bluuebunny/crossref_metadata_2025_split huggingface dataset:
FMNL-1 (Formin-Like 1) Gene in Chronic Lymphocytic Leukemia (CLL) Is Overexpressed in Young Patients with Adverse Prognostic Factors and Poor Outcome.
Authors: Calpe Eva , Favaro Patricia M.B. , Crespo Marta , Baptista Maria Joao , Muntañola Ana , Giné Eva , Saad Sara , Montserrat Emili , Bosch Francesc 
Published: October 2019

Abstract
Prognosis of patients with CLL has been traditionally assessed by using clinical parameters. Although useful, such parameters are a mere reflection of the biological diversity of CLL. In this regard, the mutational status of VH genes or ZAP-70 expression separates CLL into two clinical forms with different presenting features and outcome. Formins are multidomain proteins characterized by the presence of two conserved prolin-rich regions, namely formin homology 1 and 2. These proteins are implicated in a wide range of processes, including regulation of the cytoskeleton and in the regulation of the signal for cell survival. Formin is normally expressed in spleen, lymph node, and bone marrow cells, and it has been recently found to be overexpressed in T-cell lymphomas. The aim of this study was to analyze the expression of FMNL-1 in normal B-cell subsets and in a series of 73 patients (median age, 59 years; male/female 40/33; Binet A: 90.2%) with CLL. FMNL-1 expression was analyzed by Western Blot in separate subpopulations and by quantitative RT-PCR using expression in Jurkat as baseline. Among normal lymphocytes, FMNL-1 was only expressed in memory (CD19+CD27+) B-cells and in T-cells. In CLL cases with a low percentage of T-cells, mean of FMNL-1 expression was 2.18 AU (SD, 1.01 AU). Using an arbitrary cut-off of 3.2 AU, cases with increased expression of FMNL-1 were associated with a younger age at diagnosis (< 50 yrs), elevated lymphocyte count, high serum β2-microglobulin (β2-m) levels, increased ZAP-70 and CD38 expression, shorter time to progression, and shorter survival as compared to cases with low FMNL-1 expression. No relationship was observed with genetic abnormalities (table). In summary, among B-cell lymphoproliferative disorders, FMNL-1, a gene that regulates cell survival, is found only in CLL and its overexpression correlates with adverse clinical and biological parameters, particularly in young patients.
Variable FMNL-1 normal FMNL-1 increased p value Age < 50 yrs 15% 86% 0.0001 Lymphocyte count > 50.000/ μL 14% 57% 0.018 Increasedβ2-microglobulin 17% 66% 0.038 CD38 > 30% 28% 100% 0.001 ZAP-70≥20% 50% 100% 0.014 Time to progression (median) 5.1 yrs 0.5 yrs 0.003 Survival (median) 16.4 yrs 8.5 yrs 0.009
Prolonged Survival of Chronic Lymphocytic Leukemia Cells Induced by Sphingosine 1-Phosphate Is Mediated by the G Protein-Coupled Receptor S1P1 and Involves Erk/MAP Kinase, but Not Akt Signaling.
Authors: Seitz Gabriele , Yildirim Sedat , Boehmler Andreas , Kanz Lothar , Mohle Robert 
Published: October 2019

Abstract
Sphingosine 1-phosphate (S1P) is a bioactive lysophospholipid mediator which is generated by the hydrolysis of sphingomyelin and released predominantely by activated platelets but also by other cells upon activation. In normal blood plasma it is found in nanomolar to micromolar concentrations. By qualitative and quantitative (TaqMan) RT-PCR, mRNA expression of the S1P receptor S1P1 was detected in CLL cell lines (EHEB, MEC-1) and in all samples of primary chronic lymphocytic leukemia (B-CLL) cells, analyzed after isolation with a CD19 antibody and magnetic cell sorting. mRNA expression of other S1P receptors was less consistently found. When primary CLL cells were cultured in serum-free medium (RPMI 1640 supplemented with fatty acid-free BSA), viability decreased starting from 3 days of culture and was completely lost after 1–2 weeks depending on the individual sample. However, when the cultures were supplemented with S1P (1 nM - 5 μM) the number of viable cells was increased up to 8-fold in a time- and dose-dependent manner as measured using the WST-1 reagent. Moreover, S1P induced a rapid (maximum at 1 min) phosphorylation of the p44/42 Erk/MAP kinase in CLL cell lines and even more prominent in primary CLL cells. The time- and dose dependent phosphorylation was blocked by pretreatment (24h) of the cells with FTY720 (10–100 nM), a compound that binds predominantly to S1P1 and induces loss of cell surface expression by receptor internalization. FTY720 also abrogated the prolonged survival of CLL cells in cultures supplemented with S1P, demonstrating that both the Erk/MAP kinase phosphorylation and the consecutive increased survival were mediated by the S1P1 receptor. Activation of MAP kinase was also inhibited by pertussis toxin, which selectively blocks Gi proteins, the only class of G proteins known to be coupled to S1P1. Phosphorylation of Akt was not observed in CLL cell lines and primary CLL cells after stimulation with S1P. B-CLL cell lines showed a pronounced chemotactic response to S1P at low concentrations. In primary CLL cells however, S1P did neither induce phosphorylation of the focal adhesion kinase (FAK) nor chemotaxis. We conclude that S1P, constitutively present in the blood, prolongs survival of CLL cells involving the Erk/MAP kinase pathway. Since compounds that induce S1P1 receptor internalization like FTY720 are already used as immunosuppressants in humans, our results suggest inclusion of these drugs also in the treatment of B-CLL, where they may potentiate the effect of conventional antiproliferative therapy.
Activation by CD40 Ligand Shifts Bcl-2 Family Protein Expression from Death-by-Default to Survival-by-Default.
Authors: Willimott Shaun , Baou Maria , Huf Sarah , Wagner Simon 
Published: October 2019

Abstract
CLL cells survive and accumulate in vivo and yet leukaemic cells isolated from the peripheral blood rapidly die. We have used mouse fibroblast L-cells transfected with CD40 ligand (CD40L) in combination with IL-4 to reproduce some features of the in vivo microenvironment, which is essential for CLL cell proliferation. As anticipated we found that leukaemic cells survive and proliferate in this system. Also CD23 expression increases, but unexpectedly CD38 expression increased to varying degrees in all patients and CD5 expression decreased. Changes in CD23 and CD38 have been reported in CLL cells in lymph nodes and support the concept that we are able to some extent reproduce the in vivo microenvironment. Next we carried out a systematic analysis of Bcl-2 family proteins by Western Blot analysis. On tissue culture plastic and non-transfected L-cells Bcl-2 is the predominant pro-survival protein. The activator BH3-only protein Bim is expressed together with the sensitiser BH3-only Puma. Both pro-apoptotic Bak and Bax are expressed. By contrast on CD40L-L cell with IL-4 culture Bcl-2 expression decreases and the predominant prosurvival protein is Bcl-xL with a contribution from Mcl-1. The activator BH3-only protein is Bid rather than Bim and the sensitiser BH3 only proteins Puma, Noxa and Bad are expressed. Levels of Bak are significantly decreased but Bax expression is unchanged. Overall it appears that there is an excess of pro-survival Bcl-xL on CD40L-L cells/IL-4 with decreased amounts of Pro-apoptotic Bak. These findings are important when considering the place of treatments directed at pro-survival Bcl-2 family proteins. Large amounts of Bcl-xL is a reservoir to soak up the reduced amount of Bak suggesting that whilst Bcl-2 antagonists will tip the balance towards apoptosis in CLL cells activated by CD40L/IL-4 they are unlikely to be sufficient to cause cell death. However, leukaemic cells in the peripheral blood are dependent on Bcl-2 and have increased amounts of Bak. In this situation reduction in Bcl-2 will release sufficient Bak to cause apoptosis.
B-CLL Antibodies Comprised of Stereotypic VH1-69, D3-16, and JH3 Rearrangements Immunoprecipitate Cellular Protein(s).
Authors: Chu Charles C. , Wang Xiao Bo , Hatzi Katerina , Catera Rosa , Meffre Eric , Herve Maxime , Allen Steven L. , Rai Kanti R. , Chiorazzi Nicholas 
Published: October 2019

Abstract
B-cell chronic lymphocytic leukemia (B-CLL) is typically due to a clonal expansion of B-lymphocytes that can be divided into two classes based on the mutation status of the variable region of the antibody produced by the leukemic clone. Mutated B-CLL (antibody with ≥ 2% mutation) is associated with longer survival than unmutated B-CLL. Segregation of patients based on antibody mutation suggests a functional role for antigen binding to antibody in this disease. Furthermore, many different subsets of B-CLL antibodies have very similar CDR3 sequences, suggesting some common antigen reactivity. Subset I contains rearrangements comprised of an unmutated sequence encoded by VH1-69, D3-16, and JH3 with a nearly identical HCDR3 region. Antibodies from this subset strongly bind cytoplasmic structures in HEp-2 cells by immunofluorescence and are polyreactive against multiple autoantigens (ssDNA, dsDNA, insulin) by ELISA. In contrast, subset VI, which comprises a mutated sequence encoded by VH3-21 and JH6 resulting in a nearly identical HCDR3 that differs from that of subset I, does not bind HEp-2 cells and is not polyreactive. Because the subset I B-CLL antibodies were reactive with HEp-2 cells, we prepared HEp-2 cell extracts in order to isolate the cellular antigen recognized by these antibodies. Antibodies from both subset I (patients CLL068 and 258) and VI (patient CLL412) were recombinantly overexpressed and purified as human IgG1s. Antibody was bound to protein G agarose beads and incubated with cell extracts overnight. Bound cellular antigen was immunoprecipitated, washed, eluted, run on SDS-PAGE, and stained with Coomassie Blue. Both CLL068 and 258 antibodies immunoprecipitated protein bands at about 45 and 200 kDa. These bands have been isolated and are undergoing microsequencing analysis. In contrast, CLL412 did not immunoprecipitate any protein bands under the same conditions. Thus, unmutated subset I CLL antibodies react with common cellular protein(s), whose identity may be useful in understanding B-CLL.
Reduced Rates of B Cell Proliferation in Chronic Lymphocytic Leukemia.
Authors: Defoiche Julien , Debacq Christophe , Asquith Becca , Zhang Yan , Burny Arsène , Bron Dominique , Lagneaux Laurence , Macallan Derek , Willems Luc 
Published: October 2019

Abstract
Whether chronic lymphocytic leukemia (CLL) represents latent or proliferating disease has been intensively debated. Whilst the dogma that CLL results from accumulation of dormant lymphocytes is supported by the unresponsiveness of leukemic cells to antigens and polyclonal activators, recent in vivo kinetic measurements show that B-lymphocytes do divide at significant rates in CLL. However, B cell kinetics were not compared between CLL patients and healthy controls so it was not possible to ascertain to what extent lymphocyte kinetics were aberrant in CLL.
We compared proliferation rates of B- and T-lymphocytes in CLL patients and healthy controls, using a pulse-chase approach based on incorporation of deuterium from 6,6-2H2-glucose into DNA. We found dramatically reduced in vivo rates of CD3−CD19+ cell proliferation in CLL compared with controls (mean 0.47 versus 1.66 %/day respectively, P=0.001), equivalent to an extended half-life of circulating B-cells (147 days versus 42 days). Labeled (dividing) CD3−CD19+ cells had death rates similar to the healthy controls (2.29 versus 3.55 %/day, P=0.495). Despite such aberrant B-cells kinetics, T-cell proliferation was unaffected by CLL (1.77 versus 1.40 %/day, P=0.488). We conclude that, B-cell proliferation rates are reduced in leukemic patients compared to healthy subjects and that most circulating CD3−CD19+ cells are quiescent, long-lived cells.
Distinct Phenotype of Chronic Lymphocytic Leukaemia (B-CLL) Cells Engrafting in NOD/SCID Mice.
Authors: Ebeling Peter , Durig Jan , Grabellus Florian , Seeber Siegfried , Duehrsen Ulrich , Moritz Thomas 
Published: October 2019

Abstract
We have previously reported reliable engraftment of CD19/CD5/CD23/CD45 positive human B-CLL cells in the spleens of NOD/SCID mice (NSS-CLL cells) following transplantation of human peripheral blood-derived B-CLL (PB-CLL) cells. On histology, in the majority of the samples NSS-CLL cells formed focal aggregates which also contained various amounts (range:0–50%) of human CD45/CD3+ T cells (ASH 2005; #52). We now have further characterized these NSS-CLL cells. In our model 1x10^8 MNC are injected i.v. plus i.p. and engraftment in the spleen and other organs is analyzed after 4 (−8) weeks by flow cytometry and immunohistochemistry. In 22 CLL samples investigated in a total of 64 NOD/SCID mice 1.6±0.4x106 NSS-CLL cells representing 10.5±2.3% of total spleen cells were recovered with recovery highly reproducible on repetitive experiments (3.0 vs. 2.0; 1.6 vs. 1.5; 0.2 vs. 1.1; 0.0 vs. 0.0x106 NSS-CLL cells; n=4). In contrast to the low proliferative status reported for human PB-CLL, considerable proliferation was observed in NSS-CLL cells. Following the application of BrdU (1mg/6g d5 i.p., 1mg/ml added to drinking water d6–13; 20–27) BrdU positive NSS-CLL cells ranged from 5.3–25.0% (n=5). Immunohistological analysis showed positive staining for the proliferation marker Ki67 in 18.5±3.5% of NSS-CLL cells. In addition, markers related to proliferation/activation of B-CLL proliferation center cells such as survivin (mean MFI: 5.5±0.8 vs. 2.3±0.2; p=0.028; n=6) and CD100 (semaphorin 4d; mean MFI: 3.3±0.5 vs. 1.1±0.2; p=0.043; n=5) were significantly up-regulated in NSS-CLL vs. PB-CLL cells. When NSS-CLL cells were characterized utilizing additional adhesion/migration, differentiation and/or activation markers, only a subset of NSS-CLL cells (15.7±8.2%, n=6) expressed chemokine coreceptor (CCR) 7, while PB-CLL cells uniformly showed high CCR7 expression. Also CCR5, a migration marker in the context of non-Hodgkin lymphomas, consistently was demonstrated on a fraction of NSS-CLL cells (8.1±3.0%; n=6), whereas here no CCR5 expression was found on PB-CLL cells. No expression of human CD10, CD11c, CD127 or CD49d was detected in B-CLL cells derived from either PB- or NSS-CLL cells, while both sources, as expected, uniformly stained positive for CD40. In summary, NSS-CLL exibit a distinct phenotype as compared to PB-CLL cells and show considerable similarity to B-CLL cells in the proliferation centers of involved human tissues. For these cells not only proliferation but also increased expression of survivin and CD100 as well as down-regulation of CCR7 have been described. Therefore, we think that our model may represent a novel tool to further elucidate biological properties of human B-CLL and, in addition, may serve to devise and evaluate novel treatment strategies.
The Combination of Alemtuzumab [Continuous Intravenous Infusion (civ) Followed by Subcutaneous Injection (sc)] Plus Rituximab Has Activity in Patients (pts) with Relapsed Chronic Lymphocytic Leukemia (CLL).
Authors: Faderl Stefan , Ferrajoli Alessandra , Wierda William G. , O’Brien Susan , Ravandi-Kashani Farhad , Fayad Luis , Thomas Deborah , Kantarjian Hagop , Browning RN Mary L. , Keating MB Michael J. 
Published: October 2019

Abstract
Available data suggest enhanced activity of monoclonal antibodies in combinations. In a previous study of iv bolus alemtuzumab plus rituximab in relapsed CLL, we have shown that the combination is safe and produces a response rate of 54% following a 4-week treatment period (Blood101: 3413, 2003). As questions remain about the optimal dose, schedule and route of administration of alemtuzumab, we have been exploring alemtuzumab by civ followed by sc injections in combination with rituximab in pts with relapsed/refractory CLL who express both CD20 and CD52. Forty-five pts have been enrolled of whom 32 pts are evaluable (30 CLL, 1 CLL/SLL, 1 MZL). Six pts are too early and 7 are inevaluable [withdrawal of consent prior to treatment start (2), ineligible (2), removal from study and continuation of therapy with home physician (3)]. All pts received alemtuzumab 15mg by civ over 24 hours daily x 6 days on days 2–7 of the first course followed by 30mg sc twice weekly on days 3 and 5 of weeks 2 to 4 (one course). Rituximab was given at 375mg/m2 iv on day 1 followed by 500mg/m2 on days 8, 15, and 22. Maximum duration of therapy was 3 courses. Pts received standard premedication with acetaminophen and an antihistamine, and antiinfective prophylaxis with TMP/SMX and valacyclovir. CMV antigenemia was tested before and after completion of each course. The median age was 59 yrs (range 39–78), the median number of prior therapies was 3 (1–8), and the median b2M 3.8 mg/dL (1.7–13.6). Seventeen pts (53%) had Rai stage ≥ 3 disease. Eighteen pts (56%) were refractory to fludarabine and alkylators. All pts had prior rituximab; only 4 (13%) had received alemtuzumab. Sixteen pts (50%) responded [8 CR (25%), 1 PRn (3%), 7 PR (23%)] following one course. Response by site: peripheral blood 24/26 (92%), marrow 22/32 (69%), lymph nodes 13/26 (50%) and liver/spleen 8/9 pts (89). The combination was well tolerated with no unexpected toxicities. Most non-hematologic AEs were infusion-related and ≤ grade 2 by NCI toxicity criteria. Fevers and chills occurred in up to 56% of pts, fatigue in 34%, skin rashes in 19%, nausea in 16%, myalgias in 9%, and diarrhea in 6%. Although toxicites were more frequent with civ alemtuzumab than during the s.c. injections (with the exception of skin rashes), civ alemtuzumab was well tolerated. CMV reactivation occurred in 7 pts (22%) and was the most common infectious complication. In conclusion, the combination of civ/sc alemtuzumab plus rituximab has activity in patients with relapsed and refractory CLL. Most responses occurred after 4 weeks of therapy only. The combination is well tolerated. Side effects are predictable and manageable.
Computed Tomography (CT) Predicts Response Duration in Patients with Chronic Lymphocytic Leukemia (CLL) in Complete Response (CR) by Conventional, NCI-Working Group, Criteria.
Authors: Bosch Francesc , Muntañola Ana , Ferrer Ana , González Marcos , Gine Eva , Abella Eugenia , Gardella Santi , Briones Javier , González-Barca Eva , Escoda Lourdes , Pérez-Ceballos Elena , Asensi Antoni , Sayas Maria José , Besalduch Joan , Montserrat Emili 
Published: October 2019

Abstract
Imaging studies are not routinely performed to evaluate response to therapy in patients with CLL. To ascertain whether CT scans should be part of the maneuvers to assess response in patients with CLL we have investigated response duration according to CT results in 34 previously untreated patients (median age, 56 years; range, 38 to 65 years; males: 79%) uniformly treated with FCM (fludarabine 25 mg/m2 i.v. on days 1 to 3, cyclophosphamide 200 mg/ m2 on days 1 to 3, and mitoxantrone 6 mg/m2 i.v. on day 1, given at a 4-week intervals up to six courses) as part of a phase II study. Out of 69 patients included in the study, 64% achieved CR as defined by the NCI-Working Group criteria. Of those, 30% had abdominal lymphadenopathy as assessed by CT scans. The median follow-up of the series was 30 months. As shown in the figure, response duration was significantly shorter (54% at 20 months) in CR patients with abnormal CT (CT+ CR) than in those with no detectable lymphadenopathy (CT- CR) (95% at 20 months). Interestingly, response duration in patients with CT+ CR was not different from that of patients in PR (54% vs. 42% at 20 months, respectively; p=NS). These results demonstrate that residual abdominal lymphadenopathy negatively influences response duration in patients with CLL in whom by all other criteria a CR has been obtained. Therefore, CT scans should be performed in patients having achieved CR by conventional, NCI-Working Group criteria, and patients with residual disease by CT should be considered in PR rather than in CR.
Figure Figure
Dasatinib (SPRYCEL®) in Patients (pts) with Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia Who Are Imatinib-Resistant (im-r) or -Intolerant (im-i): Updated Results from the CA180-015 ‘START-L’ Study.
Authors: Dombret Herve , Ottmann O.G. , Rosti G. , Simonsson B. , Larson R.A. , Gollerkeri A. , Apanovitch A. , Radich J. , Coutre Steven 
Published: October 2019

Abstract
Dasatinib (SPRYCEL®, formerly BMS-354825) is a novel, oral, multi-targeted kinase inhibitor of BCR-ABL and SRC. Relapsing pts with Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph(+) ALL) who have been previously treated with chemotherapy including im have a very poor prognosis. START-L is an open label, multi-center, global phase-II study, which treated 46 im-r or im-i pts with Ph(+) ALL from January through June 2005. Dasatinib was given orally, 70 mg twice daily (BID) with escalation to 100 mg BID for poor response or reductions to 50 mg and 40 mg BID for toxicity. Response was assessed on weekly blood counts and monthly bone marrow exams, including cytogenetics. Mutation analysis was conducted at baseline and at end of study. Quantitative PCR was carried out pretreatment and at the time of complete cytogenetic response (CCyR). At the time of this update, all 46 pts had a minimum of 9 months (mo) of follow-up. Of these, 44 (96%) were im-r; due to the small number of im-i pts, data for all pts are presented. 59% of patients were male and median age was 48 years. The median time from initial Ph(+) ALL diagnosis to first dose was 18 mo. All patients had been pretreated with im including 46% with 600 mg per day or more, and 52% whose duration of im treatment was more than 12 mo. 37% of the pts had received a stem cell transplant. At baseline, 67% of pts had WBC <2000/mm3, 74% had platelets <100 × 103/mm3, and 24% had extramedullary disease outside of the spleen. In the 40 pts with baseline mutation data, im-resistant BCR-ABL mutations were observed in 78%, one with T315I. The median duration of therapy was 3.0 mo (range 0.03–16.5) for all pts and was 13.0 mo (9.2–16.5) for pts still on study. The overall complete hematologic response rate was 35%. The major cytogenetic response was 57%, including 54% CCyR. The major hematologic response (MHR) in the 31 pts with baseline mutations was 45%. The median duration of MHR was 11 mo and the median progression-free survival was 3.7 mo (95% CI 1.6–6.1). Grades 3 and 4 thrombocytopenia occurred in 13% and 67%, respectively and grades 3 and 4 neutropenia occurred in 27% and 52% of pts, respectively. Most frequent non-hematologic toxicities included diarrhea in 30% (grade 3–4, 7%), nausea in 22% (no grade 3–4), pyrexia in 22% (grade 3–4, 2%), and pleural effusion in 22% (grade 3–4, 7%) of pts. Dose was reduced in 30% and interrupted in 43% of pts, primarily due to non-hematologic toxicities. Dasatinib has important efficacy in this Ph(+) ALL pt population. Updated efficacy (including molecular response), safety, and mutational analysis data will be presented at the meeting.