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Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening hyperinflammatory syndrome that may complicate hematologic malignancies. HLH and malignancies have common clinical features, and HLH diagnostic criteria (HLH-2004/Hscore) were not validated in this specific population. We describe a case of a 72-year-old patient with a history of chronic lymphocytic leukemia stable for over 10 years who presented with fever and cytopenia. After excluding infectious etiologies and the progression of her disease, HLH was diagnosed. The patient was treated with etoposide, dexamethasone, intravenous immunoglobulin, and rituximab. Despite initial clinical improvement, the patient deteriorated and developed pulmonary aspergillosis and CNS involvement that reflected uncontrolled HLH. The patient died 45 days after her presentation. An unusual feature of this case was that HLH was not triggered by infection, disease transformation, or treatment. This case emphasizes the challenges of differentiating the development of overwhelming HLH from other complications associated with hematologic malignancy.

Background Major advances in the treatment of patients with hairy cell leukemia (HCL) have been made following the introduction of purine analogues. The major significant short-term toxicity of cladribine therapy are neutropenia and neutropenic fever (NF) which may be life-threatening. Aim In this retrospective study, we compared the incidence and duration of neutropenia and hospitalization in patients with HCL treated with cladribine followed by peg-filgrastim as primary prophylaxis versus daily filgrastim given “on demand” according to absolute neutrophil count (ANC). Methods Medical records of patients with HCL diagnosed and followed in 12 medical centers in Israel during 1985–2015 were examined for details of disease at diagnosis. The efficacy of peg-filgrastim and filgrastim was assessed by evaluating the incidence of neutropenia (ANC 9 ]/L), number and length of hospitalizations, and number of days from the last day of therapy to recovery of ANC to >1.0 × 10 [ 9 ]/L. Results The study population included 202 patients with HCL, 159 of whom (80.7%) were treated with cladribine; 78 patients (49%) required hospitalization for the administration of broad-spectrum antibiotics due to NF. Twenty-eight (19%) patients were treated with peg-filgrastim as primary prophylaxis, while 74 (64%) received filgrastim “on demand” due to neutropenia. Median length of hospitalization, and nadir duration were 8 and 18 days respectively (p = 0.71, p = 0.44). Conclusions Infectious complications post-cladribine treatment remain high. No difference was found in terms of incidence of NF, number of febrile days, and nadir duration in patients receiving primary peg-filgrastim prophylaxis compared to filgrastim given on demand. Both approaches are justifiable, and the choice remains at the physician’s discretion.

BACKGROUND Chronic lymphocytic leukemia (CLL) is frequently accompanied by immune dysregulation. AIMS In this multicenter prospective study, we investigated whether heavy + light chains (HLC: IgGκ, IgGλ, IgAκ, IgAκ, IgMκ, IgMλ) and IgG subclasses (IgG1, IgG2, IgG3, and IgG4) could be used as novel prognostic markers of immunoparesis in 105 treatment-naive patients with CLL. RESULTS Heavy + light chains immunoparesis of ≥1, ≥2, and ≥3 isotypes was evident in 74 (70%), 58 (55%), and 36 (34%) patients, respectively. Severe HLC immunoparesis was identified in 40 (38%) patients. Of the IgG subclasses, IgG1 and IgG2 were most frequently suppressed, affecting 46 (44%) and 36 (34%) patients, respectively; 63 (60%) patients had low levels of at least one IgG subclass. In multivariate analysis, severe HLC immunoparesis (hazard ratio [HR]: 36.5; P = .010) and ΣFLC ≥ 70 mg/L (HR: 13.2; P = .004) were the only factors independently associated with time to first treatment (TTFT). A risk model including these variables identified patients with 0, 1, and 2 risk factors and significantly different TTFT (P

Chronic lymphocytic leukemia (CLL) is the most common adult leukemia in the Western world. The therapeutic approach to CLL includes chemotherapeutic regimens and immunotherapy. Complement-mediated cytotoxicity, which is one of the mechanisms activated by the therapeutic monoclonal antibodies, depends on the availability and activity of the complement (C) system. The aim was to study the structure of circulating C components and evaluate the importance of C5 structural integrity for C activity in CLL patients. Blood samples were collected from 40 naïve CLL patients and 15 normal controls (NC). The Western blot analysis showed abnormal C5 pattern in some CLL patients, while patterns of C3 and C4 were similar in all subjects. Levels of the C activation markers sC5b-9 and C5a were quantified before and after activation via the classical (CP) and alternative (AP) pathways. In patients with abnormal C5, basal levels of sC5b-9 and C5a were increased while activities of the CP and of the CP C5-convertase, the immediate C5-upstream complex, were decreased compared to NC and to patients with normal C5. The data indicate a link between CP activation and apparent C5 alterations in CLL. This provides a potential prognostic tool that may personalize therapy by identifying a sub-group of CLL patients who display an abnormal C5 pattern, high basal levels of sC5b-9 and C5a, and impaired CP activity, and are likely to be less responsive to immunotherapy due to compromised CP activity.

The immunotherapy treatments offered for Chronic lymphocytic leukemia (CLL) activate various cellular and biochemical mechanisms, including the complement system. Recently it was shown that the classical complement pathway (CP) in CLL patients is persistently activated at a low level through (Michelis et al. PlosOne, 2019), and that the mechanism of chronic CP activation involves the formation of IgG-hexamers (Michelis et al. PlosOne, 2020). According to recent studies, formation of ordered IgG-hexamers occurs on cell surfaces via specific interactions between Fc regions of the IgG monomers, which occur only after antigen binding. The study investigated the formation of IgG-hexamers in CLL patients and normal controls (NC), their ability to activate complement, their incidence as cell-free and cell-bound forms and the identity of the antigen causing their formation. Sera from 30 patients and NC were used for separation of IgG-hexamers. The obtained IgG-hexamers were measure and used for assessment of CP activity. For evaluation of the presence of IgG-hexamers on blood cells, whole blood samples were stained and assessed by flow cytometry. Serum levels of IgG-hexamers were higher in patients compared to NC. Also, they activated the complement system to a higher extent in patients than in NC. Alpha 2 macroglobulin (A2M) was recognized as the antigen causing the hexamerization of IgG, and was found to be part of the hexamer structure by mass spectrometry, Western blot and flow cytometry analysis. The presence of A2M-IgG-hexamers on B-cells suggests that it may be formed on B cells surface and then be detached to become cell-free. Alternatively, it may form in the plasma and then attach to the surface of the cells. The exact course of A2M-IgG-hexamers formation in CLL should be further studied. The results in this study may be useful for improvement of current immunotherapy regimens. Disclosures Tadmor: Neopharm: Consultancy, Speakers Bureau; AbbVie: Consultancy, Speakers Bureau; Janssen: Consultancy, Speakers Bureau; Takeda: Consultancy, Speakers Bureau; Sanofi: Consultancy, Speakers Bureau; Medison: Consultancy, Speakers Bureau; 6. Novartis Israel Ltd., a company wholly owned by Novartis Pharma AG: Consultancy, Speakers Bureau. Aviv:Abbvie: Honoraria.

Background/aim In this retrospective study, we summarized the national Israeli experience with hairy cell leukemia (HCL) in a large cohort of patients with a long follow-up. Patients and methods Demographic data, and relevant laboratory and clinical parameters were analyzed, emphasizing the outcome after first-line treatment with cladribine. Results Data on 203 patients was collected from 12 medical centers during 1985-2015. Mean and median follow-up were 7.5 years and 5.18 years (interquartile range=0.1-40 years), and 5- and 10-year survival were 96% and 90.62%, respectively. The median age of diagnosis was 55.5 years for Jews and 49 years for Arabs (p=0.021), and most patients were males (81.77%); 52.2% were Ashkenazi Jews, 36.1% Sephardic Jews and 11.7% were Arab, Druze or other ethnicity. Cladribine was given to 159 patients (80.7%%) and most (62%) received intravenous (i.v.) and 38% received subcutaneous (s.c.) therapy. Overall survival and time to next treatment were not significantly different between the two schedules (i.v., s.c.). In univariate analysis of a variety of factors, only age >65 years had a negative impact on outcome, with shorter overall survival. It is of interest that Arab patients with HCL were diagnosed at an earlier age, but had a similar clinical course and outcome to both Ashkenazi and Sephardic Jews.

Chronic lymphocytic leukemia (CLL) is a disease of elderly patients. The fludarabine, cyclophosphamide, and rituximab (FCR) regimen is considered the treatment of choice for young fit patients with CLL; however, this combination is toxic for older patients. At the time this study was first planned and initiated, there was no standard chemo-immunotherapy regimen regarded as standard therapy for the less fit elderly patient with CLL. Here, we conducted a single-arm, phase II trial to examine the efficacy and safety of lower-dose fludarabine and cyclophosphamide combined with a standard dose of rituximab (LD-FCR) in elderly patients with previously untreated CLL. Forty patients received LD-FCR and were included in the efficacy analysis. Two patients treated with FC alone were only included in the safety analysis. The median age was 72.7 years (range, 65.0 to 85.0). The overall response and complete response rates were 67.5% and 42.5%, respectively. Median progression-free survival (PFS) was 35.5 months (95% CI, 29.27-41.67). Two patients (4.8%) died during the study period. Hematological toxicities and infections were the most common complications encountered; grade 3 to 4 treatment-related neutropenia occurred in 20 (47.6%) patients. During the entire study follow-up, 26 patients (61.9%) had all grades of infection including six (14.3%) with neutropenic fever and eight (19%) with grade 3 to 4 non-neutropenic infections. In conclusion, LD-FCR is an effective and relatively safe regimen for previously untreated patients with CLL. It has the advantage of being both "time and cost limited" and, even in the era of novel agents, can still be considered when planning treatment for elderly patients without high-risk biomarkers. However, recent results in fit elderly patients using the combination of bendamustine and rituximab which have achieved longer PFS with good safety profile must be taken into consideration in this regard.

Richter's syndrome (RS) is the rare development of an aggressive lymphoid malignancy in a patient with pre-existing chronic lymphocytic leukemia (CLL). Data on RS is sparse and mostly derived from case reports or small series of patients and only a few larger cohorts have been published. The purpose of this large retrospective study was to summarize our national experience with RS in CLL, examine possible risk factors, and analyze relevant demographic, laboratory and clinical parameters, including results of therapy and outcome. We first evaluated data obtained from 119 patients with RS diagnosed during 1971–2010 from 12 medical centers in Israel. The final cohort summarized consisted of 81 patients with RS who developed only diffuse large B-cell lymphoma (DLBCL) after exclusion all cases with insufficient data and those who were not DLBCL. Median overall survival from time of diagnosis of RS was 8 months; after applying the Richter score, patients could be stratified into three prognostic groups, while all other clinical and laboratory parameters evaluated had no prognostic significance. Prior therapy for CLL had no impact on RS survival (P = 0.8) and patients with therapy “naive” RS and those who had already received chemotherapy prior to developing RS, had the same survival. The addition of rituximab to chemotherapy for RS improved 2 years overall survival from 19% in the chemotherapy alone arm to 42% (P value of 0.001). Although prognosis of patients with RS remains dismal, this retrospective observation provides support for the use of chemo-immunotherapy in DLBCL-RS. Am. J. Hematol. 89:E218–E222, 2014. © 2014 Wiley Periodicals, Inc.

BACKGROUND: Bone marrow (BM) biopsies from patients with chronic lymphocytic leukemia (CLL) may show reticulin fibrosis at diagnosis, but its significance remains unclear. This study sought to assess the prognostic impact of BM reticulin fibrosis in patients with previously untreated CLL. METHODS: Data was reviewed from untreated CLL patients in the national Israel CLL database, followed during 1987 to 2012. All bone marrow biopsies were graded for reticulin fibrosis using a modified scoring system containing 4 grades (0-3), based on the European consensus report. Grade of reticulin fibrosis was correlated with overall survival (OS), outcome, and a number of well-recognized prognostic factors for CLL. RESULTS: The final cohort included 176 patients (122 males and 51 females). Median age was 63 years (range, 32-86 years) and the 5-year OS was 77.1%. Grade of BM reticulin fibrosis correlated with OS (P < .0001) and mortality (P = .001), and separated patients into 2 groups with different survival curves. Advanced reticulin fibrosis (grades 2-3) was associated with thrombocytopenia (platelet counts of < 100,000/mm3) (P = .025), anemia (P = .018), elevated β2-microglobulin < 4000 μg/mL (P = .048), and the presence of 11q deletion (P = .0015). CONCLUSIONS: There was a significant correlation between poor survival and grade of BM reticulin fibrosis. This staining procedure is easy to perform and can readily be added routinely when examining BM biopsies in CLL, because the findings do have prognostic implications. Cancer 2013. © 2013 American Cancer Society.

Chronic lymphocytic leukemia (CLL) is a disease of elderly patients. Despite the development of novel agents and new monoclonal antibodies, FCR still remains the combination chemoimmunotherapy of choice for fit patients with CLL, yielding the longest durations of remission. When this study was first started, no established chemo-immunotherapy regimen was unanimously regarded as standard therapy for less fit elderly patients with CLL; this category of patients had clearly been underrepresented in clinical trials utilizing chemo - or chemo-immunotherapy. Patients and Methods We conducted a single arm, phase II trial to assess the efficacy and toxicity of low dose fludarabine and cyclophosphamide in combination with a regular dose of rituximab (FCR-LITE) in elderly patients with therapy naïve CLL. Our intention was to deliver 6 courses of Fludarabine which was given intravenously (IV) at 12.5 mg/m2/day together with IV cyclophosphamide 150 mg/m2/day for 3 consecutive days. IV rituximab was administered on day 0 of cycle 1 at a dose of 375 mg/m2, and at 500 mg/m2 on day 1 of cycles 2-6. Categorical variables were compared in patients with and without CR using chi-square test or Fisher's exact test and continuous variables were also compared using Mann Whitney test. Duration of follow-up was recorded using reverse censoring method. Kaplan Meier curve was used to establish PFS during clinical follow-up. All statistical tests were two sided. P Results Forty patients treated with FCR-LITE were included in the efficacy analysis. The median age of the entire cohort was 72.7 years (range, 65.0 to 85.0), and 69% were male. The mean number of treatment cycles was 5.1 (range 1-6). The overall response rate was 67.5% (95% CI, 50.9%-81.4%); 17 patients (42.5%) achieved CR and 10 (25.0%) PR. Median PFS was 35.5 months (95% CI, 29.27-41.67). Two patients (4.8%) died during the study period. Reduced cumulative doses of FCR and fewer courses of treatment were both associated with lower CR rate. Hematological toxicities were the most common side effects encountered; grade-3/4 neutropenia occurred in twenty (47.6%) patients, only six (14.3%) developed neutropenic fever. Positive direct antiglobulin test (DAT), was seen in 11 patients but none of them developed autoimmune hemolytic anemia (AIHA) during treatment; two patients (4.8%) progressed to Richter's transformation and two (4.8%) had second malignancies (lung and metastatic colon carcinoma). Conclusion FCR-LITE is effective and safe for treating elderly patients with therapy-naïve CLL. It has the advantage of being both time and cost effective. In an era of novel agents, it can still be considered as suitable frontline treatment for fit elderly patients with CLL. This research was supported by roche pharmaceuticals Table. Table. Disclosures Tadmor: ABBVIE: Consultancy; ROCHE: Research Funding; NOVARTIS: Consultancy; JNJ: Consultancy; PFIEZER: Consultancy. Herishanu:JNJ: Consultancy; ABBVIE: Consultancy; ROCHE: Research Funding. Bairey:ROCHE: Research Funding; AbbVie: Consultancy; Jansen: Research Funding. Aviv:ABBVIE: Consultancy; ROCHE: Research Funding. Rahimi-Levene:ABBVIE: Consultancy. Fineman:ABBVIE: Consultancy; JNJ: Consultancy. Ruchlemer:JNJ: Consultancy; ABBVIE: Consultancy. Shvidel:JNJ: Consultancy; ROCHE: Consultancy, Research Funding; ABBVIE: Consultancy, Research Funding. Polliack:ABBVIE: Consultancy; ROCHE: Research Funding.

Introduction: Chronic Lymphocytic Leukemia (CLL) is frequently accompanied by immune dysregulation. Hypogammaglobulinemia is one of the most important immune defects encountered, and all three classes of immunoglobulins (IgG, A and M) can be involved. Recently, novel heavy+light chain (HLC) immunoassays have become available that quantify the light chain types of each immunoglobulin class (e.g. IgGk and IgGl). These assays are measured in pairs and provide information on the isotype produced by a tumour (the "involved" HLC), the non-clonal ("uninvolved") HLC, and the ratio (e.g. IgGk/IgGl) - which indicates monoclonality. HLC assays have been shown to improve monitoring of plasma cell dyscrasias, but their role in CLL is yet to be studied. Methods This is a multi-center study performed in collaboration with the Israeli CLL Study Group and involved 10 medical centers. The cohort included 122 patients with CLL and 26 healthy controls. Baseline was defined as the time the blood sample was taken. Serum samples were analyzed for levels of IgG subclasses (IgG1, IgG2, IgG3, IgG4), heavy+light chains (HLC) (IgGκ, IgGλ, IgAκ, IgAλ, IgMκ or IgMλ) and free light chains (sFLC). HLC-pair suppression was defined as an abnormal HLC ratio and uninvolved HLC levels below the normal reference range (i.e. in g/L: IgGκ HLC immunoparesis was defined as HLC isotype levels below the normal reference range, regardless of HLC ratio (i.e. HLC immunoparesis of at least 1 isotype indicates at least one of IgGκ, IgGλ, IgAκ, IgAλ, IgMκ or IgMλ below the normal reference range). Severe HLC-pair suppression or severe HLC immunoparesis was defined as a concentration of the uninvolved HLC or any HLC isotype that is suppressed by >50% below the normal reference range. The association of variables with time to first treatment (TTT) was performed with Cox proportional hazard model. Results Current analysis was performed on 105 CLL patients who had complete data available; median age was 68 years, 64% were males and Binet stage A, B, C, were 80%, 18.1% and 1.9% respectively. Median time from diagnosis to baseline measurements was 27 months (range 0-328 months). An abnormal sFLC ratio was identified in 70% (73/105) patients and summated k and λ concentrations (SFLC) were ≥70 mg/L in 18% of cases. An abnormal HLC ratio was present in 32% (34/105) patients, and 21% had HLC pair suppression of any 1 HLC isotype. HLC immunoparesis of 1, ≥2 and ≥3 isotypes was observed in 74 (70%), 58 (55%) and 36 (34%) of patients, respectively, with severe HLC immunoparesis identified in 40 patients (38%). Patients with IgG2 suppression were more frequently hospitalized due to infection with an Odds Ratio of 3.826 (p=0.031). In multivariate analysis, SFLC ≥70 mg/L and severe HLC immunoparesis were independently associated with TTT (HR 15.3, p Conclusions The findings presented here demonstrate that there is considerable potential for the use of HLC and FLC immunoassays to provide prognostic information in CLL. Figure 1. Figure 1. Disclosures Tadmor: PFIEZER: Consultancy; JNJ: Consultancy; ABBVIE: Consultancy; NOVARTIS: Consultancy; ROCHE: Research Funding. Aviv:ABBVIE: Consultancy; ROCHE: Research Funding. Herishanu:ROCHE: Research Funding; JNJ: Consultancy; ABBVIE: Consultancy. Shvidel:ROCHE: Consultancy, Research Funding; ABBVIE: Consultancy, Research Funding; JNJ: Consultancy. Rahimi-Levene:ABBVIE: Consultancy. Ruchlemer:ABBVIE: Consultancy; JNJ: Consultancy. Fogl:The Binding Site Group Ltd: Employment. Polliack:ROCHE: Research Funding; ABBVIE: Consultancy. Magal:The Binding Site: Employment. Townsend:The Binding Site Group Ltd: Employment.

Objective: Clonal B-lymphocytes of chronic lymphocytic leukemia (B-CLL) are characterized by decreased sensitivity to programmed cell death and, therefore, they accumulate in vivo. However, these malignant cells die rapidly in vitro. In the current study we concentrated on the contribution of autologous serum (AS) and lymphocyte subsets to the survival of the malignant cells in vitro. Methods: Mononuclear cells from the peripheral blood of 26 CLL patients and 24 controls were incubated overnight in the presence or absence of AS and heat-inactivated AS (HI-AS) or fetal calf serum (FCS). Also, isolated B cells were incubated at different concentrations in the presence of AS and/or isolated T cells. The level of apoptosis of CD19+ cells was measured by flow cytometry. Results: Spontaneous apoptosis of unfractionated B-CLL cells incubated with AS, FCS or without serum was significantly lower than the rate of B-cell death in the control group, in similar culture conditions. AS had an antiapoptotic effect on unfractionated B-CLL cells when compared with FCS. The rate of apoptosis of B-CLL cells was directly associated with stage. HI of AS had a variable effect, which was related to the stage of the disease. High concentrations of B cells and the addition of autologous T cells reduced the rate of apoptosis when incubated without serum. The antiapoptotic effect of T cells was most prominent in progressive stages. Conclusions: B-CLL cells exhibit decreased spontaneous apoptosis, which is partially prevented by humoral (AS) and cellular (T cells and B-CLL cells) factors. The equilibrium between apoptotic and antiapoptotic factors changes with disease progression.

Richter's syndrome (RS) is the rare development of an aggressive lymphoid malignancy in a patient with pre-existing chronic lymphocytic leukemia (CLL). Data on RS is sparse and mostly derived from case reports or small series of patients and only a few larger cohorts have been published. The purpose of this large retrospective study was to summarize our national experience with RS in CLL, examine possible risk factors, and analyze relevant demographic, laboratory and clinical parameters, including results of therapy and outcome. We first evaluated data obtained from 119 patients with RS diagnosed during 1971-2010 from 12 medical centers in Israel. The final cohort summarized consisted of 81 patients with RS who developed only diffuse large B-cell lymphoma (DLBCL) after exclusion all cases with insufficient data and those who were not DLBCL. Median overall survival from time of diagnosis of RS was 8 months; after applying the Richter score, patients could be stratified into three prognostic groups, while all other clinical and laboratory parameters evaluated had no prognostic significance. Prior therapy for CLL had no impact on RS survival (P = 0.8) and patients with therapy "naïve" RS and those who had already received chemotherapy prior to developing RS, had the same survival. The addition of rituximab to chemotherapy for RS improved 2 years overall survival from 19% in the chemotherapy alone arm to 42% (P value of 0.001). Although prognosis of patients with RS remains dismal, this retrospective observation provides support for the use of chemo-immunotherapy in DLBCL-RS.

Richter syndrome (RS) is the development of an aggressive lymphoid malignancy, in chronic lymphocytic leukemia (CLL). Most are diffuse large B-cell lymphomas (DLBCL), however in 10-15% of RS, there is transformation to Hodgkin lymphoma, termed "Hodgkin variant" (HV). In the present retrospective study we summarize the Israeli experience with HV-RS, and analyze demographic data, relevant laboratory and clinical parameters, and outcome.We collected and analyzed data from 119 patients with RS from 12 Centers in Israel during 1996-2010 and identified 16 cases with "Hodgkin's variant".The median age was 58 years, and 67% were males. The median time from CLL diagnosis to development of HV was 5.9 (range=0.8-11.9) years and the median survival was 39.5 months, compared to 9 months for the cases with RS-DLBCL.Hodgkin variant appears to differ from DLBCL-RS and is clinically less aggressive. However, compared to de novo Hodgkin's lymphoma, HV-RS has a worse prognosis.

An insect-bite-like reaction is known to occur in patients with chronic lymphocytic leukemia (CLL). Most of the literature, however, consists of isolated case reports or small case series. The aim of this retrospective study was to review the national experience with insect-bite-like reaction in a large group of patients with CLL. The study cohort of patients with these skin reactions consisted of 48 patients (25 males, 23 females) of mean age 64.8 yr (range 33-89) at skin eruption. Data on clinical, histologic, immunophenotypic, and cytogenetic characteristics, treatment, and outcome were collected from the medical files. Mean time between diagnosis of CLL and appearance of the skin lesions was 3.1 yr (range -4 to 14 yr). The eruption was not related to disease activity or the course of the hematological disease. The eruption preceded the diagnosis of CLL in 10 patients (by 0-4 yr); and followed the diagnosis in 36; in 11 patients, it occurred during therapy for CLL and in nine after therapy. Mean duration of the skin findings was 21.5 months (range 0.3-132). The eruption usually presented in summer, although it occurred also at other times of the year, and predominantly affected the upper and lower limbs, although it also appeared on unexposed areas. Treatment included local ointments, antihistaminics, oral steroids, antibiotics, phototherapy, and dapsone with varying responses. Insect-bite-like reactions is a relatively common and disturbing skin reaction in CLL patients, it may be related to the immune dysregulation accompanying CLL and further exacerbated by external factors, including actual insect bites, chemoimmunotherapy, and pyogenic infection.

Chronic lymphocytic leukemia (CLL) is the most common leukemia in the western world. Despite several advances in therapeutic options, the disease remains incurable. Recently, it was repeatedly demonstrated that statins, competitive inhibitors of 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase, have antineoplastic effects. Therefore we aimed to study the effects of simvastatin (Sim) on malignant B cells derived from patients with CLL and mechanisms of action of the drug.Purified B-CLL cells from 15 patients were cultured either alone or with Sim at concentrations of 10, 50, and 100 microM. Viability, measured by the activity of mitochondrial dehydrogenases, was reduced significantly in the cells treated with Sim at 50 and 100 microM for 24 hours (p0.005). The level of apoptosis, as measured by annexin binding to exposed phosphatidylserine moieties, increased significantly in the treated cells at concentrations higher than 50 microM for 24 hours (p0.003). The level of necrosis, as measured by propidium iodide internalization, increased significantly after 24 hours exposure to Sim at 50 microM (p0.01). The apoptotic cascade was studied by immunoblot analysis of caspases following Sim treatment. These showed cleavage of caspases 9, 8, and 3. Addition of the caspase inhibitor Z-VAD.fmk inhibited caspase 8 and 3 significantly but did not affect caspase 9.Exposure of clonal B lymphocytes from patients with CLL to simvastatin decreases viability significantly by the induction of apoptosis. The apoptosis induced by Sim is probably initiated by the mitochondrial caspase 9, which indirectly leads to activation of caspase 3 and 8.

Clonal B-lymphocytes of chronic lymphocytic leukemia (B-CLL) are characterized by decreased sensitivity to programmed cell death and, therefore, they accumulate in vivo. However, these malignant cells die rapidly in vitro. In the current study we concentrated on the contribution of autologous serum (AS) and lymphocyte subsets to the survival of the malignant cells in vitro.Mononuclear cells from the peripheral blood of 26 CLL patients and 24 controls were incubated overnight in the presence or absence of AS and heat-inactivated AS (HI-AS) or fetal calf serum (FCS). Also, isolated B cells were incubated at different concentrations in the presence of AS and/or isolated T cells. The level of apoptosis of CD19+ cells was measured by flow cytometry.Spontaneous apoptosis of unfractionated B-CLL cells incubated with AS, FCS or without serum was significantly lower than the rate of B-cell death in the control group, in similar culture conditions. AS had an antiapoptotic effect on unfractionated B-CLL cells when compared with FCS. The rate of apoptosis of B-CLL cells was directly associated with stage. HI of AS had a variable effect, which was related to the stage of the disease. High concentrations of B cells and the addition of autologous T cells reduced the rate of apoptosis when incubated without serum. The antiapoptotic effect of T cells was most prominent in progressive stages.B-CLL cells exhibit decreased spontaneous apoptosis, which is partially prevented by humoral (AS) and cellular (T cells and B-CLL cells) factors. The equilibrium between apoptotic and antiapoptotic factors changes with disease progression.

Background: The Israel National Cancer Registry (INCR) is a population based, national passive tumor registry established in 1960. Reporting by hospitals, laboratories, and other providers has been mandatory in Israel since 1982 and most cancer cases are registered on the basis of pathology reports and hospital records. In some hematopoietic malignancies where the diagnosis was not based on tissue pathology and patients initially received no inpatient treatment, cases may not have been reported to the registry, or reporting to the INCR may have been delayed, resulting in an underestimation of the true burden of disease. One diagnosis for which there is particular concern in this regard is CLL/SLL and here we used active surveillance to estimate the true incidence of CLL/SLL in Israel. Here we present the interim results Methods: We attempted to estimate the incidence of CLL in Israel more accurately,recognizing the fact that the exact incidence may never be known. The Israel Chronic Lymphocytic Leukemia Study Group, working with the Israel Center for Disease Control of the Ministry of Health, actively documented new cases of CLL/SLL in Israel for calendar years 2011 and 2012. All flow cytometry laboratories in Israel provided lists of patients with B cell clones. Israeli hematologists diagnosing CLL were asked to verify which of the B cell clones indicated a diagnosis of CLL, SLL, PLL or MBL and to fill out an internet-based reporting form. Diagnoses based on flow cytometry were verified by medical record review. Cases identified through active surveillance were pooled with cases known to INCR in order to estimate the true annual incidence of CLL/SLL and assess the completeness of the INCR data. Results: We identified 432 and 396 CLL/SLL cases for 2011 and 2012, respectively of whom 57.4% were males. The average age was 68.8. The corresponding age-adjusted[1]incidence rates per 100,000 (ASR) were 4.26 for 2011 and 3.79 for 2012. In comparison, the INCR registered 295 new CLL cases in 2011 (ASR=2.78) and 232 in 2012 (ASR=2.19), 54.5% of them males. The average age at diagnosis was 69.9. These data indicate a gap between true and reported incidence (1.48 and 1.60/100,000 in 2011 and 2012, respectively). However, it should be noted that the INCR will be fully updated for 2012 only by the beginning of 2015. Of active surveillance cases, 157 (2011) and 152 (2012) were registered in the INCR. Most cases missing in the INCR were diagnosed based on flow cytometry, peripheral blood samples and FISH (85.9% in 2011, 89.8% in 2012) without histo-pathological confirmation. Of the CLL/SLL cases existing in the INCR dataset for 2011-12 but not detected by active surveillance (138 in 2011; 80 in 2012), most (76.8% in 2011, 63.8% in 2012) had been diagnosed earlier and the remainder were coded as diagnoses other than CLL/SLL. Omitting these cases from the INCR dataset substantially increased the observed gaps in the true and the registered annual incidence of CLL/SLL (from 1.48 to 2.80 per 100,000 in 2011 and from 1.60 to 2.37 per 100,000 in 2012). Conclusions: The true incidence of CLL is unknown, but it is clear that there is under reporting to cancer registries. Completeness of CLL/SLL data requires accurate reporting of cases by hematologists and other care providers in the community. In Israel, this issue has been addressed by publishing updated guidelines for mandatory reporting stressing the requirement for reporting of hematologic malignancies. [1] Age-adjustment made on the basis of the world standard population Disclosures Ruchlemer: Roche: Research Funding.

Apoptosis, programmed cell death, occurs in a variety of cellular systems and in response to many different stimuli. In the present study we examined the ability of dexamethasone (Dex) and chlorodeoxyadenosine (2-CdA) to induce apoptosis in lymphocytes of patients with B-chronic lymphocytic leukemia (B-CLL). Lymphocytes of 29 untreated patients and 9 healthy controls were isolated and incubated for 24 hours in the presence or absence of either Dex (2 microM) (n = 15) or 2-CdA (3 microM) (n = 14). Following incubation the cells were harvested and their DNA extracted and analysed for internucleosomal DNA cleavage by UV illumination after electrophoresis on agarose slab gel containing ethidium bromide. In the Dex group, 10 patients showed dexamethasone independent spontaneous apoptosis appearing 24 hours after the start of incubation. These were the only instances of dexamethasone-enhanced apoptosis. Five patients showed no spontaneous or dexamethasone induced apopto sis. Of the 2-CdA group, 5 showed spontaneous apoptosis enhanced by 2-CdA. No spontaneous apoptosis was observed in the cells from 9 other patients, however, 2-CdA induced apoptosis in 8 cases in this group. This study shows that monitoring of apoptosis in CLL may provide important information regarding susceptibility of the cells to drug induced apoptosis.

Introduction Richter's syndrome (RS) is the rare development of an aggressive lymphoid malignancy in a patient with pre-existing, or concomitant chronic lymphocytic leukemia (CLL). This complication occurs in about 5 % of patients with CLL, and the most frequent form is the development of diffuse large B-cell lymphoma (DLBCL) and less frequently Hodgkin lymphoma (HL) or prolymphocytic leukemia (PLL). Most of the available data on RS is derived from case reports or small series of patients, and only a few larger cohorts have been published. Aim The purpose of this retrospective study was to summarize our experience with RS in CLL, examine possible risk factors and analyze relevant demographic, laboratory and clinical parameters, including outcome. Methods We collected a total of 119 patients diagnosed with RS from 12 medical centers in Israel during the period 1971-2010.We then summarized clinical, demographic and some biological features related to CLL at diagnosis and examined possible risk factors for their transformation to RS. Results Of the 119 patients with RS, 61 % were males, 82% developed DLBCL, 14% HL and 4% PLL. In terms of ethnicity: 95% were Jews (64% Ashkenazi) and only 5 % were Arabs, which is similar to the reported data for CLL in Israel. The median time from CLL diagnosis to development of RS was 60 months (range, 0-182 months), and the median overall survival from diagnosis of RS was 9.5 months;34 % of the cases developed extranodal RS, and the most frequent sites of involvement were the gastro-intestinal tract and bone marrow. These results further confirm that there are no established “sanctuary sites” of extranodal RS, which can develop in all tissues and organs. None of the conventional clinical and laboratory parameters examined and evaluated as possible risk factors in CLL were able to predict transformation to RS, including: occurrence of autoimmune phenomenon during the course of CLL, spleen size, serum beta 2 microglobulin levels, degree of CD38 positivity, and the number of previous treatments given for CLL, or the prior use of rituximab in these regimens. The only parameters, present at the time of RS diagnosis which were found to correlate with adverse prognosis, included: performance status>2 (HR- 3.45), high IPI (HR- 3.28) and high Richter score (HR-7.45). In regard to therapy for RS, patients treated with chemotherapy followed by autologous stem cell transplantation (ASCT) had a better outcome with improved overall survival (p=0.034) (Figure). Conclusions RS remains a heterogeneous entity. In this large series of patients none of the conventional clinical, epidemiological or laboratory parameters, often evaluated as possible prognostic factors in CLL, were associated with the risk of transformation to RS. In this retrospective study molecular genetic data were not available to examine their possible significance, as reported recently in other series. In terms of therapy, prior treatment of CLL with rituximab – containing regimens did not appear to impact the eventual outcome of patients who developed RS. On the other hand, ASCT did significantly improve overall survival in patients who had transformation to RS. On behalf the Israeli CLL study Group. Disclosures: No relevant conflicts of interest to declare.

Rituximab has revolutionized the treatment outcome of non-Hodgkins lymphoma (NHL) patients but its role in retreatment in all NHL patients is not yet established. Therefore, the Israel Rituximab Retreatment Study Group was organized in order to summarize retrospectively the treatment results of B-cell NHL patients who received ≥ 2 rituximab treatments. Eighty-eight NHL patients from 13 medical centers were enrolled, 39 males, 49 females with a mean age of 57.3 years (range 26–88). Sixty-four patients (73%) had indolent lymphoma (21- follicular grade I, 17 follicular grade II, 10-follicular grade III, 9-small lymphocytic, 6- MALT, 2- marginal zone, 1-lymphoplasmacytic). Eighteen patients (20%) had aggressive lymphoma [diffuse large B-cell lymphoma (DLBCL)] and 6 patients had mantle cell lymphoma. Fifty-nine patients received one regimen of chemotherapy and 23 patients received 2 chemotherapy regimens before the first treatment with rituximab. There was no significant difference in the median time to progression (TTP) after the first rituximab treatment whether the patient received no previous chemotherapy or 1 or 2 previous chemotherapy regimens (12 vs 12 vs 14 months, respectively). All patients received 2 courses of rituximab, 33 patients-3 courses, 6 patients -4 courses and 2 patients-5 courses, with a mean of 4.4 doses in each course (range 1–8). Only 45 patients received any treatment after the second course of rituximab. The first course of rituximab was administered alone in 46 patients and with chemotherapy in 42 (24 CHOP, 6 COP, 6 FC, 2 chlorambucil, 1 LMP, 1 DVIP, 1 ESHAP, 1 MACOP-B). There was no difference in the median time to next treatment (TTNT) whether the rituximab was given alone or with chemotherapy (16 and 14 months, respectively). The second course of rituximab was administered alone in 46 patients and with chemotherapy in 42 patients (13 CHOP, 11 FC, 6 COP, 5 ICE, 3 ESHAP, 2 HyperCVAD, 1 CNOP 1 VEEP). The addition of chemotherapy did not change the overall response (CR and PR) to the second rituximab treatment (72% in rituximab alone vs. 69% in rituximab with chemotherapy, in indolent lymphoma patients 76% vs. 72%, in aggressive lymphoma patients 70% vs. 62.5%, respectively). TTP after the second rituximab treatment was similar or possibly longer than after the first treatment (median 14 and 12 months, mean 23.4 and 16.2 months respectively). Conclusions: The response to a second rituximab treatment is the same whether the rituximab is given alone or combined with chemotherapy. TTP after retreatment with rituximab is as good as after the first treatment. Prospective studies should examine the treatment benefit of adding chemotherapy to a second treatment with rituximab.