Cell Chronic Lymphoproliferative Disorders Research Articles

SLAM Family Receptors in B Cell Chronic Lymphoproliferative Disorders.

The signaling lymphocytic activation molecule (SLAM) receptor family (SLAMF) consists of nine glycoproteins that belong to the CD2 superfamily of immunoglobulin (Ig) domain-containing molecules. SLAMF receptors modulate the differentiation and activation of a wide range of immune cells. Individual SLAMF receptors are expressed on the surface of hematopoietic stem cells, hematopoietic progenitor cells, B cells, T cells, NK cells, NKT cells, monocytes, macrophages, dendritic cells, neutrophils, and platelets. The expression of SLAMF receptors was studied during normal B cell maturation. Several SLAMF receptors were also detected in cancer cell lines of B-lymphoid origin and in pathological B cells from patients with B cell chronic lymphoproliferative disorders (B-CLPD), the most frequent hematological malignancies in adults. This review summarizes current knowledge on the expression of SLAMF receptors and their adaptor proteins SAP and EAT-2 in B-CLPD. Several SLAMF receptors could be regarded as potential diagnostic and differential diagnostic markers, prognostic factors, and targets for the development of novel drugs for patients with B-CLPD.

Clinical Features and Outcomes of the Patients with B-Cell Chronic Lymphoproliferative Disease in the New Drug Era

To analyze the clinical characteristics of the patients with B-cell chronic lymphoproliferative disease（B-CLPD） in the new drug era and the effect of new drug treatment on efficacy and survival. The clinical and laboratory data of 200 cases B-CLPD patients diagnosed between April 2015 and August 2021 were analyzed retrospectively. The clinical efficacy and survival of the patients under different treatments including Bruton tyrosine kinase（BTK） inhibitors， rituximab， and chemotherapy alone were analyzed. The prognostic factors affecting the survival of patients were analyzed by univarite analysis and multivariate analysis. There were 119 male（59.5％） and 81 female（40.5%） in 200 cases B-CLPD patients， the sex ratio(male/female) was 1.5∶1 with median age of 61（30- 91） years old. The distribution of subtypes were as fallows: 51 cases (25.5%) of chronic lymphocytic leukemia/small lymphocytic lymphoma（CLL/SLL）， 64（32.0%） cases of follicular lymphoma（FL）， 40（20.0%） cases mantle cell lymphoma（MCL）， 30（15.0%） cases of marginal zone lymphoma（MZL）， 10（5%） cases of lymphoplasmacytic lymphoma/waldenstrom macroglobulinemia（LPL/WM）， 5（2.5%） cases of B cell chronic lymphoproliferative disorders unclassified（B-CLPD-U） . The main clinical manifestation of 102 patients was lymph node enlargement, 32 cases were complicated with B symptoms. Among CLL/SLL patients， there were 12（23.5%） cases in Binet A and 39（76.5%） cases in Binet B/C. There were 29 patients（20.9%） in Ann Arbor or Lugano stage I-II and 110 cases（79.1%） in stage III-IV of other subtypes. The complete remission（CR） rate was 43.1%（25/58）， 40.2%（39/97）， 7.1%（1/14）， and overaIl response rate（ORR） was 87.9%（51/58）， 62.9%（61/97）， 28.6%（4/14） in the groups of BTK inhibitors， rituximab-based therapy， and chemotherapy alone. The 3-year OS rate and PFS rate in all patients was 79.2% and 72.4% respectively. The 3-year OS rate of patient with MZL, CLL/SLL, FL,WM was 94.7%, 87.7%, 86.8% and 83.3% respectively, while the 3-year OS rate of MCL was only 40.6%, which was significantly lower than other subtypes. The median OS of patients treated with BTK inhibitors and rituximab-based therapy was 20.5 and 18.5 months respectively， and the 3-year OS rate was 97.4% and 90.7%. However， the median PFS of patients receiving chemotherapy alone was 4 months， and the 1-year OS rate was 52.7%， which was statistically significant compared with the other two groups（P<0.05）. Univarite analysis showed that anemia， elevated lactate dehydrogenase， elevated β2-microglobulin， and splenomegaly were the poor prognostic factors for OS（P<0.05）， elevated lactate dehydrogenase was also poor prognostic factors for PFS（P<0.05）. Multifactor analysis showed that anemia and elevated lactate dehydrogenase were the independent poor prognostic factors for survival（P<0.05）. The clinical features of B-CLPD was various， anemia and elevated lactate dehydrogenase are the prognostic factors for poor survival. BTK inhibitors and new immunotherapy can improve the survival and prognosis of patients in the new drug era.

Role of CD200 in differentiating chronic lymphocytic leukaemia from mantle cell lymphoma and comparing its expression across various B cell chronic lymphoproliferative disorders

Multiparameter flow cytometry is a useful tool for the diagnostic evaluation of chronic lymphoproliferative disorder recently, it has been shown that CD200 may improve the distinction between chronic lymphocytic leukemia and mantle cell lymphoma, but the role of CD200 expression in atypical chronic lymphocytic leukemia and other chronic lymphoproliferative disorder remains to be established. Methods: A total of 200 samples were evaluated by Flow cytometry over a period of 2 years from august 2017 to October 2019. Cases were diagnosed according to WHO classification of tumours of hematopoietic and lymphoid tissues, 2017. Results: CD200 was strongly expressed in chronic lymphocytic leukemia and was revealed to be an excellent marker to distinguish chronic lymphocytic leukemia from mantle cell lymphoma, even in cases of atypical chronic lymphocytic leukemia. However, lack of CD200 was not an exclusive finding of mantle cell lymphoma, being also observed in other chronic lymphoproliferative disorder. Conclusion: CD200 was expressed by all cases of chronic lymphocytic leukemia as well as atypical chronic lymphocytic leukemia cases while it is negative in MCL thus it is particularly useful in distinguishing chronic lymphocytic leukemia/small lymphocytic lymphoma from other CD5+ B-cell neoplasms. CD200 is also expressed in most of the cases of hairy cell leukemia, splenic marginal zone lymphoma, and follicular lymphoma. CD200 is a useful addition to Moreau et al. panel for analysis of B cells in lymphoid tissue in Bone Marrow aspirates, and Peripheral Blood.

Clonal B-Cell Lymphocytosis with Mariginal-Zone Features: Comparison with Overt Splenic Marginal-Zone Lymphomas in 77 Patients from a Monocentric Series

Diagnosis of non-CLL B Cell Chronic Lymphoproliferative Disorders (BCLPD), defined by the detection of peripheral blood (PB) B-Cell clone with flow cytometry (FC) Matutesscore≤3, is not an infrequent event in clinical daily practice. Only a portion of these cases may be actually classified according to the 2016 revision of WHO classification of lymphoid neoplasms, as in these cases a non-bone marrow (BM) tissue biopsy is rarely available due to "liquid-only disease" (BM, PB, splenomegaly) or difficult accessible sites. The two most likely diagnosis of BCLPD according to WHO 2016 are splenic marginal-zone lymphoma (SMZL) and monoclonal B Cell lymphocytosis (MBL) with non-CLL phenotype (CD19+, CD20+, SIg+, CD5-), sometimes reported as MZL-like, defined by monotypic clonal B lymphocytes (CBL) &lt;5 x 109/l. A recent study pointed out a similar outcome regardless CBL value exceeding or not this arbitrary cut-off in strictly defined cases (absence of cytopenias, splenomegaly and adenopathy) of clonal B cell lymphocytosis with marginal-zone features (CBL-MZ) and confirmed the really indolent behaviour of this entity (Xochelli, Blood 2014). On these bases we decided to retrospectively collect clinical, immunophenotypic, molecular features and outcome of all consecutive patients (pts) with CD5- CBL-MZ owing to the Department of Hematology of University of Insubria (Varese, Italy) from 2011 to 2019 and to compare them with overt SMZL cases. Excluding 5 SMZL pts who were splenectomized, all cases were diagnosed by means of non-BM tissue biopsy, namely BM histology and PB/BM FC, FISH and molecular studies, when appropriate, and complete laboratory and imaging staging procedures (CT scan or US). The primary objective of the study was time to progression (TTP) for CBL-MZ group. Secondary objectives were overall survival (OS), progression-free survival (PFS) for treated pts and evaluation of early progression &lt;24 months (Early POD) (Luminari, Blood 2019). Overall we collected complete data of 33 pts with CD5- CBL-MZ, and 44 with SMZL (Table 1). Focusing on CBL-MZ, FC immunophenotypic findings showed a B-Cell population with Matutes score 0 to 2 in all cases, uniform expression of B Cell antigens, namely CD19, CD20 strong, FMC7 (93%), SIg (intermediate/bright in 71%), while CD10 and CD5 were consistently negative. CD23 was expressed in 18%, CD38 in 9% and CD49b in 95%. BM sinusoidal infiltration pattern was detected in 50% of cases. TP53 mutation/deletion was detected in 4/16 pts (25%) while trisomy 12 and del13q in 1 pt each. Twenty pts presented with &lt;5000/mmc CBL (MBL-MZ) and 13 with ≥5000/mmc CBL. With a median follow-up of 1.8 years, 6 pts (18%) progressed (4 in SMZL, 1 NMZL and 1 SLL; Fig 2) with a median TTP of 1.7 yrs (range 0.3- 12.3), while the remaining 27 remained completely stable. Among 4 pts treated, 2 received Rituximab-based regimen, both achieving CR and 2 with Chlorambucil both experiencing progression. Overall, 5 pts died (2 for progressive lymphoma), with an estimated 5-yrs of 80%. Concerning prognostic factors, after adjusting by competing risk method, at univariate analysis age &gt;60 yrs, CBL &gt;5000/mmc and albumin &lt;3.5 g/dl, were associated to shorter TTP, while age &gt;60 yrs, Hb &lt;11 g/dl, ECOG PS ≥2 and albumin &lt;3.5 g/dl resulted predictive of worse OS (Table 2). By applying multivariate Cox regression model, only age &gt;60 yrs and CBL&gt;5000/mmc retained prognostic significance for TTP (p&lt;0.0001), while ECOG PS ≥2 and albumin &lt;3.5 g/dl remained independently associated with inferior OS (p&lt;0.0001). Considering the SMZL group, 34 pts underwent systemic treatment (rituximab-based in 32, including R-Bendamustine in 17). With a median follow-up time of 2.8 yrs, 9 pts progressed (4 of whom within 24 months from treatment initiation), with a median PFS 2.9 yrs, and 8 died (7 due to progression, including 3 DLBCL transformations) with an estimated 5-yrs OS of 79%. Notably, Early POD SMZL pts exhibited a significant increased risk of death (p&lt;0.0001, fig.2), while in CBL-MZ did not (p=0.4). In conclusion, CBL-MZ confirmed to be associated with indolent course and preferential evolution toward SMZL, of which it can be viewed as an initial phase or a precursor entity. In contrast with previous findings, the WHO 16 CBL cut-off defining MBL (5 x 109/l), together with advanced age, retained prognostic relevance for TTP. Finally, we confirmed for the first time in an independent series of SMZL the highly prognostic impact of Early POD24. Disclosures Passamonti: Roche: Consultancy; Janssen: Consultancy, Speakers Bureau; Celgene: Consultancy, Speakers Bureau; Novartis: Consultancy, Honoraria, Speakers Bureau.

Characteristics of CD180 Expression and Its Diagnostic Value in B Cell Chronic Lymphoproliferative Disorders

To investigate the charactcristics of CD180 expression and differentiation diagnostic value in B cell chronic lymphoproliferative disorders (B-CLPD) through detecting the mean fluorescence intensity（MFI）of CD180 in different sub types of B-CLPD，using multiparameter flow cytometry (FCM). The CD180 MFI of malignant B cells in 178 patients with B-CLPD was detected by FCM. The level of CD180 MFI in various types of B-CLPD was compared to the normal control group. The level of CD180 MFI among sub-types of B-CLPD was also compared. (1) The expression levels of CD180 in B-CLPD was significantly lower as compared with the normal controls，except the spleen difuse red pulp lymphoma (SDRPL); (2) The CD180 MFI in chronic lymphocytic leukemia sCLL) was significantly lower as compared with other B-CLPD cells; (3) CD180 ware significantly overexpressed in HCL compared with MCL，LPL，and MZL (P <0.05); (4) In the spleen-derived B-CLPD，such as SMZL，HCL, HCL variation and SDRPL，the expression of CD180 has significant difference between lymphomas with or without villous. Utilizing the multiparameter flow cytometry for defecting expression of CD180 and other immunological markers can more efficiently distinguish the subtypes of B-CLPD.

Flow Cytometric Immunophenotyping Is Sensitive for the Early Diagnosis of De Novo Aggressive Natural Killer Cell Leukemia (ANKL): A Multicenter Retrospective Analysis.

Aggressive natural killer cell leukemia (ANKL) is a fatal hematological neoplasm characterized by a fulminating clinical course and extremely high mortality. Current diagnosis of this disease is not effective during the early stages and it is easily misdiagnosed as other NK cell disorders. We retrospectively analyzed the clinical characteristics and flow cytometric immunophenotype of 47 patients with ANKL. Patients with extranodal NK/T cell lymphoma, nasal type (ENKTL) and chronic lymphoproliferative disorder of NK cell (CLPD-NK), who were diagnosed during the same time period were used for comparisons. Abnormal NK cells in ANKL were found to have a distinctiveCD56bright/CD16dim immunophenotype and markedly increased Ki-67 expression, whereas CD57 negativity and reduced expression of killer immunoglobulin-like receptor (KIR), CD161, CD7, CD8 and perforin were exhibited compared with other NK cell proliferative disorders (p<0.05). The positive rates of flow cytometry detection (97.4%) was higher than those of cytomorphological (89.5%), immunohistochemical (90%), cytogenetic (56.5%) and F-18 fluorodeoxyglucose positron emission tomography/computer tomography (18-FDG-PET/CT) examinations (50%) (p<0.05). ANKL is a highly aggressive leukemia with high mortality. Flow cytometry detection is sensitive for the early and differential diagnosis of ANKL with high specificity.

Progress of response evaluation and prognosis in chronic lymphocytic leukemia: reports from the 56th American Society of Hematology annual meeting

Chronic lymphocytic leukemia(CLL)is an incurable B cell chronic lymphoproliferative disorder with a highly heterogeneous clinical course. The malignant B lymphocytes that have a mature appearance infiltrate in to lymph node, bone marrow, liver and spleen. The 56th American Society of Hematology(ASH)annual meeting explored multiple aspects including pathogenesis, treatment, response evaluation and prognosis of CLL. Clinical outcome of CLL patients can be predicted more accurately by a series of novel markers such as minimal residual disease. The progress of response evaluation and prognosis in CLL were focused in this review. Key words: Chronic lymphocytic leukemia; Response evaluation; Prognosis; American Society of Hematology annual meeting

Acquired Von Willebrand Syndrome Associated with B Cell Chronic Lymphoproliferative Disorders. Results from a Prospective Observational Study

▪Introduction. Whereas von Willebrand disease is the most common constitutional bleeding disorder, acquired von Willebrand syndrome (AVWS) is rare with an estimated prevalence between 0.04 to 0.5%(1). AVWS has been related in various pathophysiological conditions including cardiovascular diseases such as aortic stenosis, autoimmune and lymphoproliferative disorders. To our knowledge, the prevalence of AVWS in lymphoproliferative disorders has never been investigated.Methods. We conducted an observational monocentric prospective study in Caen university hospital to evaluate the incidence of AVWS in B cell chronic lymphoproliferative disorders (B-CLPD) and characterize its phenotype. Inclusion criteria were the presence of a BCLPD in patients with no personal or family history of bleeding. Every enrolled patients was tested for Biological parameters were measured including closure time, von Willebrand Antigen (VWF:Ag), ristocetin cofactor activity (VWF:RCo) and factor VIII procoagulant activity (FVIII:C). The bleeding phenotype was evaluated using Tossetto bleeding score(2). AVWS was suspected when patients presented a decrease of VWF:Ag and VWF:RCo and/or a VWF:RCo/VWF:Ag below 0.7.Results. A total of 147 patients were included with the following diagnosis: fifty five patients (37%) with chronic lymphocytic leukemia, 48 patients (33%) with monoclonal gammopathies of undetermined significance (MGUS), 22 patients (15 %) with non hodgkin B cell lymphoma, 9 patients (6 %) with multiple myeloma, 4 patients (3 %) with Waldenstrom macroglobulinemia and 9 patients with other B-CLPD. Closure times, with epinephrine and ADP, were prolonged for six patients (6/147, 4.1%) with median levels of VWF:Ag, VWF:RCo, FVIII:C and VWF:RCo/VWF:Ag ratio at 29.5 IU/dL [9-284], 11.4 IU/dL [1-140] , 42.5 [6-204] and 0.3 [0.04-0.84], respectively. Five of these 6 patients had MGUS and 1 patient presented a follicular lymphoma. Serum protein electrophoresis revealed a monoclonal component in 5 patients with a median concentration at 8.45 g/L [4.4-9.1]. Four out of these 6 patients presented mucocutaneous bleedings including menorragia, ecchymoses, epistaxis, gingival bleedings, post-operative bleedings and gastro intestinal bleedings. The median bleeding score of these six patients was 4.5 [-1 – 12]. In four patients, the biological phenotype was a type 2 von Willebrand disease, with decreased VWF:RCo/VWF:Ag ratio (<0.7), loss of high and intermediate molecular weight multimers and a decrease of von Willebrand binding to collagen (VWF:CB) (VWF:CB/VWF:Ag<0.6). Anti-von Willebrand factor inhibitor screening was negative for the six patients. Four of these 6 patients presented an accelerated clearance of VWF with increased von willebrand factor propeptide/VWF:Ag ratios.Table 1Different proposed cutoffs with their prospective sensitivity, specificity, and likelihood ratios (LR).Cutoff >Sensitivity %95% CISpecificity %95% CILR +LR -503.50100.0066.37% to 100.0%0.300.007557% to 1.652%1.000.002514.0088.8951.75% to 99.72%75.2270.24% to 79.76%3.590.155020.0077.7839.99% to 97.19%85.6781.46% to 89.24%5.430.26Conclusion: In this original prospective study, we observed AVWS for 4.1% of patients with B-CLPD. Monoclonal gammopathies of undetermined significance (MGUS) were associated with AVWS in 5 of 6 patients with AVWS. For patients with type 2 phenotypes, an increased clearance of VWF was the main mechanism of AVWS. The annual follow-up of these patients will give informations regarding the time of appearance and clinical informations for AVWS.Tiede A et al. Blood 2011.Tosetto A et al. Blood Rev 2007. DisclosuresNo relevant conflicts of interest to declare.

Expert consensus on diagnosis of B cell chronic lymphoproliferative disorders in China (2014)

Expert consensus on diagnosis of B cell chronic lymphoproliferative disorders in China (2014)

Association Of More Than One B Cell Clone Arises In Any Type Of Leukemic Chronic B Cell Disorder

The identification of more than one clone in B cell chronic lymphoproliferative disorders (B-CLPD) is known to occur occasionally. Nonetheless, to our knowledge, there is no extensive study of their incidence, their clinical and biological features, and their prognosis at diagnosis.At our institution, between 01/2006 and 12/2012, 1152 cases of chronic lymphoproliferative disorders with leukemic phase were diagnosed. Among those cases we have identified 44 cases (3,8%) with at least two B cell clones at diagnosis through systematic immunophenotyping of blood lymphocytes. We only considered cases in which both clones were of B cell origin and present in blood. Association of two haematological disorders arising from cells of different origin (i.e. myeloid or T cells) were excluded. Size of smallest second clone identified was 1% of B cells, in 11/44 cases it was less than 10% whereas in the remaining cases, both clones had up to approximately the same size.We aimed at further characterize these biclonal B-CLPD with leukemic phase. Discrimination of the clones relied on a combination of at least 2 phenotypic markers (presence or absence and not on level of expression). Diagnosis of biclonality relied on different surface immunoglobulin light chains in 21 cases. In the remaining 23 cases, displaying the same light chain, genescan analysis showed the presence of two peaks. Further cloning allowed to identify the presence of two functional rearrangements in 12 cases. Among those, IGHV were unmutated in both clones in 4 cases, mutated in both clones in 6 cases, and mixed (mutated/unmutated) in 2 cases.Based on clinical presentation, cytology and immunophenotype analysis and molecular features, the various B CLPD clones were identified the following: monoclonal B lymphocytosis (MBL) in 27 cases, chronic lymphocytic leukemia (CLL) in 14 cases, CD5+ atypical chronic B-CLPD in 22 cases, marginal zone lymphoma (MZL) in 12 cases, CD5- atypical B-CLPD in 11 cases, Waldenström macroglobulinemia (WM) in 4 cases, mantle cell lymphoma (MCL) in 3 cases and hairy cell leukemia (HCL) in 1 case.All possible associations between the various B-CLPD were seen: CD5+ B-CLPD associated with either CD5+ or CD5- B CLPD. The predominant clone was CD5+ in 56,8% cases, whereas the second clone was CD5+ in the majority of cases (82% cases). Therefore, presence of two concomitant CD5 negative clones was rare. This is exemplified in the Circos representation below (figure 1) [Display omitted] Four patients displayed an association of three clones and one patient had four.Analysis of IGHV gene sequence did not show a preferential use of any VH. Fifteen patients had unmutated IGHV genes, including CD5 negative cases. Of note, in CLL patients, 60% cases had unmutated IGHV genes, which is higher than what is normally observed in CLL at diagnosis. At presentation, most cases showed features of indolent disease; lymphnodes were present in 13/44 cases (29%), splenomegaly 10/44 cases (22%). Median Hb level was 13,2 g/dl (12,1 – 14,1 g/dl ) platelets level was 199 G/l (132 – 245 G/l). Mild cytopenias were observed in 5 patients, all with MZL, and likely related to splenomegaly.Follow up data were available for 35 cases. 23 /35 cases remained untreated with a median follow up of 16,2 months. Among those who required treatment, different chemotherapy regimen were used and 66 % patients are in remission after this first line of treatment.In conclusion, in this large prospective cohort, biclonal B CLPD were observed in less than 5% cases, and do not appear related to an adverse prognosis. The absence of predominant association, or of VH usage, and the presence of both IGHV mutated and unmutated clones do not favor a major role for the antigen to explain the presence of two clones at diagnosis in the same patient. This suggest that the mechanisms leading to the emergence of a second clone are not specific of one particular disease but may be common to all chronic lymphoproliferative disorders of B cell origin. Disclosures:No relevant conflicts of interest to declare.

An immunophenotypic and molecular diagnosis of composite hairy cell leukaemia and chronic lymphocytic leukaemia

Hairy cell leukaemia (HCL) and chronic lymphocytic leukaemia (CLL) are distinct clinicopathological B cell chronic lymphoproliferative disorders (B-CLPD). Both diseases have characteristic immunophenotypic and molecular features. The co-existence of two B-CLPD is perhaps more common than previously thought but a composite HCL and CLL has been rarely documented. A case is reported in which the morphology, integrated with an extensive immunophenotyping panel, and incorporation of the recently described HCL-associated BRAF V600E mutation, enabled the prompt diagnosis of composite HCL and CLL thus allowing appropriate treatment selection. This case serves to highlight the benefit of a multidisciplinary approach to the diagnosis of bi-clonal B-CLPD.

Aggressive mature natural killer cell neoplasms: from epidemiology to diagnosis.

Mature natural killer (NK) cell neoplasms are classified by the World Health Organization into NK/T cell lymphoma, nasal type (NKTCL), aggressive NK-cell leukemia (ANKCL) and chronic lymphoproliferative disorders of NK-cells, the latter being considered provisionally. NKTCL and ANKCL are rare diseases, with higher prevalence in Asia, Central and South America. Most NKTCL present extranodal, as a destructive tumor affecting the nose and upper aerodigestive tract (nasal NKTCL) or any organ or tissue (extranasal NKTCL) whereas ANKCL manifests as a systemic disease with multiorgan involvement and naturally evolutes to death in a few weeks. The histopathological hallmark of these aggressive NK-cell tumors is a polymorphic neoplastic infiltrate with angiocentricity, angiodestruction and tissue necrosis. The tumor cells have cytoplasmatic azurophilic granules and usually show a CD45+bright, CD2+, sCD3-, cytCD3epsilon+, CD56+bright, CD16−/+, cytotoxic granules molecules+ phenotype. T-cell receptor genes are in germ-line configuration. Epstein-Barr virus (EBV) -encoded membrane proteins and early region EBV RNA are usually detected on lymphoma cells, with a pattern suggestive of a latent viral infection type II. Complex chromosomal abnormalities are frequent and loss of chromosomes 6q, 11q, 13q, and 17p are recurrent aberrations. The rarity of the NK-cell tumors limits our ability to standardize the procedures for the diagnosis and clinical management and efforts should be made to encourage multi-institutional registries.

Pathophysiologic mechanisms and management of neutropenia associated with large granular lymphocytic leukemia

Large granular lymphocyte (LGL) syndrome includes a spectrum of clonal T cell and natural killer cell chronic lymphoproliferative disorders. These conditions are thought to arise from chronic antigenic stimulation, while the long-term survival of the abnormal LGLs appears to be sustained by resistance to apoptosis and/or impaired survival signaling. T-cell LGL (T-LGL) leukemia is the most common LGL disorder in the Western world. Despite its indolent course, the disease is often associated with neutropenia, the pathogenesis of which is multifactorial, comprising both humoral and cytotoxic mechanisms. This article addresses the pathogenesis of T-LGL leukemia and natural killer cell chronic lymphoproliferative disorder, as well as that of T-LGL leukemia-associated neutropenia. Furthermore, as symptomatic neutropenia represents an indication for initiating treatment, available therapeutic options are also discussed.

Absence of JAK-2 (Val617Phe) Point Mutations in Human Plasma Cell Myeloma.

Multiple myeloma (MM) is a terminal differentiated B cell chronic lymphoproliferative disorder characterized by the latent accumulation of plasma cells with a low proliferative index and an extended life span in bone marrow or extramedullary tissues. While studies on neoplastic cells isolated directly from MM patients or on MM-derived cell lines demonstrated that IL-6 is a major growth and survival factor for initiation of signaling in MM cells in an autocrine or paracrine manner, some MM cells independent on extracellular signaling such as IL-6 have been reported, suggesting other mechanisms may be involved in signal transduction pathways in MM cells. Accumulating evidence showed that IL-6 induces intracellular signaling through members of the signal transducers and activators of transcription (STAT) family of proteins by activating the janus kinase(JAK) family of protein tyrosine kinases, which subsequently phosphorylate and activate cytoplasmic Stat proteins. Activated Stat proteins dimerize and translocate to the nucleus, where they bind to specific DNA response elements and induce expression of Stat-regulated genes. Our previous studies demonstrated that one of the Stat family proteins, Stat3, was constitutively activated (phosphorylated) in the majority of MM patients, suggesting constitutive activation of JAK-2 kinase activities. However, other mechanisms might be also responsible JAK2 constitutive activation in MM cells, in particular in IL-6 independent MM cells. A specific point mutation of JAK-2 kinase gene has been identified recently showing its presence in higher frequency of certain types of chronic myeloproliferative disorders. The point mutation yields constitutive activation of JAK-2 kinase activity in the absence of extracellular signaling and plays an important role in the pathogenesis of those disorders. In this study, we try to determine whether JAK-2 (Val617Phe) point mutation was present in MM patients and if so, what kind of role it may play in the pathogenesis of MM. JAK-2 (Val617Phe) point mutation was analyzed by using an allele-specific polymerase chain reaction followed by separation and detection with capillary electrophoresis. The test has a detection sensitivity of up to 200 picogram mutated DNA. Bone marrow aspirates with various degrees of involvement by MM (ranged10–80%) were selected and all the cases have been verified by morphologic examination and demonstrated to have positive immunoglobulin heavy or kappa light gene rearrangements. Bone marrow specimens from total 59 MM patients were tested for JAK2 (Val617Phe) mutation and none of any MM cases with the specific point mutation were identified (0/59). The results indicated that JAK2 (Val617Phe) mutation was very infrequency or absence in MM patients, despite the fact that constitutional activation of JAK2 kinase as well as Stat3 activation was commonly seen in MM patients. The study suggests that other yet unknown mechanisms may also involve JAK2 activation, particularly in IL-6 independent MM cells.

CD11b Is Useful in the Diagnosis of Chronic Lymphocytic Leukemia/Prolymphocytic Leukemia, Mixed Chronic Lymphocytic Leukemia, and Prolymphocytic Leukemia.

CD11b is well known as an integrin, Mac-1, is often complexed with CD18, and is found on monocytes, granulocytes, and natural killer cells. It also serves as a receptor for iC3b. However, its occurrence in B cell chronic lymphoproliferative disorders is not generally recognized and has not been fully evaluated. To address this issue, a series of B cell leukemias and lymphomas referred for primary diagnosis was evaluated for the presence of CD11b. The purpose was to determine the frequency of its expression on these tumors and to evaluate its diagnostic value.Consecutive cases referred for flow cytometry as possible lymphoproliferative disease were analyzed. Included were bone marrow, peripheral blood, and lymph nodes. All cases were diagnosed according to the WHO classification based on immunophenotypic, morphologic, and clinical findings. The morphologic criteria of Melo (1986) and Bennett (1989) were used for classification of chronic lymphocytic leukemia (CLL), CLL/prolymphocytic leukemia (CLL/PLL), mixed CLL, and PLL. Cases identified as not related to chronic lymphocytic leukemia or prolymphocytic leukemia were recorded but not further analyzed. Similarly, lymph node and spleen-based tumors were excluded from the final analysis.CD11b was present on cells from 32 of 123 cases, including occasional follicular lymphoma, (5/35); mantle cell lymphoma, (1/8); diffuse large B cell lymphoma, (3/9); hairy cell leukemia, (3/5); multiple myeloma, (1/2); lymphoplasmacytic lymphoma, (2/2); nodal marginal zone lymphoma, 0/1); and splenic marginal zone lymphoma, (1/1). However, it was most consistently expressed on CLL that contained increased numbers of prolymphocytes or large cells and on PLL. A total of 16 such cases were found. Morphologic assessment showed them to include 8 CLL/PLL, 3 mixed CLL, 4 PLL, and 1 typical CLL. The typical CLL case included both large cells and prolymphocytes but did not have more than 10% PLs. Five of the 16 cases (31%) were negative for CD5, CD23, and CD38 but were positive for FMC-7. In contrast, the other 11 cases were all CD5(+) and CD23(+); 3/11 were positive for CD38; and 5/11 were positive for FMC-7. Forty-five CLLs also were identified during the study, of which 27 had sufficient data for comparison. Twenty-six of the 27 CLLs were morphologically typical. The remaining case was mixed CLL. All of the CLLs were CD11b(−), CD5(+) and CD23(+); 15/43 were CD38(+), and 6/43 were FMC-7(+).The findings show that CD11b is expressed on chronic B cell lymphoproliferative disorders. In particular, it is expressed on almost all CLL cases that contain large cells or prolymphocytes and on PLL. Inclusion of CD11b in routine screening panels of possible chronic B cell leukemiaa will improve diagnosis of these disorders.

Fludarabine: an effective treatment in patients with splenic lymphoma with villous lymphocytes.

Splenic lymphoma with villous lymphocytes (SLVL) is a B cell chronic lymphoproliferative disorder. Splenectomy and/or chlorambucil are usually regarded as the most effective treatment in SLVL patients. However, a few patients relapse and the second-line treatment remains questionable. In a retrospective study, we evaluated the efficacy and toxicity of fludarabine (FDR) in 10 SLVL patients. The median duration between diagnosis and treatment was 17 months (range, 1-30). Two patients were previously untreated. The patients received FDR 25 mg/m2/day by venous infusion for 5 days with a median of four cycles of chemotherapy (range, 2-6). All patients were assessable: five patients achieved a good and persistent response after a median follow-up of 14 months (5-31), two achieved a good response but relapsed after a follow-up of 15 and 36 months. One out of the three partial responders have a persistent response. The treatment was well tolerated. FDR appears to be an efficient therapy with a favorable toxicity profile for patients in relapse after splenectomy or resistant to CLB. Furthermore it could constitute an alternative to splenectomy in older patients. A longer follow-up and the study of a larger group of patients are warranted to confirm our findings.

Cyclin D1 overexpression allows identification of an aggressive subset of leukemic lymphoproliferative disorder.

The conjunction of clinical features, cell morphology and immunological characteristics allows an accurate diagnosis in most cases of B cell chronic lymphoproliferative disorders (CLD). However, the diagnosis remains uncertain in a small percentage of cases, often referred as to unclassified B cell proliferation or atypical chronic lymphocytic leukemia (CLL). We have studied retrospectively the 192 cases of leukemic CLD seen in our institution over a 3-year period, for which both clinical and routine biological data at presentation were available. Forty cases (20%) did not fit into any of the well-identified categories according to the FAB criteria and remained unclassified. We assessed cyclin D1 expression in all of these cases and found that 10 of them expressed a high level of cyclin D1 protein. We compared the characteristics of these 10 cases with those of the 30 cyclin D1 negative CLD. Despite non-distinctive cytological and phenotypic features, the 10 cyclin D1 positive patients exhibited a strikingly uniform clinical presentation with elevated leukocytosis, massive spleen enlargement and no superficial lymphadenopathy. Their outcome was very poor with a median survival of 10 months, contrasting with the prolonged survival of the cyclin D1 negative patients. The cytological features of tumor cells from these 10 patients with cyclin D1 positive unclassified leukemic CLD were similar to those of the circulating lymphoid cells from 15 patients with histologically proven mantle cell lymphoma (MCL) and primary or secondary blood involvement. Therefore, cyclin D1 expression allowed identification among the unclassified CLD, a subset of aggressive disorders which represent a leukemic counterpart of MCL (mantle cell leukemia). We suggest that determination of cyclin D1 expression by any technique available should be systematically included when investigating atypical CLL.

Functional Adrenocortical Adenoma in a Case with B Cell Chronic Lymphoproliferative Disorder

We describe a B cell chronic lymphocytic leukemia/lymphoma (B-CLL) patient with an adrenocortical adenoma. He was treated initially with oral cyclophosphamide and prednisolone for 2 months, followed by low-dose prednisolone for an additional 2 years, which resulted in the prompt disappearance of CLL cells. After a period of 2 years without therapy, a functional adrenocortical adenoma was found. After adrenalectomy, the CLL cells rapidly increased. Progression of the disease and clinical-hematological relapse was well controlled by low doses of steroids alone, and the patient's condition stabilized. Considering the recent data that corticosteroids can induce apoptosis of CLL cells, the above clinical course suggests that augmented glucocorticoid production from the tumor may have suppressed disease progression.

The optimal management of hairy cell leukaemia.

Hairy cell leukaemia is an uncommon B cell chronic lymphoproliferative disorder characterised by circulating lymphocytes displaying prominent cytoplasmic projections. Therapy is initiated for severe cytopenias or recurrent infections. Splenectomy, the first standard treatment, is now less commonly used as primary treatment. Interferon-alpha (IFN alpha) induces partial responses in most patients but complete responses in only a few. Adverse effects from IFN alpha are common but not life-threatening. The ability of two newer purine analogues, pentostatin (2'-deoxycoformycin) and cladribine (2-chlorodeoxyadenosine), to induce long-lasting complete remissions in the majority of patients has revolutionised the treatment of this disease. Cladribine is emerging as the treatment of choice because of its favourable toxicity profile, brief duration of treatment, high percentage of unmaintained complete remissions and low incidence of relapse.

2 Chronic lymphoproliferative disorders: Classification and diagnosis

The chronic lymphoproliferative disorders are morphologically, immunologically and clinically heterogeneous. Common features of these processes include T, B or natural killer cell immunophenotypes and terminal deoxy-nucleotidyl transferase negativity. The B cell lymphocytic disorders include B-chronic lymphocytic leukaemia, B cell prolymphocytic leukaemia, chronic lymphocytic leukaemia-prolymphocytic leukaemia, non-Hodgkin's lymphoma (including mantle cell lymphoma) in leukaemic phase, hairy cell leukaemia and splenic lymphoma with villous lymphocytes. The T cell chronic lymphoproliferative disorders include prolymphocytic leukaemia, adult T cell leukaemia-lymphoma, large granulated lymphocyte leukaemia and Sézary syndrome. Occasionally, a lymphocytic proliferation is encountered that does not satisfy the morphological or immunophenotypical criteria for any of the above categories. These processes are best left unclassified.