 ◽  
Author(s):  
Adrian Wiestner
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Absolute Lymphocyte Count ◽  
Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Cell Trafficking

AbstractChronic lymphocytic leukemia (CLL) is a malignancy of mature B cells that depend on host factors in the tissue microenvironment for survival and proliferation. In vitro, CLL cells rapidly undergo apoptosis unless microenvironmental factors are provided that support their survival. Signaling pathways activated in the microenvironment in vivo include the B-cell receptor (BCR) and NF-κB pathways. Thus, CLL is a disease “addicted to the host” and is dependent on pathways that promote normal B-cell development, expansion, and survival; this is particularly true in the case of the BCR signaling cascade. Small-molecule inhibitors of kinases that are essential for BCR signal transduction abrogate the stimulating effects of the microenvironment on CLL cells. The orally administered tyrosine kinase inhibitors fostamatinib and ibrutinib and the phosphatidylinositol 3-kinase inhibitor GS-1101 have induced impressive responses in relapsed and refractory CLL patients, mostly with moderate side effects. Reductions in lymphadenopathy and splenomegaly are seen within weeks and are frequently accompanied by a transient rise in absolute lymphocyte count that is asymptomatic and probably the result of changes in CLL cell trafficking. This review discusses the biologic basis for kinase inhibitors as targeted therapy of CLL and summarizes the exciting early clinical experience with these agents.

scholarly journals Responsiveness of chronic lymphocytic leukemia B cells activated via surface Igs or CD40 to B-cell tropic factors

Blood ◽  
1992 ◽  
Vol 80 (12) ◽  
pp. 3173-3181
Author(s):  
AC Fluckiger ◽  
JF Rossi ◽  
A Bussel ◽  
P Bryon ◽  
J Banchereau ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Lymphocytic Leukemia ◽  
Interleukin 4 ◽  
Aureus Strain ◽  
Cross Linking ◽  
Cell Recovery

Recent studies performed in the laboratory have established that interleukin-4 (IL-4) used in combination with anti-CD40 monoclonal antibody (MoAb) 89 presented on Ltk- mouse fibroblasts stably expressing human Fc gamma RII/CDw32 (referred to as the CD40 system) sustains long-term proliferation of normal human B cells. In the present study, B-cell chronic lymphocytic leukemias (B-CLLs) activated through slgs or CD40 were examined for their capacity to proliferate and differentiate in response to various cytokines. Our results indicate that the outcome of IL-4 stimulation on the in vitro growth of B-CLL depends on the signalling pathway used for their activation. Whereas IL-4 did not display any growth-stimulatory effect on B-CLL activated by Ig cross-linking agents, it could stimulate DNA synthesis and enhance the viable cell recovery when leukemic B cells were cultured in the CD40 system. Most B-CLL samples were induced for IgM synthesis upon Staphylococcus aureus strain Cowan I stimulation. This Ig response was potentiated by IL-2 and antagonized by IL-4. Anti-CD40 MoAb used alone or in combination with cytokines (IL-1 alpha to IL-6, interferon gamma, tumor necrosis factor gamma, and transforming growth factor beta) failed to induce Ig secretion from B-CLL cells. No evidence for Ig isotype switching was obtained with the cytokines listed above, regardless of the mode of activation. Taken together, our results suggest that B-CLL cells can be partially released from their apparent maturation block by IL-2 and Ig cross-linking agents. In contrast, combinations of IL-4 and cross-linked anti-CD40 antibodies induced entry of B-CLL cell into cycle, but poorly stimulated their differentiation into Ig secreting cells.

The RNA Binding Protein Musashi 2 Is up-Regulated in the Proliferative B-Cell Fraction of Chronic Lymphocytic Leukemia Clones

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4149-4149
Author(s):  
Florencia Palacios ◽  
Xiao-Jie Yan ◽  
Jaqueline C. Barrientos ◽  
Jonathan E. Kolitz ◽  
Steven L. Allen ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Cell Division ◽  
B Cells ◽  
B Cell ◽  
Lymphocytic Leukemia ◽  
Ki 67 ◽  
Post Transcriptional Regulation

Abstract Chronic Lymphocytic Leukemia (CLL) is an incurable disease in which most of the tumor cells in the blood are arrested in G0/G1 stages of the cell cycle with only a minimal number displaying proliferative activity. In this regard, our group has found by gene expression profiling (GEP) that the proliferative fraction (PF) of CLL cells is enriched in the intraclonal subset marked by CXCR4dim CD5brite expression. Indeed, this subset differs by more than 1000 genes from the CXCR4brite CD5dim resting fraction (RF). The genes over-expressed in the PF relate to replication and migration as well as regulation of gene expression. One of these genes is Musashi 2 (MSI2). Of note, MSI2 is expressed at the highest levels in IGHV unmutated CLL (U-CLL) clones and their PFs. Normally, MSI2 binds mRNA and blocks translation of proteins, playing an important role in post-transcriptional regulation. In addition, MSI2 has been linked to proliferation of normal and malignant stem cells, tumorigenesis, and poor prognosis. In CLL, high MSI2 mRNA expression has been identified in patients with worse prognosis. Nevertheless, nothing is known about the function of MSI2 in CLL cells. Therefore, we have begun to study the biological role of MSI2 in B-CLL cells and its possible association with B-cell proliferation and CLL disease progression. Fist, we studied MSI2 protein expression by flow cytometry in CD19+ B cells from healthy donors (HD) and CD5+CD19+ cells from CLL patients, observing an up-regulation in CLL compared to HD. Also, we documented higher MSI2 expression in U-CLL compared to IGHV-mutated (M-CLL) CLL as well as HD. Within the leukemic clone, we observed that MSI2 expression was highest in the PF, lower in the intermediate (INT) fraction (defined as CXCR4int CD5int), and much lower in the RF (PF>INT>RF). The PF expressed 40% more MSI2 than the RF, suggesting that the highest amounts of MSI2 protein is in dividing and recently-divided cells of the clone. Since CLL B-cell proliferation occurs in the microenvironment of lymphoid organs, presumably delivered by external signals, we tested whether such signals could stimulate MSI2 expression. After stimulating CLL cells with TLR9 agonist + IL15 + IL2 in vitro MSI2 protein was up-regulated form 0.3 to 2.5 fold. In addition, the increase in MSI2 protein was associated with an enhancement in Ki-67+ cells and in phosphorylation of MAPK/ERK and AKT signaling components, measured by flow cytometry. These results suggest that signals from the microenvironment that induce cell growth and proliferation lead to MSI2 synthesis in CLL B cells. In order to study a possible association between MSI2 expression and cell division, we labeled CLL PBMCs with a dilutable cell tracer, CFSE, and then stimulated them in vitro with TLR9 agonist + IL15 + IL2. These studies indicated that MSI2 protein synthesis was increased in the activated cells and that MSI2 protein levels increased with each cell division. However, it was also clear that this increase was not directly associated to the extent of cell replication as CLL B cells from only 10% of the patients underwent 4 cycles of cell division. Since we observed an increase in MSI2 and Ki-67 expression after stimulation in all patients' clones but did not detect replication of CLL cells in all patients, we studied the effects of in vitro stimulation on cell cycle entry and completion and how this related to MSI2 expression. Experiments using propidium iodide to evaluate DNA content of PBMCs showed that in vitro stimulation increased the percentage of cells in S phase (5-25%) compared to control cells without activation (<5%), whereas only a small fraction of cells entered the M/G2 phases, with or without activation (<1% and <0.5%, respectably) suggesting that only a small portion of the cells completed the cell cycle and divided. Hence, MSI2 synthesis corresponds with DNA replication and not cell division, suggesting that MSI2 could be an important molecule involved in entry into and/or in the early phases of the cell cycle. These results, and the facts that MSI2 plays an important role in post-transcriptional regulation and is associated with cell proliferation and poor prognosis in cancer, suggest that a better understanding of the role of MSI2 in CLL patients will provide clues to understanding the birth and growth of CLL B cells and to identifying and designing new therapeutic strategies for the disease. Disclosures No relevant conflicts of interest to declare.

scholarly journals Acalabrutinib: A Selective Bruton Tyrosine Kinase Inhibitor for the Treatment of B-Cell Malignancies

2021 ◽  
Vol 11 ◽  
Author(s):  
Hussein A. Abbas ◽  
William G. Wierda
Keyword(s):  
Adverse Events ◽  
Tyrosine Kinase ◽  
B Cell ◽  
Kinase Inhibitor ◽  
Progression Free Survival ◽  
Lymphocytic Leukemia ◽  
B Cell Malignancies ◽  
Bruton Tyrosine Kinase ◽  
Btk Inhibitor

Bruton tyrosine kinase (BTK) is a validated target for treatment of B-cell malignancies, and oral inhibitors of BTK have emerged as a standard of care for these diseases. Acalabrutinib is a second generation, highly selective, potent, covalent BTK inhibitor that exhibits minimal off-target activity in in vitro assays, providing the potential to improve tolerability over the first-in-class BTK inhibitor, ibrutinib. Acalabrutinib was approved for the treatment of relapsed/refractory mantle cell lymphoma (MCL) and chronic lymphocytic leukemia (CLL) in the US in 2017 and 2019, respectively. Acalabrutinib is also undergoing trials for other B-cell malignancies, both as monotherapy and in combinations. In this review, we discuss results from clinical trials evaluating the efficacy and safety of acalabrutinib in patients with CLL, MCL, and Waldenstrom’s macroglobulinemia. Recent phase 3 data showed that acalabrutinib improved progression-free survival (PFS) compared with rituximab plus idelalisib or rituximab plus bendamustine in patients with relapsed/refractory CLL, and acalabrutinib with or without obinutuzumab improved PFS compared with chlorambucil plus obinutuzumab in patients with treatment-naïve CLL. Overall, acalabrutinib had a tolerable safety profile, with most adverse events being grade 1/2 severity (most commonly headache and diarrhea) and a low rate of discontinuation due to adverse events.

The Novel Cancer Drug Seliciclib Engages the Mitochondrial Apoptosis Pathway Via the Mcl-1/Noxa Axis in CLL.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2983-2983
Author(s):  
Delfine Hallaert ◽  
Rene Spijker ◽  
Ingrid A.M. Derks ◽  
Margot Jak ◽  
Bart de Goeij ◽  
...  
Keyword(s):  
B Cell ◽  
Rna Polymerase Ii ◽  
Kinase Inhibitor ◽  
Lymphocytic Leukemia ◽  
Gene Profiling ◽  
Cancer Drug ◽  
Caspase 9 ◽  
Apoptotic Pathway ◽  
Apoptosis Pathway

Abstract Seliciclib (cyc202, R-roscovitine) is a novel cyclin dependent kinase inhibitor currently in pre-clinical testing. It indirectly inhibits RNA polymerase II and has been shown to be effective against B-cell malignancies by inducing apoptosis and Mcl-1 degradation. The aim of this study was to dissect the underlying apoptotic pathway(s) of seliciclib in chronic lymphocytic leukemia (B-CLL). Treatment of CLL cells with &gt;10 μM seliciclib in vitro induced apoptosis within 24 hours, irrespective of IgVH mutation status (n=20). Gene profiling by means of RT-MLPA, did not reveal involvement of p53 as indicated by the absence of Puma upregulation. None of the &gt;30 other apoptosis genes tested were induced; instead signals for certain labile mRNAs (Mcl-1, A1/Bfl-1, PI-9) were clearly decreased upon seliciclib. Detailed investigation of B cell lines overexpressing various anti-apoptotic proteins (Bcl-2, caspase-9-DN, Flip, FADD-DN), indicated that seliciclib activated the mitochondrial but not the death receptor pathway. Neither mitochondrial activation nor the rapid degradation of Mcl-1 protein could be prevented by caspase inhibition (zVAD) or overexpression of inactive caspase-9 (DN). This indicated that Mcl-1 decline is an upstream, caspase-independent event. Immuno-precipitation demonstrated that pro-survival Mcl-1 is engaged by pro-apoptotic Noxa and Bim. The specific contribution of Noxa was confirmed by RNAi, resulting in inhibition of apoptosis after seliciclib, but not after staurosporin, CD95- or BCR triggering. These findings demonstrate that seliciclib induces rapid, p53-independent apoptosis via Mcl-1 degradation and mitochondrial activation. The involvement of Noxa, which is highly expressed in CLL, suggests this BH3-only protein may be a novel therapeutic target.

Pharmacological Inhibition of PIM Kinases In Chronic Lymphocytic Leukemia Cases with Unfavorable Prognosis Markers

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2468-2468
Author(s):  
Beatriz Herreros ◽  
James R. Bischoff ◽  
Maria E. Rodriguez ◽  
Margarita Sanchez-Beato ◽  
Elena Domenech ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Therapeutic Target ◽  
Kinase Inhibitor ◽  
Annexin V ◽  
B Cell Lymphoma ◽  
Lymphocytic Leukemia ◽  
Pim Kinases

Abstract Abstract 2468 Introduction: Pim kinases are a family of oncogenic kinases that has been demonstrated to play a role in B-cell development and lymphomagenesis, and proposed as Chronic Lymphocytic Leukemia (CLL) therapeutic target. An increased expression of PIM1 and PIM2 has been found in subsets of CLL and other B-cell lymphoma types. In order to further explore PIM inhibition as a rational therapeutic target, we decided to test a new pan-Pim kinase inhibitor, ETP-39010, developed by the Experimental Therapeutics Programme at the CNIO. Materials and methods: Blood samples from 16 CLL patients were collected by the Tumor Bank Unit at the CNIO. Samples were processed in order to separate B cells (RosetteSep® Human B cell enrichment Cocktail, Stem cell Technologies). Sensitivity to the compound was analyzed by EC50 calculations using the Cell Titer Glo® commercial kit from Promega. Gene expression data were normalized and preprocessed using GEPAS utility available at http://gepas.bioinfo.cipf.es/. Differentially expressed genes and pathways were obtained using T-Rex (GEPAS) or GSEA (http://www.broadinstitute.org/gsea/) bioinformatic tools respectively. Apoptosis was measured by Annexin V/ PI staining and cell cycle was studied by PI staining using flow cytometry. Results: A significant variability in ETP-39010 sensitivity was found in this series of B-CLL samples, with EC50 values ranging from 1,3 nM to 22,6 nM, consistently with the heterogeneous PIM expression levels observed in CLL cases. A higher sensitivity to the compound was identified in samples with markers of unfavorable outcome, such as ZAP70 positivity (p<0.05) or unmutated IGHV (p=0.09). The series was divided into sensitive and resistant samples according to the EC50 value (above or below the median of the series). Gene expression studies showed that sensitive samples expressed higher levels of LPL, another prognosis marker that has been shown to be related with IGVH somatic mutation load. Pharmacodynamic studies demonstrated that ETP-39010 was able to induce apoptosis in sensitive CLL samples, without cell cycle changes. Furthermore, treatment with ETP-39010 was related with the downregulation of genes involved in protein metabolic processes, transport, signal transduction and cellular biosynthetic processes (T-Rex and Babelomics analysis), as well as to downregulation of several pathways related to metabolism (GSEA analysis). Conclusion: Inhibition of PIM kinases by ETP-39010 induces apoptosis in CLL samples. An increased sensitivity to PIM inhibition has been found in CLL cases with unfavourable prognostic markers, such as unmutated immunoglobulin heavy chain (IGHV) and increased expression of ZAP70 and LPL, pointing out PIM kinases as a potential therapeutic target for unmutated CLL. Disclosures: Garcia-Marco: ROCHE: Consultancy, Honoraria, Research Funding.

scholarly journals Responsiveness of chronic lymphocytic leukemia B cells activated via surface Igs or CD40 to B-cell tropic factors

Blood ◽  
1992 ◽  
Vol 80 (12) ◽  
pp. 3173-3181 ◽  
Author(s):  
AC Fluckiger ◽  
JF Rossi ◽  
A Bussel ◽  
P Bryon ◽  
J Banchereau ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Lymphocytic Leukemia ◽  
Interleukin 4 ◽  
Aureus Strain ◽  
Cross Linking ◽  
Cell Recovery

Abstract Recent studies performed in the laboratory have established that interleukin-4 (IL-4) used in combination with anti-CD40 monoclonal antibody (MoAb) 89 presented on Ltk- mouse fibroblasts stably expressing human Fc gamma RII/CDw32 (referred to as the CD40 system) sustains long-term proliferation of normal human B cells. In the present study, B-cell chronic lymphocytic leukemias (B-CLLs) activated through slgs or CD40 were examined for their capacity to proliferate and differentiate in response to various cytokines. Our results indicate that the outcome of IL-4 stimulation on the in vitro growth of B-CLL depends on the signalling pathway used for their activation. Whereas IL-4 did not display any growth-stimulatory effect on B-CLL activated by Ig cross-linking agents, it could stimulate DNA synthesis and enhance the viable cell recovery when leukemic B cells were cultured in the CD40 system. Most B-CLL samples were induced for IgM synthesis upon Staphylococcus aureus strain Cowan I stimulation. This Ig response was potentiated by IL-2 and antagonized by IL-4. Anti-CD40 MoAb used alone or in combination with cytokines (IL-1 alpha to IL-6, interferon gamma, tumor necrosis factor gamma, and transforming growth factor beta) failed to induce Ig secretion from B-CLL cells. No evidence for Ig isotype switching was obtained with the cytokines listed above, regardless of the mode of activation. Taken together, our results suggest that B-CLL cells can be partially released from their apparent maturation block by IL-2 and Ig cross-linking agents. In contrast, combinations of IL-4 and cross-linked anti-CD40 antibodies induced entry of B-CLL cell into cycle, but poorly stimulated their differentiation into Ig secreting cells.

B Cell Receptor signaling and genetic lesions in TP53 and CDKN2A/CDKN2B cooperate in Richter Transformation

Blood ◽  
2021 ◽  
Author(s):  
Supriya Chakraborty ◽  
Claudio Martines ◽  
Fabiola Porro ◽  
Ilaria Fortunati ◽  
Alice Bonato ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
B Cell ◽  
Critical Role ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Richter Syndrome

B cell receptor (BCR) signals play a critical role in the pathogenesis of chronic lymphocytic leukemia (CLL), but their role in regulating CLL cell proliferation has still not been firmly established. Unlike normal B cells, CLL cells do not proliferate in vitro upon engagement of the BCR, suggesting that CLL cell proliferation is regulated by other signals from the microenvironment, such as those provided by Toll-like receptors or T cells. Here, we report that BCR engagement of human and murine CLL cells induces several positive regulators of the cell cycle, but simultaneously induces the negative regulators CDKN1A, CDKN2A and CDKN2B, which block cell cycle progression. We further show that introduction of genetic lesions that downregulate these cell cycle inhibitors, such as inactivating lesions in CDKN2A, CDKN2B and the CDKN1A regulator TP53, leads to more aggressive disease in a murine in vivo CLL model and spontaneous proliferation in vitro that is BCR-dependent but independent of costimulatory signals. Importantly, inactivating lesions in CDKN2A, CDKN2B and TP53 frequently co-occur in Richter syndrome, and BCR stimulation of human Richter syndrome cells with such lesions is sufficient to induce proliferation. We also show that tumor cells with combined TP53 and CDKN2A/2B abnormalities remain sensitive to BCR inhibitor treatment and are synergistically sensitive to the combination of a BCR and CDK4/6 inhibitor both in vitro and in vivo. These data provide evidence that BCR signals are directly involved in driving CLL cell proliferation and reveal a novel mechanism of Richter transformation.

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