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Splenic diffuse red pulp lymphoma is an indolent small B-cell lymphoma recognized as a provisional entity in the World Health Organization 2008 classification. Its precise relationship to other related splenic B-cell lymphomas with frequent leukemic involvement or other lymphoproliferative disorders remains undetermined. We performed whole-exome sequencing to explore the genetic landscape of ten cases of splenic diffuse red pulp lymphoma using paired tumor and normal samples. A selection of 109 somatic mutations was then evaluated in a cohort including 42 samples of splenic diffuse red pulp lymphoma and compared to those identified in 46 samples of splenic marginal zone lymphoma and eight samples of hairy-cell leukemia. Recurrent mutations or losses in BCOR (the gene encoding the BCL6 corepressor) - frameshift (n=3), nonsense (n=2), splicing site (n=1), and copy number loss (n=4) - were identified in 10/42 samples of splenic diffuse red pulp lymphoma (24%), whereas only one frameshift mutation was identified in 46 cases of splenic marginal zone lymphoma (2%). Inversely, KLF2, TNFAIP3 and MYD88, common mutations in splenic marginal zone lymphoma, were rare (one KLF2 mutant in 42 samples; 2%) or absent (TNFAIP3 and MYD88) in splenic diffuse red pulp lymphoma. These findings define an original genetic profile of splenic diffuse red pulp lymphoma and suggest that the mechanisms of pathogenesis of this lymphoma are distinct from those of splenic marginal zone lymphoma and hairy-cell leukemia.

CD19 is widely used as a marker of B-cell lineage because it is expressed as early as the pre-B stage and repressed only during terminal differentiation into plasma cells.[1][1] This transmembrane glycoprotein is an essential co-receptor of the B-cell receptor (BCR),[2][2] although it can also be

Dysregulation of MYC is the genetic hallmark of Burkitt lymphoma (BL) but it is encountered in other aggressive mature B-cell lymphomas. MYC dysregulation needs other cooperating events for BL development. We aimed to characterize these events and assess the differences between adult and paediatric BLs that may explain the different outcomes in these two populations. We analysed patterns of genetic aberrations in a series of 24 BLs: 11 adults and 13 children. We looked for genomic imbalances (copy number variations), copy-neutral loss of heterozygosity (CN-LOH) and mutations in TP53, CDKN2A, ID3 (exon 1), TCF3 (exon17) and CCND3 (exon 6). Young patients displayed more frequent 13q31.3q32.1 amplification, 7q32q36 gain and 5q23.3 CN-LOH, while 17p13 and 18q21.3 CN-LOH were only detected in adult BLs. ID3 mutations were present in all adult samples, but only in 42% of childhood cases. CCND3 and ID3 double-hit mutations, as well as 18q21 CN-LOH, seemed to be associated with poorer outcome. For the first time, we report different genetic anomalies between adult and paediatric BLs, suggesting age-related heterogeneity in Burkitt lymphomagenesis. This may explain the poorer prognosis of adult BLs. Additional studies are needed to confirm these results in the setting of clinical trials.

International audience; In splenic marginal zone lymphoma (SMZL), specific and functional Toll-like Receptor (TLR) patterns have been recently described, suggesting their involvement in tumoral proliferation. Splenic diffuse red pulp lymphoma with villous lymphocytes (SDRPL) is close to but distinct from SMZL, justifying here the comparison of TLR patterns and functionality in both entities. Distinct TLR profiles were observed in both lymphoma subtypes. SDRPL B cells showed higher expression of TLR7 and to a lesser degree TLR9, in comparison to SMZL B cells. In both entities, TLR7 and TLR9 pathways appeared functional, as shown by IL-6 production upon TLR7 and TLR9 agonists stimulations. Interestingly, circulating SDRPL, but not SMZL B cells, constitutively expressed CD86. In addition, stimulation with both TLR7 and TLR9 agonists significantly increased CD80 expression in circulating SDRPL but not SMZL B cells. Finally, TLR7 and TLR9 stimulations had no impact on proliferation and apoptosis of SMZL or SDRPL B cells. In conclusion, SMZL and SDRPL may derive from different splenic memory B cells with specific immunological features that can be used as diagnosis markers in the peripheral blood.

To the editor: Most hematologic malignancies are preceded by a premalignant condition. In myeloid neoplasms, genetically defined premalignant conditions such as clonal hematopoiesis of indeterminate potential[1][1] reflect the clonal dominance of hematopoietic stem cells harboring oncogenic

MALT1, a paracaspase, forms together with CARD11 and BCL10 a trimolecular complex, and, both as a scaffold protein constitutively associated with BCL10 and as a protease, is essential in B- and T-c...

International audience; Splenic Diffuse Red Pulp Lymphoma (SDRPL) has been recently introduced as a provisional entity but differential diagnosis with other splenic lymphomas is needed to be clarified since the therapeutic approaches are distinct. Recently described recurrent mutations or CD180 expression appear useful for differential diagnosis. We completed our previous description in a larger cohort including 53 patients selected on the presence of characteristic villous cells in peripheral blood (PB) and a specific immunophenotype. Immunoglobulin heavy variable (IGHV), BRAF, MYD88, and NOTCH2 mutations were determined and CD180 and BRAF expressions were assessed. Most cases (79%) were IGHV mutated with an overrepresentation of IGHV3-23 (19%) and IGHV4-34 (21%). MYD88 L265P and NOTCH2 mutations were observed in one case each, whereas no BRAF V600E mutation or expression was found. All cases demonstrated a high CD180 expression. Those results strengthen the concept that SDRPL does emerge as a new lymphoma entity distinct from the other splenic lymphomas with circulating lymphocytes

Non-Hodgkin's lymphomas and lymphoproliferative disorders include a high number of heterogeneous entities, described in the 2008 WHO classification. This classification reflects the crucial role of a multidisciplinary approach which integrates cytogenetic results both for the notion of clonality and for differential diagnosis between these entities. The prognostic impact of some cytogenetic abnormalities or genome complexity is also confirmed for many of these entities. Novel provisional entities have been described, such as BCLU (B-cell lymphoma unclassifiable with features intermediate between diffuse large B-cell lymphoma and Burkitt lymphoma) for which karyotype is critical to distinguish BCLU from Burkitt's lymphoma. The karyotype can be established from any tumour or liquid infiltrated by lymphoma cells. Recent adaptations of technics for cellular cultures according to the subtype of known (or suspected) lymphoma have significantly improved the percentage of informative karyotypes. Conventional karyotypes remain the best technical approach recommended for most of these subtypes. Interphase and/or metaphase FISH also represents a solid and rapid approach, because of the significant number of recurrent (sometimes specific) rearrangements of these entities. Next generation sequencing technologies contribute to enrich genomic data and substantially improve the understanding of oncogenic mechanisms underlying these lymphoid malignancies. Some molecular biomarkers are already part of the diagnostic process (for example, somatic mutation of MYD88 in Waldenstrom disease) thus reinforcing the essential principle of a multidisciplinary approach for the diagnosis of all the mature lymphoid malignancies.

We previously showed that complex karyotypes (CK) and chromosome 13q abnormalities have an adverse prognostic impact in childhood Burkitt lymphomas/leukemias (BL) and diffuse large B-cell lymphomas (DLBCL). The aim of our study was to identify recurrent alterations associated with MYC rearrangements in aggressive B-cell lymphomas with CK. Multicolor fluorescence in situ hybridization (M-FISH) was performed in 84 patient samples (59 adults and 25 children), including 37 BL (13 lymphomas and 24 acute leukemias), 12 DLBCL, 28 B-cell lymphomas with intermediate features (DLBCL/BL), 4 B-cell precursor acute lymphoblastic leukemias (BCP-ALL), and 3 unclassifiable B-cell lymphomas. New (cytogenetically undetected) abnormalities were identified in 80% of patients. We also refined one-third of the chromosomal aberrations detected by karyotyping. M-FISH proved to be more useful in identifying chromosomal partners involved in unbalanced translocations and in revealing greater complexity of 13q rearrangements. Most of the newly identified or refined recurrent alterations involved 1q, 13q and 3q (gains/losses), 7q and 18q (gains), or 6q (losses), suggesting that these secondary aberrations may play a role in lymphomagenesis. Several patterns of genomic aberrations were identified: 1q gains in BL, trisomies 7 in DLBCL, and 18q-translocations in adult non-BL. BCP-ALL usually displayed an 18q21 rearrangement. BL karyotypes were less complex and aneuploid than those of other MYC-rearranged lymphomas. BCP-ALL and DLBCL/BL were associated with a higher rate of early death than BL and DLBCL. These findings support the categorization of DLBCL/BL as a distinct entity and suggest that BL with CK are indeed different from other aggressive MYC-rearranged lymphomas, which usually show greater genetic complexity. © 2012 Wiley Periodicals, Inc.

'Splenic red pulp lymphoma with numerous basophilic villous lymphocytes' (SRPL), recently described, is characterized by clinical, morphologic, immunologic, cytogenetic and molecular features distinct from SMZL/SLVL and HCL. In particular, the intensity of CD11c staining (expressed as fluorescence intensity -RFI-) in SRPL is significantly different from the RFI in SMZL/SLVL and HCL. Moreover the use of a scoring system based on the expression of CD11c, CD22, CD76, CD38 and CD27 appears to improve the differential diagnosis between SRPL and SMZL/SLVL and emphasizes that SRPL is an entity closed to but distinct from SMZL/SLVL. Copyright © 2010 John Wiley & Sons, Ltd.

Purpose Patients with diffuse large B-cell lymphoma (DLBCL) usually relapse early following diagnosis but some relapses happen at 5 years or later. Few data exist regarding clinical characteristics and outcome of these patients. Patients and Methods We performed a retrospective analysis of all patients from two centers in Lyon, France, between 1985 and 2003 who had a biopsy-proven relapse 5 years or later following diagnosis of DLBCL. All available biopsies were reviewed and immunohistochemistry was completed. Results Among 1,492 patients with DLBCL, 54 were eligible. At diagnosis, 63% of patients had stage I-II, 82% had low/low-intermediate International Prognostic Index (IPI) score, 65% had extranodal involvement, 24% had an indolent component associated with DLBCL, 57% had germinal center phenotype, and 43% had non-germinal center phenotype. Median time from diagnosis to relapse was 7.4 years (range, 5 to 20.5 years). At time of relapse, 83% had DLBCL histology, and 17% had indolent histology. Having an indolent component at diagnosis was associated with indolent histology at relapse (P = .028). Five-year event free-survival (EFS) was 17% for patients with DLBCL relapse and 61% for patients with indolent relapse (P = .027). Five-year overall survival was 27% for patients with DLBCL and 75% for patients with indolent relapse (P = .029). For DLBCL relapse, 3-year EFS was 56% versus 18% with autologous stem-cell transplantation or not, respectively (P = .0661). Conclusion Patients with DLBCL who had a late relapse usually had localized stage, favorable IPI score, and extranodal involvement at diagnosis. The outcome of patients with DLBCL at time of relapse remains poor, and aggressive treatment such as autologous stem-cell transplantation should be pursued whenever possible. Biopsy at relapse is essential because some patients relapse with indolent histology.

Since the initial description of splenic marginal zone lymphoma (SMZL) in 1992, an increasing number of publications have dealt with multiple aspects of SMZL diagnosis, molecular pathogenesis and treatment. This process has identified multiple inconsistencies in the diagnostic criteria and lack of clear guidelines for the staging and treatment. The authors of this review have held several meetings and exchanged series of cases with the objective of agreeing on the main diagnostic, staging and therapeutic guidelines for patients with this condition. Specific working groups were created for diagnostic criteria, immunophenotype, staging and treatment. As results of this work, guidelines are proposed for diagnosis, differential diagnosis, staging, prognostic factors, treatment and response criteria. The guidelines proposed here are intended to contribute to the standardization of the diagnosis and treatment of these patients, and should facilitate the future development of clinical trials that could define more precisely predictive markers for histological progression or lack of response, and evaluate new drugs or treatments.

Analysis of BCL-6, CD95, PIM1, RHO/TTF and PAX5 mutations in splenic and nodal marginal zone B-cell lymphomas suggests a particular B-cell origin

Chromosomal breakpoints affecting immunoglobulin (IG) loci are recurrent in many subtypes of B-cell lymphomas. However, despite the predominant B-cell origin of the Hodgkin and Reed-Sternberg (HRS) cells in classical Hodgkin lymphoma (cHL), the presence of chromosomal translocations in IG loci has not yet been systematically explored. Therefore, we have investigated a series of cHL for chromosomal breakpoints in the IGH (n = 230), IGL (n = 139), and IGK (n = 138) loci by interphase cytogenetics. Breakpoints in the IGH, IGL, or IGK locus were observed in the HRS cells of 26 of 149 (17%), 2 of 70, and 1 of 77 evaluable cHLs, respectively. The IG partners could be identified in eight cHLs and involved chromosomal bands 2p16 (REL), 3q27 (BCL6, two cases), 8q24.1 (MYC), 14q24.3, 16p13.1, 17q12, and 19q13.2 (BCL3/RELB). In 65 of 85 (76%) cHLs evaluable for an IGH triple-color probe, the HRS cells showed evidence for a (partial) deletion of the IGH constant region, suggesting the presence of class switch recombination (CSR). Furthermore, analyses with this probe in cases with IGH breakpoints indicated that at least part of them seem to be derived from CSR defects. Our results show that chromosomal breakpoints affecting the IG loci are recurrent in cHL. (Cancer Res 2006; 66(21): 10332-8)

Primary low-grade B-cell lymphomas of the skin are separated into marginal zone and follicle center lymphomas according to the recent World Health Organization-European Organization for Research and Treatment of Cancer classification, with distinct histologic and immunohistochemical profiles. Some cases remain difficult to distinguish. The degree of relationship with their extracutaneous counterparts is currently being investigated on clinical, histologic and molecular grounds. Cytogenetic analysis using fluorescence in situ hybridization was performed on 12 frozen samples of infiltrated skin that had been classified as marginal zone lymphoma (MZL). Chromosomal changes known to be recurrently observed in systemic MZL of the mucosa-associated lymphoid tissue type, and in follicular center lymphoma were analyzed. These included trisomy for chromosomes 3, 7, 12, and 18 as well as t(14;18) IGH/BCL2, t(14;18) IGH/MLT1, and t(11;18) API2/MLT1 translocations. Complementary molecular search of IGH/BCL2 rearrangement using a polymerase chain reaction technique and of API2/MLT1 mRNA expression by reverse transcriptase-polymerase chain reaction were performed. Two cases showed evidence of trisomy 3 at levels varying from 14% to 20% of the analyzed cells. No other chromosomal abnormalities were found with those techniques in the remaining cases. These results demonstrate that known recurrent chromosomal abnormalities rarely occur in primary cutaneous MZLs and suggest the possibility of a variety of initial oncogenic events leading to a common downstream pathway. These data also underline that fluorescence in situ analysis on routine skin punch biopsies represents a reliable tool for the detection of chromosomal changes, but requires consistent dermal infiltration.

Waldenstrom's macroglobulinemia (WM) is considered in the World Health Organization classification as a clinical syndrome associated with monoclonal immunoglobulin (Ig) M secretion, mainly observed in patients with lymphoplasmacytic lymphoma (LPL) and occasionally with other small B-cell lymphomas. Some authors consider it a rare distinct lymphoproliferative disorder with primary bone marrow infiltration and IgM monoclonal gammopathy. As LPL shares important morphologic and immunophenotypic overlaps with marginal zone B-cell lymphomas (MZLs) in cases showing plasmacytic maturation, it remains unclear if they constitute unique or distinct entities. Both diseases are composed of lymphocytes, lymphoplasmacytoid cells, and tumoral plasma cells with a surface (s) IgM-positive sIgD+/ cytoplasmic IgMpositive CD19+ CD20+ CD27+/ CD5 CD10 CD23 phenotype, without a specific marker. Extranodal mucosa-associated lymphoid tissue (MALT) lymphoma, nodal MZL (NMZL), and splenic MZL (SMZL) are distinct entities displaying common morphologic, immunophenotypic, and genetic characteristics. MALT lymphoma is clearly distinct from LPL, although bone marrow infiltration and IgM paraprotein are not rare. Splenic MZL and NMZL are incompletely characterized, but a plasmacytoid/plasmacytic differentiation, autoimmune manifestations, and monoclonal component are frequent in both diseases. Bone marrow involvement is constant in SMZL and present in 60% of NMZLs. Molecular IgVH gene analysis has confirmed this heterogeneity, particularly within SMZL, with mutated and unmutated cases. Further studies are needed to clarify the pathogenesis of these MZLs and their relationship with LPL.

Identification and validation of seven genes, as potential markers, for the differential diagnosis of small B cell lymphomas (small lymphocytic lymphoma, marginal zone B cell lymphoma and mantle cell lymphoma) by cDNA macroarrays analysis

We retrospectively analyzed 26 patients with thyroid lymphoma (TL). Patients were mostly females, with a median age of 59 yr, presenting a rapidly growing nodular goiter with or without cervical adenopathy, without symptoms related to lymphoma for 81% and hypothyroidism in 61%. A previous history of Hashimoto thyroiditis was observed in 11 patients. Six different subtypes of lymphoma were observed: 13 of 26 (50%) had diffuse large B cell lymphoma, 6 (23%) mucosa- associated lymphoid tissue (MALT) lymphoma, 3 (12%) had follicular lymphoma, 2 (7%) had Hodgkin's disease, 1 (4%) had small lymphocytic lymphoma, and 1 (4%) had Burkitt's lymphoma. Diffuse large B cell lymphoma patients presented a compressive multinodular goiter, cervical adenopathy (66%), disseminated disease (50%), and poor performance status, with a poor prognosis (5-yr survival at 44%) despite a treatment based on a multidrug regimen. MALT lymphoma arose in patients with previous history of Hashimoto disease, was localized in all but 1, and was biologically associated with hypothyroidism and a high level of serum antithyroid antibodies. With total thyroidectomy, prognosis was good (5-yr survival at 100%). We did not find any routine clinical or biological parameters that could predict the evolution from Hashimoto's thyroiditis to MALT lymphoma. In conclusion, we confirmed the histological heterogeneity of TL corresponding to different clinical presentations and different prognoses.

Some lymphoma subtypes, particularly marginal zone lymphoma (MZL), are associated with autoimmune diseases or chronic infections.[1][1],[2][2] Mucosa-associated lymphoid tissue (MALT) lymphomas derive from an acquired lymphoid tissue in the context of a chronic inflammation induced either by a

International audience; Follicular Lymphoma (FL) is the second most common non-Hodgkin lymphoma (NHL) subtype and its course is heterogeneous. At diagnosis, some patients with FL manifest a detectable leukaemic phase (FL-LP), but this feature has been seldom described and is poorly characterized. Among 499 patients diagnosed with FL in Lyon-Sud hospital, 37 (7·4%) had characteristic FL-LP (by cytological blood smears and flow cytometric analysis). In addition, 91/1135 FL patients from the PRIMA study presented FL-LP at study entry. In order to evaluate the outcome of this Lyon-Sud cohort, FL-LP patients were matched with 111 newly diagnosed FL without LP according to the Follicular Lymphoma International Prognostic Index (FLIPI) score, age and treatment. Presence of FL-LP was associated with shorter progression-free survival (PFS) and overall survival (OS) (P = 0·004 and P = 0·031, respectively). Presence of FL-LP and high FLIPI score remained independent prognostic factors in a Cox model for time to progression (TTP). A number of circulating lymphoma cells (CLC) >4 × 109/l was the most significant predictor for a shorter TTP in this Cox model. The prognostic impact of FL-LP on TTP was validated in the PRIMA cohort (P = 0·0004). In conclusion, FL-LP is a rare event associated with shorter PFS and patients with CLC >4 × 109/l have a poorer outcome. These patients should be monitored carefully to consider alternative therapeutic options.

Mucosa-associated lymphoid tissue (MALT) lymphomas most frequently involve the gastrointestinal tract and are the most common subset of extranodal non-Hodgkin lymphoma (NHL). Here we describe overexpression of BCL10, a novel apoptotic signalling gene that encodes an amino-terminal caspase recruitment domain (CARD), in MALT lymphomas due to the recurrent t(1;14)(p22;q32). BCL10 cDNAs from t(1;14)-positive MALT tumours contained a variety of mutations, most resulting in truncations either in or carboxy terminal to the CARD. Wild-type BCL10 activated NF-kappaB but induced apoptosis of MCF7 and 293 cells. CARD-truncation mutants were unable to induce cell death or activate NF-kappaB, whereas mutants with C-terminal truncations retained NF-kappaB activation but did not induce apoptosis. Mutant BCL10 overexpression might have a twofold lymphomagenic effect: loss of BCL10 pro-apoptosis may confer a survival advantage to MALT B-cells, and constitutive NF-kappaB activation may provide both anti-apoptotic and proliferative signals mediated via its transcriptional targets.

Mantle cell lymphoma (MCL), small lymphocytic lymphoma (SLL), and marginal zone lymphoma (MZL) are small B-cell non-Hodgkin lymphomas (NHLs) that may be difficult to distinguish. In order to identify specific proteomic biomarkers, differential proteomic analysis of these three NHLs was performed using surface enhanced laser desorption/ionization-time of flight mass spectrometry (SELDI-TOF-MS). Whole cell lysates obtained from 18 MCL, 20 SLL, and 20 MZL biopsies were applied on two different ProteinChips (cationic and anionic). Hierarchical clustering and discriminating scores combined with an innovative bio-informatics microdissection strategy allowed us to distinguish specific lymphoma proteomic signatures based on the expression of 37 protein peaks. SELDI-assisted protein purification combined with nano-liquid chromatography (LC) quadrupole-time of flight tandem mass spectrometry (Q-TOF MS/MS) was used to identify proteins overexpressed in both MCL and SLL tumors. Among them two histones, H2B and H4, were identified in MCL tumor biopsies and the signal recognition particle 9 kDa protein, SRP9, in SLL tumor biopsies.

Background Classically, splenic marginal zone B-cell lymphoma is characterized by the absence of CD5 expression. Cases of apparent splenic marginal zone B-cell lymphoma showing CD5 expression, as diagnosed by blood studies, have been described; however, in the absence of histological evidence, the correct diagnosis of these cases is controversial because of possible confusion with other CD5-positive small B-cell neoplasms.Design and Methods We report a series of 24 CD5-positive, t(11;14)-negative cases of splenic marginal zone B-cell lymphoma diagnosed by flow cytometry studies of blood and histologically proven on spleen sections. Clinical data as well as morphological, immunological, cytogenetic and molecular characteristics were assessed to evaluate the similarities and differences of these cases with those of classical CD5-negative splenic marginal zone B-cell lymphoma.Results The CD5 expression detected in blood by flow cytometry was confirmed in most cases by immunohistochemistry on spleen sections. In general, cases of CD5-positive and CD5-negative splenic marginal zone B-cell lymphoma did not appear different and, in particular, they showed similar karyotypic changes such as 7q deletion, trisomy 3, trisomy 18 and biased IGHV usage (i.e. VH1-2). The main differences were a higher lymphocyte count at diagnosis (8.15×109/L versus 3.90×109/L; P=0.005) and more frequent diffuse bone marrow infiltration (34% versus 8%; P=0.03) in the CD5-positive group. A tendency to a more mutated IGHV status in the CD5 positive cases was observed (80% versus 54.5%; (P=0.11). No significant differences in outcome were found in relation to CD5 expression.Conclusions This study confirms the existence of cases of CD5-positive splenic marginal zone B-cell lymphoma and shows that these cases are closely related to classical splenic marginal zone lymphoma. Whether or not CD5-positive splenic marginal zone B-cell lymphoma constitutes a true subset obviously requires the study of more cases.

The t(14;18)(q32;q21) involving the immunoglobulin heavy chain locus (IGH) and the MALT1 gene is a recurrent abnormality in mucosa-associated lymphoid tissue (MALT) lymphomas. However, the nucleotide sequence of only one t(14;18)–positive MALT lymphoma has been reported so far. We here report the molecular characterization of the IGH-MALT1 fusion products in 5 new cases of t(14;18)–positive MALT lymphomas. Similar to the IGH-associated translocations in follicular and mantle cell lymphomas, the IGH-MALT1 junctions in MALT lymphoma showed all features of a recombination signal sequence–guided V(D)J-mediated translocation at the IGH locus. Furthermore, analogous to follicular and mantle cell lymphoma, templated nucleotides (T-nucleotides) were identified at the t(14;18)/IGH-MALT1 breakpoint junctions. On chromosome 18, we identified a novel major breakpoint region in MALT1 upstream of its coding region. Moreover, the presence of duplications of MALT1 nucleotides in one case suggests an underlying staggered DNA-break process not consistent with V(D)J-mediated recombination. The molecular characteristics of the t(14;18)/IGH-MALT1 resemble those found in the t(14;18)/IGH-BCL2 in follicular lymphoma and t(11;14)/CCND1-IGH in mantle cell lymphoma, suggesting that these translocations could be generated by common pathomechanisms involving illegitimate V(D)J-mediated recombination on IGH as well as new synthesis of T-nucleotides and nonhomologous end joining (NHEJ) or alternative NHEJ repair pathways on the IGH-translocation partner.

The presence of circulating villous lymphocytes (VLs) in lymphoma patients usually points to splenic marginal zone B-cell lymphoma (SMZL), even if the VLs can be found occasionally in other small B-cell lymphomas. However, those cells are variably described, and detailed cytologic characterization is often lacking. We identified lymphoma cases with numerous basophilic VLs among the large group of splenic lymphoma with VLs, and for further delineation, 37 cases with this particular cytology were analyzed. Patients, predominantly older men, presented with moderate lymphocytosis and splenomegaly without pancytopenia. The monoclonal B cells expressed IgM + D, IgM + G, IgM or IgG, as well as CD76 and CD11c, frequently CD103, and rarely CD123. Spleen sections were peculiar, with atrophic white pulp and a monomorphic diffuse lymphoma infiltration in a congested red pulp. Bone marrow infiltration was interstitial and intrasinusoidal without extensive fibrosis. Cytogenetic analysis showed a frequent absence of clonal aberrations (68%). Most cases (79%) were IgH mutated, with an overrepresentation of VH3 and VH4 gene families. These results, as well as the clinical evolution, show that those lymphoma cases represent a homogeneous group distinct from SMZL and reminiscent of hairy cell leukemia variant, perhaps corresponding to a separate lymphoma entity.

We retrospectively analyzed 77 patients with pathologically diagnosed angioimmunoblastic T-cell lymphoma from a single city. There were 43 men and 34 women; the median age was 64.5 years (range, 30-91 yr). Average time between first symptoms of the disease and diagnosis was 3.6 months. At diagnosis, peripheral nodes were present in all but 1 patient, and were generalized in 90% of cases. Constitutional symptoms were reported in 77% of cases and spleen enlargement in 51%. A cutaneous eruption--morbilliform, urticarial, or more polymorphic--was present in 45% of patients; in one-third of them, the eruption occurred after drug administration. Other clinical manifestations included pleuritis (22%); arthralgia or arthritis (17%); ear, nose, and throat involvement (14%); central or peripheral neurologic manifestations (10%); and ascites (5%). Most patients presented with advanced disease at diagnosis (bone marrow involvement in 60% of cases). The main laboratory abnormalities were elevated lactate dehydrogenase levels (71%), inflammatory syndrome (67%), hypergammaglobulinemia (50%), anemia (51%), and lymphopenia (52%). Auto- or disimmune manifestations were reported in one-third of patients: autoimmune hemolytic anemia was present at diagnosis in 19% of patients and thrombocytopenic purpura in 7%. Documented vasculitis was described in 12% of cases. Clonality was analyzed in lymph nodes in 47 patients: T-cell and B-cell clones were found in 45 (96%) and 20 (45%) patients, respectively. Chromosomal abnormalities were identified in 62% of cases: trisomies 3, 5, 18, 19, additional X chromosome, and deletion of chromosome 7 were the most common abnormalities. The current study underlines the diversity of presenting manifestations of angioimmunoblastic T-cell lymphoma.

Purpose Some diffuse large B-cell lymphomas (DLBCL) present at diagnosis with associated morphologic features of small B-cell non-Hodgkin's lymphoma (NHL) and may arise from the transformation of a previously unknown indolent low-grade lymphoma. The characteristics and prognosis of these particular DLBCL are not well known. Patients and Methods The strict morphologic review of consecutive DLBCL patients diagnosed over 12 years in our department (Hematology Department, Centre Hospitalier Lyon-Sud, Lyon, France) allowed to retrieve 60 DLBCL that could be have occurred from the transformation of marginal zone B-cell NHL (32 patients), follicular NHL (22 patients), and small lymphocytic NHL (6 patients). We compared them to 180 matched patients of de novo DLBCL. Results Patients median age was 55 years and presented the following clinical characteristics: poor performance status in 33%, disseminated disease in 97%, more than one extranodal site in 50%, and increased lactate dehydrogenase level in 55%. Complete remission with multidrug chemotherapy regimens was achieved in 60% of the patients, but 48% relapsed: 28% with aggressive and 20% with indolent histology, respectively. Overall survival (OS) and freedom-from-progression rates at 5 years were 57% and 33%, respectively. The matched-control analysis showed that patients with transformed NHL at diagnosis had lower complete response to chemotherapy (P = .004) and higher progression rate (P = .03), whereas no difference was observed in OS (P = .21). Conclusion Compared to de novo DLBCL, transformed NHL at diagnosis have similar overall survival but lower complete response to initial treatment and higher risk of indolent relapses.

Contains fulltext : 50323.pdf (Publisher’s version ) (Closed access) The molecular pathogenesis of mantle cell lymphomas (MCL), a subset of B-cell non-Hodgkin's lymphomas with a poor prognosis, is still poorly understood. In addition to the characteristic primary genetic alteration t(11;14)(q13;q32), several further genetic changes are present in most cases. One of the most frequent genomic imbalances is the deletion of 1p22.1-p31.1 observed in nearly one-third of MCL cases. This might indicate the presence of tumor suppressor gene(s) in this critical region of deletion. Quantitative microsatellite analysis (QuMA) is a real-time PCR-based method to detect DNA copy number changes. Since QuMA has the resolving power to detect subtle genomic alterations, including homozygous deletions, this may help to identify candidate tumor suppressor genes from deleted regions. To gain more insight into the molecular pathogenesis of MCL, QuMA was performed on genomic DNA from 57 MCL cases. Eight microsatellite loci mapping to the chromosomal region 1p22.3 were analyzed. Losses were observed in 51 of the 57 ( approximately 89.5%) samples. Two cases showed a homozygous deletion at the locus containing the gene SH3GLB1, which plays a key role in Bax-mediated apoptosis. Two hotspots with copy number losses were detected at chromosomal localizations 85.4 and 86.6 Mb encompassing BCL10 and CLCA2. Both the genes seem to be attractive candidates to study tumor suppressor function in MCL.

The immunological profile of lymphoproliferative disorders is usually conserved whatever the involved site, thus allowing a reliable diagnosis from peripheral blood analysis, especially in small lymphocytic lymphoma/chronic lymphocytic lymphoma (SLL/CLL). Here we present a case wherein the cytology and immunophenotype of blood specimen and bone marrow argue in favor of SLL/CLL with a typical Matutes score (5/5), whereas the cyto-histology and immunophenotype of spleen specimen led to the diagnosis of splenic marginal zone B-cell lymphoma (SMZL). Moreover genomic analysis showed that the splenic cells displayed a SMZL signature. Whereas these data suggested the presence of 2 B-cell clones, the study of the mutational status of IgVH gene in blood and spleen demonstrated the presence of a single clone, which likely developed simultaneously along two distinct ways of differentiation according to the anatomic site suggesting here the predominant role of a micro-environmental factor in cell differentiation. Although rare, this kind of event must be kept in mind as a cause of discrepancies between diagnoses from different sites.

Non-germinal center small B-cell lymphomas represent a heterogeneous group of non-Hodgkin lymphomas, the most frequent histologic subtypes being small lymphocytic lymphoma (SLL), splenic marginal zone B-cell lymphoma (MZL), and mantle cell lymphoma (MCL). In order to identify genomic signatures specific for each disease, we analyzed 128 primary tumors using high-density microarrays. Several clusters of genes significantly discriminated the 3 histologic subtypes. Genes associated with cell adhesion, angiogenesis, and inhibition of apoptosis were up-regulated in SLL. Genes associated with intracellular signaling via the AKT1 pathway were up-regulated in splenic MZL. Genes associated with cell cycle control and multidrug resistance were up-regulated in MCL. Using 44 genes selected within the gene clusters discriminant for the 3 lymphoma subtypes, we generated a class prediction score that allowed us to classify the 3 entities in 96% of the cases, including borderline cases. Whereas specific transcriptional profiles easily distinguished all MZL samples, SLL samples, and most of the MCL samples into separate groups, few MCL cases exhibited MZL-type transcriptional profiles. This study demonstrates that SLL, splenic MZL, and MCL possess specific transcriptional profiles that may be relevant to the pathogenesis and the diagnosis of these histologic subtypes.

Summary In the World Health Organization classification system, splenic marginal-zone lymphoma (splenic MZL) is described as an indolent B-cell lymphoma, which generally presents as splenomegaly with involvement of the bone marrow and peripheral blood. Presence of disease in peripheral lymph nodes and extranodal locations is uncommon. Splenic MZL is characterised by micronodular infiltration of the spleen with marginal-zone differentiation; the immunophenotype is usually lgM+ lgD+/− cytoplasmic-lg-/+ pan B antigens+ CD5- CD10- CD23- CD43-/+ cyclin D1-; and the most common genetic abnormalities are deletions at 7q22-7q32. Most patients with splenic MZL live for a long time but classic prognostic factors cannot distinguish between patients who are likely to have good and poor outcomes. However, immuno-logical events, such as haemolytic anaemia and immune thrombocytopenia, or the presence of a monoclonal component, are significantly associated with shorter survival. Splenectomy is considered the first-line treatment of choice for splenic MZL; it results in only partial remission, but responses are generally sufficient for correcting cytopenia, improving quality of life, and increasing survival.

Splenic marginal zone B-cell lymphoma (MZL), with or without villous lymphocytes, is an indolent lymphoma, presenting with massive splenomegaly, generally associated with bone marrow dissemination. In patients requiring therapy, splenectomy has been reported as the treatment of choice. We reviewed the cases of 81 patients with splenic MZL. Patients presented with stage IV disease at diagnosis in 95% of the cases. Autoimmune events (hemolytic anemia, immune thrombocytopenia, acquired coagulation disorders, positive Coomb's test) were observed in 16 patients, and a monoclonal (M) serum component was detected in 46% of the patients. Twenty patients did not receive any initial treatment at diagnosis. Splenectomy was proposed in 79% of the treated patients, with adjuvant chemotherapy in 47% of patients. Median survival was 10.5 years and was significantly shorter in the presence of an M component, an elevated b2-microglobulin level, leukocyte count > 20000/microL, and lymphocytes > 9000/microL. Disease progression was significantly more frequent in patients presenting an immunological event or an M component. Seventy percent of the patients had persistent involvement of bone marrow and/or peripheral blood after splenectomy. Disease progression was significantly more frequent in partial responders than in complete responders (P < 0.005), but overall survival, risk of histologic transformation, and risk of death from lymphoma were not different in the 2 groups. Moreover, patients with cytopenia at diagnosis treated by splenectomy alone rapidly recovered normal hematological parameters. We conclude that splenectomy is an efficient treatment for splenic MZL, but that it may be delayed until the occurrence of symptoms or cytopenia.

Marginal zone B-cell lymphoma (MZL) is a recently individualized lymphoma that encompasses mucosa-associated lymphoid tissue (MALT) lymphoma, splenic lymphoma with or without villous lymphocytes, and nodal lymphoma with or without monocytoid B-cells. If the clinical description and outcome of MALT lymphoma is well known, this is not the case for the other subtypes. We reviewed 124 patients presenting non-MALT MZL treated in our department to describe the morphologic and clinical presentation and the outcome of these lymphomas. Four clinical subtypes were observed: splenic, 59 patients; nodal, 37 patients; disseminated (splenic and nodal), 20 patients; and leukemic (not splenic nor nodal), 8 patients. These lymphomas were usually CD5-, CD10-, CD23-, and CD43-, but the detection of one or, rarely, two of these antigens may be observed. Bone marrow and blood infiltrations were frequent, except in the nodal subtype, but these locations were not associated with a poorer outcome. Splenic and leukemic subtypes were associated with a median time to progression (TTP) longer than 5 years, even in the absence of treatment or of complete response to therapy. Nodal and disseminated subtypes were associated with a median TTP of 1 year. However, in all these subtypes, survival was good with a median survival of 9 years, allowing these lymphomas to be classified as indolent. Because of the retrospective nature of this analysis, no conclusion may be drawn on the therapeutic aspects, but conservative treatments seem recommended for leukemic and splenic subtypes.

Mantle cell lymphoma (MCL) patients represent a difficult prob-lem, sometimes to establish the diagnosis but mostly becauseof their refractoriness to standard lymphoma treatments. Whichtreatments to apply and to whom is not yet defined. In thisstudy, we attempted to analyze the clinical features, to identifythe major prognostic factors, and to evaluate the outcome of121 MCL patients treated in our institution between 1979 and1997. Clinical data, treatment modalities, and InternationalPrognostic Index (IPI) score were evaluated. Median age was 63years. Patients usually presented with advanced stage disease(87%), disseminated lymph nodes (57%), bone marrow involve-ment (79%), but with a good performance status (PS) (81%).Lymphocytosis .4000/ml and/or peripheral blood involvementwas present in 36% of cases, and gastrointestinal disease in18%. The t(11;14)(q13;q32) and/or bcl-1 rearrangement wasdetected in 47/57 studied cases. Median overall survival (OS)was 3.12 years and a longer survival was significantly associa-ted with younger age (,70 years), good PS (,2), localized dis-ease (stage I–II), fewer than two extra-nodal sites, absence ofspleen or peripheral blood involvement, normal serum LDH andb2-microglobulin levels, and hemoglobin level greater than12 g/dl. However, the IPI failed to identify patients with longerOS and in a multiparametric analysis, only older age, hemoglo-bin less than 12 g/dl, poor PS, and blood involvement wereassociated with a poorer outcome. Treatment modalities hadno impact on survival with 75% of patients relapsing or pro-gressing. Our data showed that the poor outcome of MCLpatients is mainly related to adverse patient characteristics, ahighly disseminated tumor, and some unknown parametersassociated with the refractoriness to standard therapy.Keywords: mantle cell lymphoma; prognostic factors; outcome

Introduction Among recently discovered B cell activators responsible for signaling events leading to B-cell activation and maturation, of particular interest are the Toll-like receptors (TLRs). TLR expression is heterogeneous and variable among B-cells. In addition, abnormal TLR levels/signaling may play an important role in the pathogenesis of lymphoma, particularly in splenic marginal zone lymphoma (SMZL), since TLR pathways are recurrently targeted by genetic changes in this lymphoma. Methods Frozen spleen tissue specimens from patients with SMZL (n=13) and splenic diffuse red pulp lymphoma with villous lymphocytes (SDRPL, n=5) were analyzed for the expression of TLR1 to TLR10 in comparison to control cases (traumatic spleen, n=7) using multi-parametric flow cytometry and quantitative mRNA Taqman assay. To identify the B-cell subset, the samples were also stained with anti-CD19 and anti-CD3. All cases were studied by morphological, immunological, cytogenetic and molecular analysis. Results The TLR profile obtained at protein level by flow cytometry was closely related to that obtained at mRNA level. The SMZL/SDRPL B-cells expressed all TLRs, but with variable levels of protein expression (low for TLR1, TLR2, TLR3, TLR8, TLR9, TLR10, and high for TLR4, TLR5, TLR6 and TLR7). On the other hand, distinct TLRs profiles were observed according lymphoma subtypes. The SDRPL cases showed indeed a significant TLR2 under-expression and TLR4/TLR7 over-expression in comparison to SMZL cases and control B-cells. SMZL cases exhibited a significant TLR4/TLR8 over-expression in comparison to control B-cells. These TLR profiles were not associated with specific cytogenetic features (presence of del7q or trisomy 3) and/or immunological profile (expression of the CD11c/CD27/isotype of immunoglobulin). But, in both entities, TLR4 expression was higher in cases with mutated IGHV than in cases with unmutated IGHV. The overexpression of TLR7 MyD88-dependent signaling molecules has been reported as pathogenic mechanism for autoimmune diseases; however no more autoimmune disease or circulating auto-antibodies were associated with SDRPL cases as compared to SMZL cases. As previously reported, all cases but one SMZL case presented unmutated MyD88 (L265P mutation), and MyD88 protein evaluated here by flow cytometry was similarly expressed in both entities. While TLR7 stimulation is known to induce CD38 expression, all SDRPL cases were CD38 negative; while in SMZL cases, higher TLR7 expression was observed in CD38 positive as compared to CD38-negative cases. This may suggest the existence of an abnormal TLR7 pathway in SDRPL as opposed to SMZL. Conclusion TLR profiling should allow a better understanding of the mechanisms involved in SMZL/SDRPL pathogenesis. Present data suggest an abnormal TLR pathway involving NF-kB and/or MyD88 in the SDRPL entities. Disclosures: No relevant conflicts of interest to declare.

Abstract 1594 Background: Follicular Lymphoma (FL) is the most frequent low-grade NHL. Clinical course is heterogeneous, some patients (pts) presenting an indolent clinical course with overall survival (OS) over 15 y and others developing a more aggressive disease with shorter survival. The Follicular Lymphoma International Prognostic Index (FLIPI) or FLIPI-2 are commonly used to predict pts outcome, but fail to identify pts with a really poor prognosis. At diagnosis, few FL pts present with detectable leukemic phase (FL-LP) and this characteristic has been seldom described. Aim: The aim of the study was to describe the clinical features and outcome of FL-LP pts. Results: Among 499 pts diagnosed with FL according to the WHO criteria in the Centre Hospitalier Lyon Sud (transformation and grade 3b excluded) and treated between 01/1992 and 01/2012, 37 (7.4%) had characteristic FL-LP detected by cytological blood smears analysis with confirmation by flow cytometry (kappa/lambda clonality). Median age was 58 y and FLIPI score repartition was 4 pts in low, 16 in intermediate, and 17 in high risk groups. Splenomegaly was present in 23 pts, high tumour burden (GELF criteria) in 11, B symptoms in 8, and ECOG PS>1 in 3. Seven pts had anaemia, 17 platelets UNL, and 11 LDH >UNL. The circulating lymphoma cells expressed the CD10 in 29/37 cases and surface Ig expression was detected in 31/35 cases, mainly IgM or IgG isotype. The median count of circulating lymphoma cells was 1.95.109/L, ranging from 0.6 to 129.109/L, 15 pts having count >4.109/L and 6 pts >10.109/L. Cytogenetic data were available for 21 pts, 20 carried the t(14;18), or its variant t(18;22), 16 of them having complex karyotype. Two pts were on watchful waiting for 24 and 82 m and 35 received a chemotherapy regimen at diagnosis including rituximab in 27 cases. Overall response rate was 83% (29/35) with 23 CR/CRu. Median progression-free survival (PFS) was 29 m. PFS and OS estimates were 37% and 86% at 5 y and 31% and 68% at 10 y, respectively. Splenomegaly (P=.035), high tumour burden (P=.017), lymphoma cells count greater than 4.109/L (P=.028), β2-m>UNL (P=.036), and thrombocytopenia (4.109/L (HR 5.92; P=.000916) as independent prognostic factors. After progression, 12 pts received high-dose therapy (HDT) with HSCT as salvage with a long second PFS (68% at 10 y) compared to 8.3 m median PFS in first line. Pts not receiving HDT at salvage had a median second PFS of 27 m. To further evaluate the impact of FL-LP on pts outcome, a 1:3 matched analysis was performed. The FL-LP 37 pts were successfully matched with 111 newly diagnosed FL without FL-LP according to FLIPI score, age, treatment type (abstention vs chemotherapy, with or without rituximab) and treatment period (before or after 2000). In these 111 matched pts, 5- and 10-y OS was 97% and 91%, respectively. Considering all 148 pts, high FLIPI score, presence of FL-LP, and β2-m>UNL were all significantly associated with a worse PFS. In a Cox regression model for PFS (120/148 pts with complete data), high FLIPI score (P=.0034; HR=2) and presence of FL-LP (P=.0085; HR=2.2) remained independently associated with shorter PFS. High FLIPI score and presence of FL-LP were also associated with a shorter OS. Interestingly pts with less than 4.109/L of circulating lymphoma cells had a similar PFS than those without. When circulating lymphoma cells >4.109/L as variable (see abstract figure) instead of FL-LP were tested in the Cox regression model for PFS including β2-m>UNL and FLIPI score as variables, the most significant predictor for a shorter PFS was circulating lymphoma cells >4.109/L (P=.0004; HR=3.56) as compared to FLIPI score (P=.051; HR=1.6) and β2-m (P=.09; HR=1.52). Conclusion: The presence of circulating lymphoma cells in FL is a rare event and is associated with shorter PFS independently of FLIPI score and β2-m level. A validation of these findings on pts from the PRIMA study is on progress. However, this population is not homogenous and pts with circulating lymphoma cells >4.109/L have a poorer outcome. Although ∼1/3 of the pts experience long term PFS, these pts should be monitored carefully during and after first line treatment to consider HSCT as a therapeutic option to achieve a sustained response. Disclosures: No relevant conflicts of interest to declare.

Within the group of indolent B cells lymphomas derived from cells of the marginal zone, the pathogenesis of Splenic Marginal Zone Lymphoma (SMZL) remains incompletely characterized, in contrast to MALT lymphoma. However numerous findings recently contributed to a better description of this lymphoma, including its possible association with hepatitis C and the recurrence of 7q deletion. Furthermore, the peculiar pattern of Ig genes mutations and the biased usage of some segments (IGHV1-2) is suggestive of a T-independent Ag driven proliferation, at least at initial steps. The clinical heterogeneity is also marked with a lack of reproducible prognostic factors able to characterize the cases with a more aggressive course or histological transformation. MicroRNA (miRNA) are small non-coding RNA that have a post transcriptional regulation role by inhibiting translation of mRNA in protein and their abnormal expression was already found to contribute to the oncogenesis of various leukemias and lymphomas. The aim of the study was then to identify a miRNA profile of SMZL and to assess if specific miRNAs were associated with biological or clinical characteristics. MiRNA expression profile of SMZL was obtained by quantitative RT-PCR technology (Taqman microRNA Assays Human Panel, Applied Biosystems) which enabled to analyze 365 miRNAs. Six frozen spleen specimens of SMZL (including 2 cases with histological transformation) and 5 frozen specimens of non tumoral spleen (traumatic) were used to compare miRNA levels of expression (2−□□Ct method). Out of these 365 miRNAs, 95 were found to be reproducibly detectable in those samples and were further analyzed. Three miRNA were uniformly overexpressed in SMZL samples as compared to non tumor samples: miR-155 (mean fold change = 3.1), miR-451 (mean = 2.37), miR-486 (mean = 2.7). Conversly, 4 miRNAs were uniformly underexpressed: miR-127 (mean = 0.32), miR-139 (mean = 0.32), miR-335 (mean = 0.26), miR-411 (mean = 0.25). Interestingly, 1 miRNA, miR-21 was specifically found overexpressed only in the 2 transformed cases of SMZL (mean = 3.49) in comparison with the 4 non transformed cases (mean = 0.94). Those results were confirmed in a larger cohort of SMZL patients using a simplex real time PCR (Taqman microRNA Assays Human, Applied Biosystems) on frozen or paraffin embedded samples with a specific overexpression of miR-21 in 10 transformed cases (mean = 14.4), absent in the 6 non transformed cases (mean = 0.9) (Mann Whitney test: p =.0022). MiR-155 plays a key role in B cell differentiation and was already found overexpressed in other lymphomas subtypes. Little is know about the other miR overexpressed except for MiR- 21. MiR-21 is commonly overexpressed in solid tumors and the level of its expression is correlated in tumor growth and metastatic dissemination. It was also found overexpressed in chronic lymphocytic leukaemia and DLBCL (diffuse large B cell lymphoma). In opposite to DLBCL, our results seem to link miR-21’s overexpression with aggressive forms of SMZL. MiR-21 regulates multiple genes associated with apoptosis, migration and invasiveness. Altogether, these data points towards a specific miRNA expression pattern in SMZL and suggests that miR-21 could regulate important oncogenic pathways in transformed SMZL lymphomas.

Chromosomal translocations involving the MYC oncogene (8q24) are known to occur in Burkitt lymphomas/leukemias (BL) but also in 5–30% of diffuse large B-cell lymphomas which are usually highly aggressive (8q24 DLBCL). Two recent conventional cytogenetics (CC) studies showed that secondary chromosomal abnormalities have a negative prognostic impact, especially13q abnormalities (frequently described as a deletion and usually associated with a complex karyotype, i.e. > 3 chromosomal alterations), mainly in childhood mature B-cell lymphomas, and in a less extent 7q, 3q, 22q and chromosome 17. M-FISH was applied to 120 (74% adults and 26% children) high grade B-cell non-Hodgkin lymphomas (86% BL, 14% 8q24 DLBCL) carrying MYC rearrangement and complex karyotype and/or 13q abnormality in order to find recurrent and/or cryptic chromosomal alterations. Abnormal metaphases were available in 96 (80%) cases. We described ‘new’ (not seen in CC) chromosomal rearrangements in 50 (52%) cases, refined those seen by CC in 28 (29%) cases and confirmed CC abnormalities in 18 (19%) cases. M-FISH allowed to characterize 55 structural 13q abnormalities in 42 patients (21 ‘new’ alterations): 24 der(13q) leading to partial del(13q) and partial gain of different partner chromosomes [mainly 1q or 7q], 15 del(13q) [of 2 minimal regions : q14 & q31q34], 13 gains (only 2 seen by CC) and 3 balanced translocations. Combined results of CC and M-FISH showed that the most frequent abnormalities among patients with complex karyotype involved 1q, 7q, Xq, 3q, 18q, 6q, 17p and chromosome 22 (excluding 8q24 translocations). 79 1q abnormalities were detected in 54 patients (26 % ‘new’): mostly gains (minimal amplified region: q22q31) due to unbalanced translocations with chromosomes 13, 22, 7 (59%) or duplications (25%). 55 partial 7q gains were observed in 42 patients (22% ‘new’), mostly +7 or unbalanced translocations with 13q, 6q or 1q. The other most common abnormalities were: t(14;18) in 8q24 DLBCL, del(6q), der(3q) with various partners leading to partial loss of 3q, monosomy 17 or del(17p) and numerical changes of chromosomes 22 (monosomy) and X. In conclusion, this study confirms the contribution of M-FISH in refining CC results in highly aggressive 8q24 B-cell lymphomas: ‘new’ rearrangements were identified especially in 1q (leading to partial 1q gains), 18q, 6q (leading to partial 6q deletions), 13q ; partial 13q gains were underestimated by CC and both 13q deletions and gains were more heterogeneous than expected. We are characterizing these prognostic additional chromosomal abnormalities with SNP-CHIPS 50K array (Affymetrix) to look for candidate genes and/or cellular pathways involved in Burkitt lymphomagenesis in cooperation with oncogenic effect of MYC. °on behalf of the GFCH (Groupe Francophone de Cytogénétique Hématologique) and the BCG-Ho (Belgian Cytogenetic Group of Hematology and Oncology).

Glutathione-S-transferase π (GSTπ), whose gene is located in 11q13, is belonging to a complex multigene family of enzymes that may detoxify electrophilic xenobiotics. Recent gene expression profiling analysis showed that GSTπ transcript is belonging to the mantle cell lymphoma (MCL) signature and constitutes one of the most expressed mRNA in MCL[*][1]. Furthermore it has been reported by immunohistochemistry that overexpression of cyclin D1 in MCL was associated with a high level of GSTπ protein expression. We then looked for the specificity of a high level of GSTπ protein expression in MCL, and particularly when comparing to marginal zone lymphoma (MZL) with t(11;14). Material and Methods: Immunochemistry analysis of GSTπ and cyclinD1 expression was performed 111 non Hodgkin’s lymphoma including MCL, MZL, small lymphocytic lymphoma (SLL), follicular lymphoma (FL), and diffuse large B-cell lymphoma (DLBCL) cases with or without t(11;14). Results: A strong GSTπ expression in more than 50% of the cells was observed in 98% of the MCL. None of the MZL cases with t(11;14) expressed GSTπ, except one case that expressed a weak signal in less than 20% of the cells. None of the MZL without t(11;14) expressed GSTpi. Only two (13%) FL graded 3 according to OMS classification expressed GSTπ in less than 50% of the cells. A high GSTπ expression (>50% cells stained) was detected in 29% of the DLBCL and 43% of the SLL. | | n | GST π expression >50% | GST π expression 5–50% | π GST expression

Splenic Marginal Zone Lymphoma (MZL) is described as an indolent lymphoma with a long term survival. However classical prognostic factors can not distinguish between patients who are likely to have good or poor outcomes. Moreover histological progression occurs in 10–20% of cases at the time of recurrence, but may be present at diagnosis, inducing a shorter survival with a median around 2 years. New model based on molecular understanding are needed to better discriminate these patients for their prognosis and disease evolution. Biopsy samples of splenic MZL from 43 patients treated in one institution and 8 with transformed splenic MZL were examined for gene expression using a nylon cDNA microarray consisting of 7, 000 human genes. Two of them came from a group of matched pair of splenic MZL and the transformed counterpart. An additional group comprising 4 transformed follicular lymphomas and 1 transformed small lymphocytic lymphoma were also analysed. Hierarchical clustering realized with all samples allowed to visualized patterns of gene expression already described in our previous work1 and corresponding to precise functions (proliferation, early response...), cell or tissue subtype (T cells, stroma …) or lymphoma subtype (MZL, MCL, SLL). The ability of individual genes to distinguish 1/ MZL and transformed-MZL subtypes, and 2/ alive patients and dead patients was calculated using a supervised method (discriminating score and bootstrap resampling). This allowed us to construct molecular predictors of survival and histological transformation. Transformation discriminating genes regrouped 73 genes related to cell proliferation such as CDC10, CDK4, PRKDC, PLCG2, LDHA, MCL2, and NPM1. P53 was down-expressed in all the transformed samples. Survival discriminating genes regrouped 26 genes from a unique cluster related to cell proliferation such as LDHA, NME1, MYC, ENO1. Although these genes seem to participate to the same function, only one gene was found discriminating the 2 parameters (LDHA). Gene ontology analysis using GOminer showed that the histological transformation was more related to cyclin-dependent protein kinase and that the survival was more related to metabolism. We used 26 genes to construct a predictor for survival in MZL patients. This gene-based predictor allowed predicting survival (p