Lymphoplasmacytic Differentiation Associated with Lenalidomide Therapy in Patients with Chronic Lymphocytic Leukemia.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4412-4412
Author(s):  
Marays Veliz ◽  
Lynn C. Moscinski ◽  
Ling Zhang ◽  
Eduardo M. Sotomayor ◽  
Javier Pinilla-Ibarz
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Mechanism Of Action ◽  
Conflicts Of Interest ◽  
Selection Effect ◽  
Lymphocytic Leukemia ◽  
Significant Activity ◽  
Immune Mediated ◽  
Immune Synapses ◽  
Initiation Of Therapy

Abstract Abstract 4412 Background Due to significant activity of lenalidomide in chronic lymphocytic leukemia (CLL), research is ongoing to better characterize its mechanism of action. Recent laboratory studies of cells from CLL patients have shown that lenalidomide could exert its antiproliferative activity by decreasing the presence of survival-promoting cytokines. It has also been shown to facilitate immune-mediated ADCC by restoring the defective ability of T- and NK-cells to form immune synapses with tumor B-cells (Ramsay et al JCI 2008). However, no direct cytotoxic effect of lenalidomide has yet been demonstrated in CLL cells. Here, we suggest a new potential mechanism of action based on morphologic evaluation of bone marrow samples from patients with CLL receiving treatment with lenalidomide. Methods Bone marrow biopsy and aspirations from 13 patients with relapsed or refractory CLL, who are participating in a phase II clinical trial of lenalidomide and rituximab, and 1 patient who received lenalidomide outside of study, were reviewed before initiation of therapy and every 2 months thereafter. Results Of 14 CLL samples evaluated, 3 samples contained cells with lymphoplasmacytoid features in both bone marrow (figure 1) and peripheral blood, which had not been present prior to initiation of treatment. Of the 3 samples containing lymphocytes with plasmacytoid features, 2 were obtained from patients who received more than 6 months of therapy with the lenalidomide/rituximab combination, and 1 was taken from the patient receiving lenalidomide, after one month of therapy. Long term morphologic effects of lenalidomide therapy on lymphocytes from patients with CLL remain unknown. However, we hypothesize that the observed changes in morphology after lenalidomide treatment could represent either a differentiation phenomenon, as a potential antitumor mechanism in CLL cells, or perhaps represent just a selection effect. Conclusions An exact direct antitumor mechanism of lenalidomide on CLL cells remains unknown. Our observations suggest that lenalidomide may promote terminal differentiation of lymphocytes thus proposing an additional mechanism by which lenalidomide exerts its anti-tumor activity in CLL. Disclosures: No relevant conflicts of interest to declare.

Erythroid Differentiation Regulator 1 (ERDR1) From Nurse-Like Cells Promotes the Survival of Chronic Lymphocytic Leukemia Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1372-1372
Author(s):  
Hendrik W. Van Deventer ◽  
Robert Mango ◽  
Jonathan Serody
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Conflicts Of Interest ◽  
Erythroid Differentiation ◽  
Mesenchymal Cells ◽  
Lymphocytic Leukemia ◽  
Leukemia Cells ◽  
Wild Type

Abstract Abstract 1372 Background: Chemotherapy resistance in chronic lymphocytic leukemia (CLL) can be mediated by anti-apoptotic signals produced by stromal or nurse-like cells. Developing strategies to overcome this resistance is hindered by the lack of suitable “stromal” targets responsible for these signals. We have discovered that erythroid differentiation regulator 1 (ERDR1) may be a candidate target for such a strategy. In this study, we show Erdr1 is generated by several stromal cell types including bone marrow stromal cells, fibrocytes, and nurse-like cells. Furthermore, inhibition of stroma-generated Erdr1 results in increased apoptosis of co-cultured CLL cells. Methods/Results: We initially identified Erdr1 on an Affymetrix array that compared the gene expression of wild type and CCR5-/- mice with pulmonary metastasis. The increased expression of Erdr1 in the wild type mice was particularly pronounced in the pulmonary mesenchymal cells. Therefore, these cells were transfected with one of two shRNAs (shRNA #9 or shRNA#11) and the survival of these cells was compared with mesenchymal cells transfected with a non-targeted control vector. After 15 days in culture, the control cells expanded normally; however, no significant expansion was seen in either the shRNA#9 or shRNA#11 transfected cells. These differences in cellular expansion were associated with differences in apoptosis. 21.4+1.6% of the Erdr1 knockdown cells were annexin V+ compared to 11.2+1.9% of the non-targeted control (p<0.03). Using GFP as a marker for transfection, we were also able to show that knockdown of Erdr1 increased the apoptosis of surrounding non-transfected mesenchymal cells. Thus, Erdr1 is a critical protein for the survival of stromal cells. Further analysis of the mesenchymal cell subpopulations revealed the greatest expression of Erdr1 in the CD45+, thy1.1+/− fibrocytes. When compared to CD45- fibroblasts, the fibrocytes expressed CCR5 and increased Erdr1 expression by 14.2+/−2.9 fold when treated with the CCR5 ligand CCL4. Given the similarities between fibrocytes and nurse-like cells, we went on to measure the effect of Erdr1 inhibition on CLL cells. In these experiments, stable Erdr1 knockdown and control clones were selected after the transfection of the bone marrow stromal cell line M2-10B4. These clones were then co-cultured with primary CLL cells. At 96 hours, leukemia cells co-cultured with the control lines had expanded by 1.33 + 0.9 compared to 0.74 + 0.22 fold in the knock-down lines (p<0.03). As before, the lack of cellular expansion was associated with an increase in apoptosis. To further show the relevance of these findings to CLL, we demonstrated that human fibrocytes and nurse-like cells expressed mRNA and protein for ERDR1 in all patient samples tested. Implications for the treatment of human disease: Our data demonstrate that ERDR1 is a critically important protein for the survival of nurse-like cells. These data suggest that targeting ERDR1 or the upstream pathway through CCR5 might be a novel approach for the treatment of CLL. Disclosures: No relevant conflicts of interest to declare.

Heat-Shock Protein Signature Is Associated with Refractory Chronic Lymphocytic Leukemia Cells From Different In Vivo Microenvironments,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3866-3866
Author(s):  
Payal Gupta ◽  
Amit K. Mittal ◽  
Dennis D Weisenburger ◽  
Philip Bierman ◽  
Shantaram S Joshi
Keyword(s):  
Bone Marrow ◽  
Heat Shock ◽  
Chronic Lymphocytic Leukemia ◽  
Heat Shock Protein ◽  
Peripheral Blood ◽  
Treatment Cycle ◽  
Conflicts Of Interest ◽  
Lymphocytic Leukemia ◽  
Activation Markers ◽  
Protein Signature

Abstract Abstract 3866 Chronic Lymphocytic Leukemia (CLL) is a monoclonal B-cell disorder with accumulation of leukemic cells in peripheral blood, bone marrow and lymphoid organs. It presents with a heterogeneous clinical course. Many patients survive long periods of time without any need for treatment, whereas other patients show resistance to treatment or relapse soon after administration of therapy. Although some prognostic markers such as mutational status of immunoglobulin variable heavy chain, chromosomal abnormalities, CD38 levels, or ZAP-70 expression may help predict at initial diagnosis which patients will have more aggressive disease, the exact factors that can determine chances of remission in CLL are still not clear, making treatment challenging. Furthermore, CLL remains an incurable disease, necessitating a way for controlling its progression. Identifying novel molecular signatures associated with refractory CLL disease may help devise targeted treatment strategies and thus may prolong survival times and prevent the progression of CLL in relapsed patients. Considering this, we performed gene expression profiling (GEP) on peripheral blood (PB), bone marrow (BM) and lymph node (LN) samples collected at the time of diagnosis. We divided CLL samples into 3 groups based on their response to treatment; i) Stable CLL group: asymptomatic patients requiring no treatment, ii) Treated but stable CLL group: patients required treatment but had stable disease for at least one year after the end of the treatment cycle, and iii) Relapsed CLL: patients who relapsed within a year of end of the treatment cycle. Significance analysis of microarray (SAM) revealed that the heat-shock protein (HSP) signature (HSJ2, HSP70, HSP90, HSP60, HSP10, HSP 105, HSP40, HSP27, HSPA2, HSJ1, HSF4, HSPCA), BCR signaling pathway (JUN, NFATC4, NFKBIE, PPP3CB, TRAF3, CD81, CCT4), activation markers (CD81, CD83) and MMPs (MMP3, MMP9) were overexpressed in relapsed PB-CLL (n=3) compared to stable PB-CLL (n=6) and treated but stable PB-CLL (n=10). Overexpression of heat-shock protein signature genes were further observed in additional relapsed PB-CLL (n=6) group compared to other two PB-CLL (n=22) group. Interestingly, the HSP signature was consistently overexpressed in relapsed BM-CLL (n=6) and LN-CLL (n=12) compared to stable and treated but stable BM-CLL (n=11) and LN-CLL (n=3) groups. HSPs are considered chaperones of tumorigenesis and known to enhance survival, migration, and proliferation of tumor cells which may contribute to relapse in patients. Furthermore, the HSPs genes (HSP90 and HSP70) were significantly overexpressed in LN-CLL as compared to PB-CLL which implies important role of the microenviroment in rendering CLL refractory. To investigate the link between the expression of the individual genes with the aggressiveness of the disease, Kaplan-Meier log-rank tests were performed. We found that the higher expression of HSP90A, HSP90B, HSJ, and MMP9 were significantly (p<0.05) associated with shorter time to treatment. In summary, our study suggests that HSP genes are overexpressed in refractory CLL patients and thus are promising targets to improve clinical outcome. Disclosures: No relevant conflicts of interest to declare.

Ibrutinib (PCI 32765) Rapidly Improves Platelet Counts in Chronic Lymphocytic Leukemia / Small Lymphocytic Lymphoma (CLL/SLL) Patients and Has Minimal Effects On Platelet Aggregation

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1789-1789 ◽  
Author(s):  
Mohammed Farooqui ◽  
Jay Nelson Lozier ◽  
Janet Valdez ◽  
Nakhle Saba ◽  
Ajunae Wells ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Chronic Lymphocytic Leukemia ◽  
Conflicts Of Interest ◽  
Thrombus Formation ◽  
Lymphocytic Leukemia ◽  
Significant Activity ◽  
Small Lymphocytic Lymphoma ◽  
Open Label ◽  
Platelet Counts ◽  
Pre Treatment

Abstract Abstract 1789 INTRODUCTION: Ibrutinib (PCI 32765) is an orally administered covalent inhibitor of Bruton's Tyrosine Kinase (BTK). Ibrutinib has significant activity in chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) and is typically well tolerated (Byrd ASCO 2011, O'Brien ASH 2011). Rarely serious bleeding in patients concurrently on oral anticoagulation has been reported but was not related to thrombocytopenia (O'Brien ASH 2011). However, grade 1 or 2 ecchymosis/contusion is a frequent adverse event in patients on ibrutinib. In addition to being essential for B cell receptor signaling BTK is also involved in the signaling of the glycoprotein (GP)VI and GPIV von Willebrand (vW) receptors (Liu, Blood 2006). Thus, it is possible that ibrutinib could increase the bleeding risk by interfering with thrombus formation. In addition, lymphoproliferative disorders and some drugs have been associated with acquired vW-disease (AvWD). METHODS AND PATIENTS: In an ongoing single center, open label phase II trial we treat CLL/SLL patients with ibrutinib 420 mg daily on 28 day cycles (NCT01500733). We measured platelet (PLT) function on the PFA-100 instrument, vW-factor (vWF) antigen levels and activity (vWF-Ag/vWF-Act), and factor VIII (FVIII) on baseline, days 2 and 28. Here we report on effects of ibrutinib on platelet counts and function in 25 patients who completed >2 cycles. RESULTS: PLT counts prior to treatment ranged from 36 k/μl to 256 k/μl with a median of 102 k/μl. Twelve (48%) patients had a pre-treatment PLT count <100 k/μl. Median PLT counts for days 14, 28, and 56 increased to 140, 137, and 135 k/μl, respectively (P<.01). 76% of patients showed an increase after only 2 weeks on drug (median increase 25 k/μl (range 4–183 k/μl) that was sustained at subsequent timepoints. On day 14, 6 patients (24%) had a decrease in PLT count by a median of 13 k/μl from baseline; of these, 3 had a pre-treatment PLT count of <100 k/ul and 1 developed grade III thrombocytopenia (42 k/μl) that resolved to >100 k/μl by day 56. 20% (5 of 25) of patients reported grade 1 spontaneous ecchymosis with no correlation to platelet count, PFA testing, or vWF measurements. Of note we performed lymph node core biopsies in 35 patients taking ibrutinib with minimal bruising. Only 2 patients had more extensive local bruising/ecchymosis at the biopsy site. In 19 patients PFA-100 measurements of epinephrine (EPI) and adenosine diphosphate (ADP) stimulated platelet aggregation times were available (test requires PLT count >100 k/μl). Median changes in closure times with EPI and ADP on treatment were not significantly different from baseline (See table). Four (21%) patients started with abnormally prolonged EPI closure times (one on aspirin, one on ibuprofen; discontinued with the start of ibrutinib) which resolved by day 28 in 3 and decreased in 1. Three (16%) patients had a prolongation of EPI closure times on day 2 that resolved by day 28 in 2 and decreased in 1. All closure times on ADP were low or normal. No patients with abnormal PFA testing demonstrated spontaneous ecchymosis. From baseline to day 28 vWF-Act, vWF-Ag and FVIII decreased (P<0.05; n=24). All 3 values were high normal to elevated prior to treatment and decreased to normal on treatment. CONCLUSION: This preliminary report does not identify any significant ibrutinib effect on platelet function. PLT counts improved rapidly in the majority of patients and when seen transient decreases have been minimal. Three patients (16%) developed an abnormal reading in PLT function tests on treatment but none developed spontaneous echymosis or bleeding. The observed normalization of mildly elevated baseline levels of vWF and FVIII seems most consistent with a reduction in acute phase reactants and there was no evidence for AvWD on ibrutinib. The apparent functional tolerance of BTK inhibition in platelets is likely attributable to redundancy in the affected signaling pathways. This work was supported by the Intramural Research Program of NHLBI, NIH. We thank our patients for participating in these research studies. Disclosures: No relevant conflicts of interest to declare.

scholarly journals A Patient with Chronic Lymphocytic Leukemia and Bone Localization: A Case Report

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 5301-5301
Author(s):  
Iolanda Donatella Vincelli ◽  
Patrizia Cufari ◽  
Said al Sayyad ◽  
Carmelo Tuscano ◽  
Natale Porta ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Bone Marrow Biopsy ◽  
Peripheral Blood ◽  
Conflicts Of Interest ◽  
White Blood Cells ◽  
Lymphocytic Leukemia ◽  
Foot Pain ◽  
Staging Systems ◽  
The Right

Abstract Metastatic disease of the bone is a rare complication of chronic lymphocytic leukemia (CLL), it may be result from richter's transformation or metastatic from non lymphoid malignancies. CLL is the most common form of adult leukemia, with the median age of 70 years at diagnosis [Siegel et al. 2013]. The diagnosis is established by blood counts, blood smears, and immunophenotyping of circulating B-lymphocytes.The result is the increased number of lymphocytes in the peripheral blood, leukocytosis with absolute lymphocytosis, the increase of the lymphnodes, the increase in size of the spleen. The diagnosis of chronic lymphocytic leukemia B requires the presence of Clonal B cells in the peripheral blood at or above 5,000 / ul for at least 3 months. Typing immunophenotypical pathological lymphocytes are positive for surface antigens CD5, CD19, CD23, weakly positive for CD20 and CD22, generally negative FMC7 and CD79b; also expressing surface immunoglobulins. The Rai and Binet staging systems, which are established by physical examination and blood counts, have been recognized as standards for deciding whether to begin treatment. Patients with active or symptomatic disease or with advanced Binet or Rai stages require therapy. For fit patients, chemoimmunotherapy with fludarabine, cyclophosphamide, and rituximab represents the current standard therapy. For unfit patients, treatment with an anti-CD20 antibody (obinutuzumab, rituximab, ofatumumab) plus a milder chemotherapy (Chlorambucil) may be applied. At relapse, if the treatment-free interval exceeds two to three years, the initial treatment may be repeated, if the disease relapses earlier, drugs such as bendamustine (plus rituximab), alemtuzumab, lenalidomide, ofatumumab, ibrutinib, or idelalisib, must be choosen. Patients with a del(17p) or TP53 mutation can be treated with ibrutinib or a combination of idelalisib and rituximab. in relapsing patients with TP53 mutations or del(17p) or patients that are refractory to repeated chemoimmunotherapies, an allogeneic SCT may be considered [Hallek M 2015]. In this article we show a case of a 66-year-old man with CLL and a bone localization. In 2011 diagnosis of CLL, Rai Stage 0, Binet Stage A. Principal characteristics at diagnosis: HB 13.2 g /dl, White Blood Cells 15.800 / mm3, lymphocytes 61%, neutrophils 32%, monocytes 4%, platelets 141.000/mm3; normal hepatic end renal function; flowcytometric immunophenotyping of the peripheral blood revealed B-cell CLL; prognostic factors: CD38 negative, ZAP70 positive, rearrangement of the immunoglobulins mutated; FISH: negative; CT chest / abdomen / pelvis: presence of multiple aorto-pulmonary and axillary adenopathies (max diameter of 2 centimeters); bone marrow biopsy: infiltration of CLL equal to 60% of global cellularity. The patient was only observed until January 2015, when he was hospitalized due to acute anemia, requiring supportive therapy, and right foot pain . So it was decided to re-evaluate the whole disease in order to decide whether to start chemotherapy. The disease was staged again with instrumental and laboratory tests: presence of renal insufficiency, egd and colonoscopy negative, Coombs' test negative, bone marrow biopsy confirmed the diagnosis of chronic lymphocytic with bone marrow infiltration of 90%, abdomen ultrasound showed only moderate splenomegaly. On February, persistence of right foot pain and appearance of swelling, assessed by the orthopedic as a suspected algic and dystrophic syndrome. So he suggested to perform scintigraphy which revealed: pronounced inflammatory osteometabolic reaction of the right tibia/fibula/ankle third distal which could be referred, in the first evaluation, to algic and dystrophic syndrome. However, a local biopsy was performed: localization of chronic lymphocytic leukemia. On March 2015 a total body TC showed 2 nodular calcifications in the right lung lobe, multiple right paratracheal, barety space, aortopulmonary and axillary adenopathies. Prostate size increased. In order to study carefully the liver and prostate lesions, an ultrasound abdomen was performed that documented only enlarged spleen, normal size liver, free of focal disease, increased prostate due to symmetric bilobate hypertrophy . After the second cycle of chemotherapy, prolonged thrombocytopenia, so he continues only with a radiotherapy program. Disclosures No relevant conflicts of interest to declare.

Absence of Caveloin-1 Leads to Delayed Development of Chronic Lymphocytic Leukemia in Eµ-TCL1 Mouse Model

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2196-2196
Author(s):  
Ashima Shukla ◽  
Christine E Cutucache ◽  
Karan Rai ◽  
Siddharth Rai ◽  
Rene Opavsky ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Lymph Nodes ◽  
Lymphocytic Leukemia ◽  
Clinical Heterogeneity ◽  
Knock Down ◽  
Immune Synapses

Abstract Background: Chronic Lymphocytic Leukemia (CLL) is the most common adult leukemia in the United States. Clinical heterogeneity, a characteristic feature of CLL is a major problem in the clinical management of this currently incurable leukemia. We and others have demonstrated that the tissue microenvironment, specifically the lymph node (LN), influence the biological and clinical behavior including the clinical heterogeneity of CLL. Using gene expression profiling of CLL cells from peripheral blood (PB), bone marrow (BM) and LNs, we identified Cav-1 a member of the Tolerogenic Signature (genes associated with host immune tolerance) as one of the candidate genes which might be involved in the pathogenesis of CLL. We found that Cav-1 levels were significantly elevated (11 fold) in CLL cells from LNs compared to BM and PB. Cav-1 is the major element of caveolae, which are flask-shaped membrane invaginations. Cav-1 is involved in multiple cellular processes like the regulation and transportation of cellular cholesterol and lipids, clathrin independent endocytosis and signal transduction leading to oncogenesis or tumor suppression. We have previously shown that knock down of Cav-1 results in a significant decrease in cell migration and proliferation of primary human CLL cells in vitro. We have also demonstrated that knock down of Cav-1 prevents CLL cells from forming immune synapses. These immune synapses are important for the interaction between the CLL cells and their tumor microenvironment. These results suggest that Cav-1 protect CLL cells from undergoing apoptosis and enhances their migration in vitro. Objectives and Methodology: To understand the precise role of Cav-1 in leukemic progression in vivo, we crossed Cav-1-/- mice to Eµ-TCL1 mice, which is a well-established transgenic murine model for CLL. The offspring were observed and evaluated for the development of CLL. These mice were sacrificed at the age of 12, 24, 36 and 40+ weeks and peripheral blood, bone marrow and spleen and were examined for the presence of CD5+B220+CD19+ CLL cells using flow cytometry. Spleen, lymph nodes, liver, lungs and kidney were evaluated for the presence of CLL cells using H&E staining of histologic slides. Results: To study the role of Cav-1 in Eµ-TCL1, we isolated splenic B cells and measured the expression of Cav-1. We observed a gradual increase in the expression of Cav-1 in splenic B cells from Eµ-TCL1 mice at age of 12, 24 and 36 weeks when compared with wild type mice. This suggested that Cav-1 might be playing a role in CLL progression in Eµ-TCL1 mice. Therefore, to study the role of Cav-1 in CLL disease progression we decreased the expression of Cav-1 in vivo by breeding Eµ-TCL1 with Cav1 knockout mice. We generated Eµ-TCL1-Cav1-/+ and Eµ-TCL1-Cav1-/- mice to study the effect of Cav-1 knock down in aggressiveness of CLL in vivo. We have shown that Cav-1 is overexpressed in CLL cells from patients with poorer clinical outcome and protects CLL cells from undergoing apoptosis. Therefore, we analyze the number of CLL cells in Eµ-TCL1-Cav1-/+ and Eµ-TCL1-Cav1-/- mice. We observed a significant reduction in the number of B220+CD5+ CLL cells population in bone marrow and spleen of Eµ-TCL1-Cav1-/+ and Eµ-TCL1-Cav1-/- mice when compared with Eµ-TCL1-Cav1wt/wt mice. We have previously shown that Cav-1 is important for CLL cells migration in vitro. Therefore, to study its effect in vivo we analyzed infiltration of CLL cells in spleen, lymph nodes, liver, kidney and lungs in these mice. There was no or significant decrease in tumor infiltration of CLL cells in spleen, lymph nodes, liver, lungs and kidney in Eµ-TCL1-Cav1-/+ and Eµ-TCL1-Cav1-/- mice when compared with Eµ-TCL1-Cav1wt/wt alone. Next, we wanted to examine the effect of Cav-1 knock down on splenomegaly and hepatomegaly. We found that there was a significant decrease in splenomegaly and hepatomegaly in Eµ-TCL1-Cav1-/+ and Eµ-TCL1-Cav1-/- mice. The spleen and liver size of Eµ-TCL1-Cav1-/+ and Eµ-TCL1-Cav1-/- mice was significantly reduced when compared with Eµ-TCL1 mice. Together these results suggest that high expression of Cav-1 in CLL cells leads to enhance proliferation and promotes disease progression in Eµ-TCL1 mice. Disclosures No relevant conflicts of interest to declare.

scholarly journals Study of the Quantitative, Functional, Cytogenetic and Immunoregulatory Properties of Bone Marrow Mesenchymal Stem Cells in Patients with B-Cell Chronic Lymphocytic Leukemia

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3861-3861
Author(s):  
Charalampos Pontikoglou ◽  
Maria-Christina Kastrinaki ◽  
Mirjam Klaus ◽  
Christina H. Kalpadakis ◽  
Pavlos Katonis ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Conflicts Of Interest ◽  
Ex Vivo ◽  
Study Groups ◽  
Lymphocytic Leukemia ◽  
Specific Gene ◽  
Proliferative Potential ◽  
Fish Analysis

Abstract Abstract 3861 The microenvironment in both the bone marrow (BM) and lymph nodes has clearly been implicated in the biology of B-chronic lymphocytic leukemia (B-CLL). The non-hematopoietic components of the BM microenvironment originate from a rare population of multipotent progenitor cells, currently referred to as mesenchymal stem/stromal cells (MSCs). The latter have been shown to support hematopoiesis and to affect B cell proliferation and differentiation, thereby setting the stage for studying MSCs' putative role in CLL pathogenesis. However, this particular field of research has not been extensively investigated and the question as to whether patient MSCs differ from their normal counterparts has not been properly answered. The aim of the present study is to explore whether BM-derived MSCs from CLL patients harbor intrinsic abnormalities, which in turn might contribute to the pathophysiology of the disease. BM MSCs were thus isolated from 11 patients with B-CLL (Rai stage 0-III) and 16 age- and sex-matched healthy individuals and their quantitative, functional and cytogenetic characteristics were comparatively assessed. BM MSCs were expanded and re-seeded for a total of 6 passages. Adherent cells from both study groups displayed the same spindle-shape morphology and showed a similar expression of CD29, CD44, CD73, CD90 and CD105 while being negative for CD14, CD34 and CD45. Even though MSC cultures could be established and serially replated from all CLL patients, their growth rate over passages was significantly reduced compared to cultures generated from normal individuals (P < 0.0001). These findings were further substantiated by the MTT assay according to which the number of live cells, at a representative passage (P2), remained significantly lower in CLL patients compared to controls (P <0.0001). The survival characteristics of CLL-derived MSCs at P2 were further evaluated using flow cytometry and 7AAD staining. A statistically significant increase in the proportion of both early and late apoptotic cells was thus documented in B-CLL-derived MSCs as compared to their normal counterparts (P=0.0007 and P=0.0045, respectively). These data suggest that the impaired proliferative potential of CLL-derived MSCs can be attributed, at least in part, to increased cell apoptosis. The frequency of MSCs within the BM mononuclear cell (BMMC) fraction, estimated via the standard CFU-F assay, was significantly reduced in patients, as compared to normal controls (P=0.0039) and this might also be explained by the increased apoptotic rate of CLL-derived MSCs, as well as by BM infiltration by the malignant cells. Regarding MSC differentiation capacity, patient MSCs exhibited normal osteogenic and adipogenic potential as evidenced by cytochemical staining (Von Kossa and Oil Red, respectively) and also by the quantification of osteogenesis (RUNX2, ALP)- and adipogenesis (PPARG, aP2)-related specific gene expression. Furthermore, CLL-derived MSCs did not differ from their normal counterparts in the ability to suppress mitogen-induced T-cell proliferative responses. Since B-CLL cells undergo spontaneous apoptosis in vitro, unless cultured on direct contact with marrow stroma, we evaluated whether ex vivo expanded patient MSCs confer a survival advantage to the malignant clone. Yet, MSCs from both study groups exerted a similar anti-apoptotic effect on CLL cells. CLL-derived MSCs exhibited aberrant production of cytokines that are crucial for B-cell differentiation, survival and apoptosis. More specifically, they secreted significantly lower levels of SDF-1 (P=0.002) and TGF-β1 (P<0.001), as compared to their normal counterparts. Furthermore, CLL-derived MSC supernatants were shown, for the first time, to contain significantly higher levels of APRIL (P=0.0002) and lower levels of BAFF (P=0.0036), as compared to normal MSCs. Finally FISH analysis demonstrated that CLL-derived MSCs did not share any of the cytogenetic abnormalities with CLL cells, thereby suggesting that they are not part of the leukemic clone. In conclusion, ex-vivo expanded B-CLL-derived MSCs harbor intrinsic qualitative and quantitative abnormalities that may be implicated in disease development and/or progression. We anticipate that our observations will contribute to delineating B-CLL biology and will hopefully provide important clues for the design of appropriate microenvironment-targeted therapies. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Elevated FANCA Expression Marks a Worse Prognosis in Chronic Lymphocytic Leukemia. Evidence of a New Role of FANCA in the Stability of p53 By a Neddylation-Related Mechanism

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5541-5541
Author(s):  
Carlos Pipaon ◽  
Sara Bravo Navas ◽  
Iñigo Romon ◽  
Eulogio Conde ◽  
Lucrecia Yanez San Segundo
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Conflicts Of Interest ◽  
Clinical Manifestations ◽  
Lymphocytic Leukemia ◽  
Worse Prognosis

Abstract Background: Chronic lymphocytic leukemia (CLL) is the most common hematological malignancy in western countries. It is characterized by a failure in the mechanisms of apoptosis that leads to an accumulation of mature B cells in peripheral blood, bone marrow and lymphoid organs. With exceptions, CLL is considered incurable and some patients show a worse prognosis related to the expression of certain cytogenetic or molecular markers such as p53 dysfunction (17p13 deletion or TP53 inactivation), 11q23 deletion, unmutated IgVH and the presence of a complex karyotype. FANC proteins have been related to chromosomal instability and alterations in the mechanisms of p53 activation, control of cell cycle and apoptosis. Germline mutations in any of the 20 FANC genes known so far generates Fanconi Anemia, a syndrome characterized by a extraordinary proneness to cell apoptosis leading to a progressive bone marrow aplasia and pancytopenia. Some FANCA proteins aggregate in response to DNA damage forming the FANCcore complex that mediates the monoubiquitination of FANCD2 and FANCI, thus activating the mechanism to repair stalled replication forks. In addition, individual FANC proteins have been involved in functions out of the FANCcore. This is the case of FANCA, that has recently been involved in the neddylation of CXCR5 and beta-2-microglobulin, processes reported to be altered in CLL. Hypothesis: Given the fragmentary information connecting FANC proteins with cellular processes altered in CLL, like apoptosis, cell cycle or neddylation, we hypothesize a role of these proteins in the diagnosis or prognosis of CLL. Methods: We analyzed the expression of 5 FANC genes in a cohort of 160 patients of CLL by quantitative RT-PCR. Statistical analysis were carried out to establish relations between FANC genes deregulation and clinical manifestations. We also investigated the role of the FANCA gene in primary circulating B-lymphocytes from CLL patients by either gain- or loss-of-function approaches. Results: Our data identified a group of CLL patients with high expression of FANCA in peripheral B-CLL cells, and we stablished its relationship with the deletion of 11q23 and a worse prognosis. When we investigated the molecular mechanisms of this bad prognosis, we observed a reduction in the mRNA expression of two p53 target genes, p21 and ∆Np73, in CLL primary cells transfected with FANCA. Luciferase studies demonstrated an impairment of p53 function by FANCA. Moreover, we obtained evidence of a cooperation between FANCA and the NEDD8-interacting protein NUB1L in the destabilization of p53. Conclusion: These results point to FANCA as a bad prognosis marker in CLL and unveils a new role of this protein aside from its role in the FA-BRCA DNA repair pathway. Disclosures No relevant conflicts of interest to declare.

Chronic Myeloid Leukemia Developing in a Patient with B-Cell Chronic Lymphocytic Leukemia – a Case Report

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4421-4421
Author(s):  
Krystyna M Zawilska ◽  
Lucyna Malendowicz-Portala
Keyword(s):  
Bone Marrow ◽  
Chronic Myeloid Leukemia ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Myeloid Leukemia ◽  
Conflicts Of Interest ◽  
Chronic Phase ◽  
Typical Feature ◽  
Lymphocytic Leukemia ◽  
Cell Chronic Lymphocytic Leukemia

Abstract Abstract 4421 The coexistence of B-cell chronic lymphocytic leukemia (CLL) and chronic myeloid leukemia (CML) in the same patient is rare. A 61-year-old man developed a lymphocytosis with morphologic and immunophenotypic feature of B-CLL (stage I according to the modified Rai classification), without indications for treatment. Ten months later he presented with a markedly elevated leukocytes count and splenomegaly. Myeloblasts, promyelocytes, myelocytes and metamyelocytes appeared in his peripheral blood. A bone marrow aspirate was hypercellular with an increased proportion of the myeloid series in all maturative stages; the percentage of lymphocytes was 3%. The immunophenotypic study demonstrated the typical feature of chronic phase of CML, simultaneously 2% of CD5+ CD19+ cells have been found. Unstimulated bone marrow culture shoved a 46,XY,t(9;22)(q34;q11.2) karyotype, and interphase FISH detected the presence of BCR/ABL fusion with 4,55×106 of p 210 and 1,55×103of p 190 copies/ml. Previously it has been shown that these two different hematological malignancies derive from distinct progenitors. Disclosures: No relevant conflicts of interest to declare.

Phase II multicenter study of human CD52 antibody in previously treated chronic lymphocytic leukemia. European Study Group of CAMPATH-1H Treatment in Chronic Lymphocytic Leukemia.

1997 ◽  
Vol 15 (4) ◽  
pp. 1567-1574 ◽  
Author(s):  
A Osterborg ◽  
M J Dyer ◽  
D Bunjes ◽  
G A Pangalis ◽  
Y Bastion ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Phase Ii ◽  
Opportunistic Infections ◽  
Lymphocytic Leukemia ◽  
World Health ◽  
High Dose ◽  
Significant Activity ◽  
Who Grade ◽  
Median Response

PURPOSE CAMPATH-1H is a human immunoglobulin G1 (IgG1) anti-CD52 monoclonal antibody (MAb) that binds to nearly all B- and T-cell lymphomas and leukemias. We report the results of a multicenter phase II trial that used CAMPATH-1H in previously chemotherapy-treated patients with chronic lymphocytic leukemia (CLL). MATERIALS AND METHODS Twenty-nine patients who had relapsed after an initial response (n = 8) or were refractory (n = 21) to chemotherapy were treated with CAMPATH-1H administered as a 30-mg 2-hour intravenous (IV) infusion thrice weekly for a maximum period of 12 weeks. RESULTS Eleven patients (38%) achieved a partial remission (PR) and one (4%) a complete remission (CR) (response rate, 42%; 95% confidence interval [CI], 23% to 61%). Three of eight patients (38%) with a relapse and nine of 21 refractory patients (43%) responded to CAMPATH-1H therapy. CLL cells were rapidly eliminated from blood in 28 of 29 patients (97%). CR in the bone marrow was obtained in 36% and splenomegaly resolved completely in 32%. Lymphadenopathy was normalized in only two patients (7%). The median response duration was 12 months (range, 6 to 25+). World Health Organization (WHO) grade IV neutropenia and thrombocytopenia developed in three (10%) and two patients (7%), respectively. Neutropenia and thrombocytopenia recovered in most responding patients during continued CAMPATH-1H treatment. Lymphopenia (< 0.5 x 10(9)/L) occurred in all patients. Two patients had opportunistic infections and four had bacterial septicemia. CONCLUSION CAMPATH-1H had significant activity in patients with advanced and chemotherapy-resistant CLL. The most pronounced effects were noted in blood, bone marrow, and spleen. Preferential clearance of blood may allow harvesting of uncontaminated blood stem cells for use in high-dose chemotherapy protocols.

scholarly journals Aberrant Expression of TLR2, TLR7, TLR9, Splicing Variants of TLR4 and MYD88 in Chronic Lymphocytic Leukemia Patients

2021 ◽  
Vol 10 (4) ◽  
pp. 867
Author(s):  
Katarzyna Skorka ◽  
Paulina Wlasiuk ◽  
Agnieszka Karczmarczyk ◽  
Krzysztof Giannopoulos
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Cytotoxic T Cells ◽  
Lymphocytic Leukemia ◽  
Aberrant Expression ◽  
Downstream Signaling ◽  
Splicing Variants ◽  
High Level ◽  
Time To First Treatment ◽  
First Time

Functional toll-like receptors (TLRs) could modulate anti-tumor effects by activating inflammatory cytokines and the cytotoxic T-cells response. However, excessive TLR expression could promote tumor progression, since TLR-induced inflammation might stimulate cancer cells expansion into the microenvironment. Myd88 is involved in activation NF-κB through TLRs downstream signaling, hence in the current study we provided, for the first time, a complex characterization of expression of TLR2, TLR4, TLR7, TLR9, and MYD88 as well as their splicing forms in two distinct compartments of the microenvironment of chronic lymphocytic leukemia (CLL): peripheral blood and bone marrow. We found correlations between MYD88 and TLRs expressions in both compartments, indicating their relevant cooperation in CLL. The MYD88 expression was higher in CLL patients compared to healthy volunteers (HVs) (0.1780 vs. 0.128, p < 0.0001). The TLRs expression was aberrant in CLL compared to HVs. Analysis of survival curves revealed a shorter time to first treatment in the group of patients with low level of TLR4(3) expression compared to high level of TLR4(3) expression in bone marrow (13 months vs. 48 months, p = 0.0207). We suggest that TLRs expression is differentially regulated in CLL but is similarly shared between two distinct compartments of the microenvironment.

Sign in / Sign up

Export Citation Format

Share Document