Light Chain IgLV3-21 Is Associated with Poor Prognosis in Chronic Lymphocytic Leukemia Patients Independently of the Presence of Heavy Chain IgHV3-21

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3218-3218
Author(s):  
Basile Stamatopoulos ◽  
Thomas Smith ◽  
David Sims ◽  
Andreas Heger ◽  
Hélène Dreau ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heavy Chain ◽  
Light Chain ◽  
Poor Prognosis ◽  
Mrna Translation ◽  
Lymphocytic Leukemia ◽  
The Other ◽  
Mutational Status ◽  
Initial Cohort ◽  
Independent Cohort

Abstract Background : Chronic Lymphocytic Leukemia (CLL) is characterized by a very heterogeneous clinical course and the immunoglobulin heavy-chain gene (IgHV) mutational status is currently considered the gold standard of prognostication: unmutated (UM) immunoglobulin heavy chain region (IgHV) is associated with a poor prognosis while patients with mutated IgHV (M) have more indolent disease. An exception are patients with IgHV3-21/IgLV3-21 who have poor prognosis irrespectively of the IgHV mutational status. Interestingly, IgHV3-21 is co-expressed with IgLV3-21 in the majority of cases. However, little is known about IgLV3-21: indeed this light chain has never been characterized independently of IgHV3-21 in terms of gene expression and prognostic impact. Methods: Based on a cohort of 32 patients with aggressive CLL, we used total RNAseq data of highly purified leukemic cells to define gene expression profiles and IgHV/IgLV rearrangements for each patient. Gene set enrichment analysis was correlated with treatment-free (TFS) and overall (OS) in the initial cohort of 32 patients and in an independent cohort of 255 patients where IgLV3-21 positivity was determined by real-time PCR and confirmed by Sanger sequencing. Results: Among the 32 initial CLL patients, 9 patients had an IgLV3-21 rearrangement, but only 1 patient carried the IgHV3-21 rearrangement. The other patients had VH1-24/69, VH3-9/66/23/48/53 and VH4-59 heavy chain rearrangements. RNAseq expression profiling yielded 1457 transcripts and 789 genes that were differentially expressed between IgLV3-21 patients and the other patients. Within the differentially expressed genes, 68% were upregulated while 32% were downregulated at least 1.5 fold. Gene set analysis revealed enrichment of genes related to translation enhancement (ribosome, translational reactome, peptide chain elongation, RNA metabolism - P<0.0001) and MYC target genes (P=0.0003), in line with recent finding showing that BCR stimulation can increase global mRNA translation including MYC-specific mRNA translation. In the initial cohort of 32 patients, IgLV3-21 patients had a median TFS of 17 months compared to 44 months in patients with another light chain (P=0.0270). Similarly, IgLV3-21 patients had a shorter median OS (88 months vs >192 months, P=0.0287).We validated these results in an independent cohort of 255 patients with 31 (12%) IgLV3-21 patients and 10 (4%) with IgHV3-21 (of which 8/10 also carried the light chain IgLV3-21 rearrangement). IgLV3-21 patients presented a median TFS/OS of 29/183 months compared to non IgLV3-21 patients who had a median TFS/OS of 88/292 months (P=0.0003/P=0.0142). In addition, when IgHV3-21 patients (n=10) were compared to IgLV3-21 only patients (n=23), no statistical difference was observed in terms of TFS or OS. Interestingly, if patients were classified according the IgHV mutational status, both IgHV3-21 and IgLV3-21 patients displayed a prognosis similar to UM patients: median TFS was 144, 32, 23, 48 months for M, UM, IgHV3-21 and IgLV3-21 patients, respectively (Figure A- P<0.0001). Similar results were observed for OS with a median OS of 292, 112, 128 and 241 months for M, UM, IgHV3-21 and IgLV3-21 patients, respectively (Figure B - P<0.0001). If all IgLV3-21 (n=31) were considered independently of their heavy chain, median TFS (29 months) were similar to UM patients (32 months, P=0.5536) and statistically different from M patients (144 months - P<0.0001, Figure C). Similar results were observed for OS (Figure D). Conclusions: Our results highlight for the first time the importance of light chain IgLV3-21 in CLL in terms of its differential gene expression profile and prognosis: IgLV3-21 is associated with a translational enhancement gene signature and confers a poor prognosis similar to UM patient irrespectively of the heavy chain IgHV3-21 or the IgHV mutational status. Figure Figure. Disclosures Schuh: Gilead: Consultancy, Honoraria, Research Funding; Roche, Janssen, Novartis, Celgene, Abbvie: Consultancy, Honoraria.

Analysis with the LightCycler System® Identifies a Highly Significant Correlation between ZAP-70 mRNA Expression and Immunoglobulin Variable Heavy Chain Gene Mutational Status in Chronic Lymphocytic Leukemia.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3259-3259
Author(s):  
Frank Dicker ◽  
Christine Ebenbichler ◽  
Karin Mann ◽  
Annemarie Linder ◽  
Susanne Schnittger ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Mrna Expression ◽  
Heavy Chain ◽  
Poor Prognosis ◽  
Somatic Mutations ◽  
Lymphocytic Leukemia ◽  
Mutational Status ◽  
Variable Heavy

Abstract The presence or absence of somatic mutations in the immunoglobulin variable heavy chain (IgVH) region of chronic lymphocytic leukemia (CLL) B cells has prognostic information already at the time of diagnosis. The lack of somatic mutations in about 50% of the CLL patients significantly separates those with poor prognosis from those with a more indolent course that carry somatic mutations in their IgVH gene. However, the analysis of this marker is time consuming. Global gene expression analysis of CLL cells identified the expression of the T cells marker zeta chain associated protein (ZAP-70) as a surrogate marker for an unmutated IgVH. In order to analyze ZAP-70 expression in CLL cells using a relative quantification approach, we developed an assay* on the LightCycler® instrument. Here, we analyzed a cohort of 100 samples for ZAP-70 mRNA and compared the results with the IgVH mutational status. Mononuclear cells were isolated from blood (n=80) or from bone marrow (n=20). To exclude ZAP-70-expressing T and natural killer cells from analysis, cells were enriched for B cells to a purity of &gt; 99% by CD19 magnetic beads positive selection and mRNA was isolated using a MagNA Pure LC instrument. cDNA was reverse transcribed from an equivalent of 5x 106 cells and used for the analysis of the IgVH status as described (Blood. 2003;101:2049–2053) as well as for the analysis of ZAP-70 expression with a LightCycler® instrument. ZAP-70 expression levels as the target gene were normalized to the houskeeping gene ABL using relative quantitation. mRNA of the Jurkat cell line was used as a calibrator. In preliminary experiments, ABL was selected from a total of 4 different housekeeping genes based on its expression levels, which were in the range of ZAP-70 expression, and based on the resulting correlation to the IgVH status. Thus, results for ZAP-70 gene expression are given as normalized ratio. ZAP-70 mRNA expression (n=100) revealed a highly significant correlation to an unmutated IgVH and low ZAP-70 expression correlated to a mutated IgVH (r2=0,692, Spearman). A cut off of 0.5 ZAP-70 expression was optimal to distinguish samples with mutated from unmutated IgVH, resulting in 80 concordant samples (80%). Of the 11 mutated IgVHs with discordant ZAP-70 expression 7 (64%) had borderline mutation status and 4 (36%) expressed the poor prognosis VH3-21 gene. 7(78%) of the 9 unmutated IgVHs with low ZAP-70 expression were characterized by adverse cytogenetic features. In summary, the results of the present study show that the LightCycler® system based quantitative RT-PCR is suitable to achieve a highly significant correlation between the IgVH status and ZAP-70 expression in CLL samples. Unlike the flow cytometric detection of ZAP-70 protein expression, which uses ZAP-70 of endogenous T cells as an internal control, the assay* developed by us might be easier to standardize between laboratories and therefore, might be applicable for future routine applications. * For research use only. Not available in distribution.

Angiopoietin-2 Expression in B-Cell Chronic Lymphocytic Leukemia: Association with Clinical Outcome and Immunoglobulin Heavy-Chain Mutational Status.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2780-2780
Author(s):  
Rossana Maffei ◽  
Silvia Martinelli ◽  
Ilaria Castelli ◽  
Rita Santachiara ◽  
Elena Morandi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Chronic Lymphocytic Leukemia ◽  
Clinical Outcome ◽  
Heavy Chain ◽  
Lymphocytic Leukemia ◽  
Adverse Clinical Outcome ◽  
Mutational Status ◽  
Angiopoietin 2 ◽  
Normal Controls ◽  
Cell Chronic Lymphocytic Leukemia

Abstract B-cell Chronic Lymphocytic Leukemia (B-CLL) follows an extremely variable clinical course. For some patients CLL is an indolent disease that never progresses to the point of requiring therapy and these patients have a survival time similar to age-matched controls. On the contrary, other patients experience rapidly deteriorating blood count and organomegaly which requires prompt treatment. The overall survival (OS) times range from months to decades. B-CLL patients can be divided into two subgroups on the basis of the presence or absence of somatic mutations in the specific immunoglobulin heavy-chain variable region (IgVH) genes used by leukemic cells. Patients with unmutated IgVH genes usually have an advanced stage and an unfavourable cytogenetic features, require therapy and have a short survival. The biological reasons of the different behaviour of Ig-mutated and Ig-unmutated leukemic clones have not been fully elucidated yet. Angiogenesis is a very complex network which is closely regulated by the orchestrating functions of many angiogenic factors. The balance of cellular expression of all these angiogenic factors determines vascular stabilization or angiogenic remodelling and sprouting or vessel regression. Increasing evidence shows that neovascularization plays a role in the biology of chronic lymphocytic leukemia. The goal of this study was to evaluate the angiogenic status of Ig-mutated and Ig-unmutated CLL in the attempt to identify a possible role of angiogenesis in the adverse clinical outcome of Ig-unmutated CLL patients. So, we first performed a large scale gene-expression analysis on 29 B-CLL patients using microarrays comprising about 20,000 probes and 208 angiogenesis-related genes. We identified 64 up-regulated genes in Ig-unmutated CLL relative to Ig-mutated CLL. Among them, we found angiopoietin-2 (Ang-2) as one of the highest differentially expressed gene (p=3.02x10−6). Then, we evaluated the Ang-2 expression both at transcript and protein level in a wide cohort of CLL, in normal controls and in other haematological malignancies. The data showed an extremely wide range of Ang-2 expression in B-CLL: Ang-2 high-expressing cases were characterized by advanced Binet stage (p=0.032) and significantly shorter progression-free survival than Ang-2 low-expressing subset (median, 16 vs. 146 months) (p=0.006). Moreover, there was a strong correlation between the IgVH mutational status and the Ang-2 gene expression level (p<0.0001). Ig-mutated CLL exhibited very low Ang-2 expression, absolutely similar to the levels measured in healthy controls (median, 0.27 vs. 0.32, p=0.959). On the contrary, Ig-unmutated CLL expressed up to one hundred-fold higher levels of Ang-2 than normal controls(median, 38.61 vs. 0.32, p=0.002). Moreover, Ang-2 was up-regulated in all investigated CML, ALL and AML samples. We observed extremely high levels of expression in CML and ALL (median Ang-2 mRNA, 3948.96 and 190.00, respectively), whereas moderately high levels of Ang-2 were found in AML patients (median, 16.21). These data suggest that increased angiogenesis due to high Ang-2 expression may be involved in adverse clinical outcome of Ig-unmutated CLL patients.

scholarly journals Relation of Gene Expression Phenotype to Immunoglobulin Mutation Genotype in B Cell Chronic Lymphocytic Leukemia

2001 ◽  
Vol 194 (11) ◽  
pp. 1639-1648 ◽  
Author(s):  
Andreas Rosenwald ◽  
Ash A. Alizadeh ◽  
George Widhopf ◽  
Richard Simon ◽  
R. Eric Davis ◽  
...  
Keyword(s):  
Gene Expression ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Germ Line ◽  
Gene Expression Signature ◽  
Lymphocytic Leukemia ◽  
Common Mechanism ◽  
Human Leukemia ◽  
Mutational Status ◽  
Cell Chronic Lymphocytic Leukemia

The most common human leukemia is B cell chronic lymphocytic leukemia (CLL), a malignancy of mature B cells with a characteristic clinical presentation but a variable clinical course. The rearranged immunoglobulin (Ig) genes of CLL cells may be either germ-line in sequence or somatically mutated. Lack of Ig mutations defined a distinctly worse prognostic group of CLL patients raising the possibility that CLL comprises two distinct diseases. Using genomic-scale gene expression profiling, we show that CLL is characterized by a common gene expression “signature,” irrespective of Ig mutational status, suggesting that CLL cases share a common mechanism of transformation and/or cell of origin. Nonetheless, the expression of hundreds of other genes correlated with the Ig mutational status, including many genes that are modulated in expression during mitogenic B cell receptor signaling. These genes were used to build a CLL subtype predictor that may help in the clinical classification of patients with this disease.

scholarly journals Arrangement of the disulphide bridges in human low-Mr kininogen

1987 ◽  
Vol 247 (1) ◽  
pp. 15-21 ◽  
Author(s):  
J Kellermann ◽  
C Thelen ◽  
F Lottspeich ◽  
A Henschen ◽  
R Vogel ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Heavy Chain ◽  
Light Chain ◽  
The Other ◽  
Critical Importance

The arrangement of the disulphide bridges in human low-Mr kininogen has been elucidated. Low-Mr kininogen contains 18 half-cystine residues forming nine disulphide bridges. The first and the last half-cystine residues of the amino acid sequence form a disulphide loop which spans the heavy- and the light-chain portion of the kininogen molecule. The other 16 half-cystine residues are linked consecutively to form eight loops of 4-20 amino acids; these loops are lined up in the heavy-chain portion of the kininogen molecule. In this way, a particular pattern of disulphide loops is formed which seems to be of critical importance for the inhibitor function of human kininogen.

scholarly journals Phosphorylation of lymphocyte myosin catalyzed in vitro and in intact cells.

1982 ◽  
Vol 93 (2) ◽  
pp. 261-268 ◽  
Author(s):  
M Fechheimer ◽  
J J Cebra
Keyword(s):  
Heavy Chain ◽  
Light Chain ◽  
Lymphocytic Leukemia ◽  
Leukemia Cells ◽  
Intact Cells ◽  
Surface Immunoglobulin ◽  
Cell Extracts ◽  
Lymphocyte Populations ◽  
Immunologic Reactions

Myosin has been isolated from guinea pig B-lymphocytic leukemia cells (L2C). The myosin has been enzymatically phosphorylated and dephosphorylated in vitro using both heterologous and lymphocyte-derived enzymes. Both the heavy chain and 20,000-dalton light chain of lymphocyte myosin are phosphorylated in vitro. Phosphorylation of myosin enhances actin-activated ATPase activity. Phosphorylation of myosin in murine lymphocytes was analyzed by use of a novel technique for rapid immunoprecipitation of myosin from cell extracts. Both the heavy chain and 20,000-dalton light chain of myosin are phosphorylated in intact cells. Addition of antibody reactive with cell-surface immunoglobulin to lymphocyte populations enriched for B cells stimulates locomotion of these cells and also increases the quantity of 32P isolated in association with the 20,000-dalton light chain of lymphocyte myosin, when 32Pi was present in the medium. In addition, an unidentified, phosphorylated polypeptides with a molecular mass of 22,000 daltons is co-isolated with myosin from cells by rapid immunoprecipitation. These results are consistent with the hypothesis that phosphorylation of myosin may contribute to regulation of movements performed by lymphocytes which are related to their participation in immunologic reactions.

Strikingly Homologous Immunoglobulin Gene Rearrangements and Poor Outcome in VH3-21-Utilizing Chronic Lymphocytic Leukemia Independent of Geographical Origin and Mutational Status.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 175-175
Author(s):  
Fiona Murray ◽  
Mia Thorselius ◽  
Alexander Krober ◽  
Ulf Thunberg ◽  
Gerard Tobin ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Light Chain ◽  
Geographical Origin ◽  
Lymphocytic Leukemia ◽  
Gene Rearrangements ◽  
Immunoglobulin Gene ◽  
Mutation Status ◽  
Mutational Status ◽  
Significant Difference ◽  
Vh Genes

Abstract We recently reported that Swedish VH3-21-utilizing chronic lymphocytic leukemia (CLL) patients showed restricted immunoglobulin gene features and poor prognosis despite VH mutation status. To investigate whether VH3-21+ CLLs have similar characteristics in different parts of the world, we analyzed the VH and VL gene rearrangements in 90 patients from Sweden, Germany, Italy, USA, Finland and Australia and correlated these data with survival. Sixty-three percent of cases exhibited mutated VH genes and 37% had unmutated VH genes. Fifty patients (56%) displayed a short and homologous heavy-chain CDR3, many of these with the amino acid motif, DANGMDV. Also, a highly biased Vλ2-14 usage was evident in 73% of patients with a restricted light-chain CDR3, QVWDS(S/G)SDHPWV. Combined restricted heavy- and light-chain CDR3s were found in patients from all included countries. Although VH3-21+ CLLs have a remarkably predominant λ-expression, analyses of kappa deleting element showed a conserved rearrangement order of the light-chain loci. The overall survival was poor in the VH3-21+ cohort (median survival 88 months) with no significant difference in relation to mutation status or homologous/non-homologous CDR3. In summary, highly restricted B-cell receptors and worse outcome characterize VH3-21+ CLLs independent of geographical origin and mutation status. VH3-21 usage should now be included in prognostic stratification of CLL when assessing mutation status.

Serum BAFF (B-Cell Activating Factor of the TNF Family) Compared with Immunoglobulin Heavy-Chain Mutation Status, ZAP-70 and CD38 as a Predictor of Time to First Treatment in Early B-Cell Chronic Lymphocytic Leukemia.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3093-3093
Author(s):  
Stefano Molica ◽  
Gaetano Vitelli ◽  
Giovanna Cutrona ◽  
Giovanna Digiesi ◽  
Rosanna Mirabelli ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Heavy Chain ◽  
Immunoglobulin Heavy Chain ◽  
Serum Levels ◽  
Lymphocytic Leukemia ◽  
B Cell Activating Factor ◽  
Mutational Status ◽  
Time To First Treatment ◽  
Cell Chronic Lymphocytic Leukemia

Abstract We analyzed the correlation between well-established biological parameters of prognostic relevance in B-cell chronic lymphocytic leukemia [CLL] (i.e, mutational status of the immunoglobulin heavy chain variable region [IgVH], ZAP-70- and CD38-expression) and serum levels of BAFF (B-cell activating factor of the TNF family) by evaluating the impact of these variables on the time to first treatment [TFT] in a series of 69 previously untreated Binet stage A B-cell CLL patients. By using a commercial ELISA (R & D Systems, USA) we found that higher levels of BAFF characterized more frequently patients with Rai stage 0 (P=0.008) and mutated IgVH (P=0.03). In contrast, peripheral blood lymphocytosis (P=0.06), serum β2-m (P=0.159), LDH (P=0.333) and percentage of ZAP-70-positive (P=0.242) or CD38-positive B-CLL cells (P=0.142) did not reflect circulating levels of BAFF. The relationship among various bio-pathological parameters, analyzed by the multiple correspondence analysis (MCA), showed two different clinico-biological profiles. The first, characterized by higher BAFF serum levels (i.e., > 336 ng/mL), presence of mutation in the IgVH, low percentage of CD38-positive B-CLL cells (< 30%) and low LDH was associated with a stable pattern of disease generally not requiring therapy. The second, defined by lower BAFF levels, absence of mutation in the IgVH, high percentage of CD38- positive B-CLL cells and high LDH was associated with a more progressive pattern of disease and a shorter TFT. After a median follow-up time of 35 months (range, 2–120 months) 26 (37.6%) out of 69 patients experienced a need for chemotherapy. Kaplan-Meier estimates of patientsTFT, plotted after searching the best cut-off for BAFF (i.e., 336 ng/mL), demonstrated that low BAFF concentration was associated with a shorter TFT (median TFT 36 months) while median was not reached by patients with BAFF levels higher than 336 ng/mL (P<0.0001). Along with lower serum levels of BAFF (Hazard Ratio [HR], 0.19; P<0.0001), the univariate Cox proportional hazard model identified absence of mutation in IgVH (HR, 0.17; P<0.0001), CD38-positivity (HR, 3.32; P=0.01) and lower platelet count (HR, 0.19; P=0.03) as predictor of shorter TFT. Finally, in multivariate analysis only mutational status of IgVH (HR, 0.25; P=0.007) and serum concentration of BAFF (HR, 034; P=0.04) affected significantly TFT. Our results indicate that in early B-cell CLL clinico-biological profile including among other parameters BAFF may provide a useful insight into the complex interrelationship of prognostic variables and semplify their interpretation. The possible presence of BAFF isoform in B-CLL could peraphs account for the unexpected correlation between low soluble BAFF levels and poor clinical outcome in patients with early disease.

Extensive Intraclonal Diversification in a Subgroup of Chronic Lymphocytic Leukemia Patients with Stereotyped IGHV4-34/IGKV2-30 B cell Receptors: Implications for Ongoing Interactions with Antigen.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2337-2337
Author(s):  
Lesley-Ann Sutton ◽  
Efterpi Kostareli ◽  
Anastasia Hadzidimitriou ◽  
Nikos Darzentas ◽  
Athanasios Tsaftaris ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Heavy Chain ◽  
Conflicts Of Interest ◽  
Lymphocytic Leukemia ◽  
B Cell Receptors ◽  
Valuable Insight ◽  
Cell Receptors ◽  
Mutational Status ◽  
The Impact

Abstract Abstract 2337 Poster Board II-314 Several studies indicate that the development of chronic lymphocytic leukemia (CLL) may be influenced by antigen (Ag) recognition through the clonotypic B cell receptors (BCRs). However, it is still unclear whether Ag involvement is restricted to the malignant transformation phase or whether the putative Ag(s) may continuously trigger the CLL clone. Valuable insight into these issues may be gleaned from the study of intraclonal diversification (ID) within the immunoglobulin (IG) genes through ongoing somatic hypermutation (SHM). Definitive data regarding ID within IG genes in CLL remains limited and conflicting. In the present study we systematically explored the presence of ID within IG genes of CLL, not only at cohort level but also in subgroups defined by BCR stereotypy and IG gene mutational status. We thus conducted a large-scale subcloning study of both IG heavy and light variable genes, in a total of 1496 and 1008 subcloned sequences from 71 and 56 CLL cases, respectively. The analysis was intentionally biased to cases expressing IGHV4-34/IGKV2-30 IGs (subset #4) and IGHV3-21/IGLV3-21 IGs (subset #2) that exhibit distinctive, subset-biased SHM patterns. PCR reactions were run using the high-fidelity Accuprime Pfx polymerase and at least 14 colonies/case were analyzed. All “non-ubiquitous” sequence changes from the germline were evaluated and recorded as follows: (i) unconfirmed mutation (UCM) - a mutation observed in only one subcloned sequence from the same sample (ii) confirmed mutation (CM) - a mutation observed more than once among subcloned sequences from the same sample. Analysis of heavy chain sequences revealed that 40% (28/71) of cases carried intraclonally diversified IGHV-D-J genes with CMs amongst subclones, whilst 32% (23/71) of cases carried only UCMs. The remaining 28% (20/71) of cases carried sets of identical IGHV-D-J subcloned sequences. Although most cases showed no or low levels of ID, an intense and, likely, functionally driven ID was evident in selected cases, especially those belonging to subset #4. The distinct ID in subset #4 was statistically significant when compared to all other groups defined by IGHV gene usage and mutation status, BCR stereotypy or heavy chain isotype. Subsequent analysis of the clonotypic light chains revealed that the impact of ID was generally low, with the outstanding exception again relating to subset #4. In fact, of 22 IGKV-J rearrangements exhibiting CMs, 11 (50%) utilized the IGKV2-30 gene and notably 10/11 (91%) of these were expressed by subset #4 cases. In such cases, the expressed IGKV2-30 gene was affected by an active and precisely targeted ID, analogous to their partner IGHV4-34 gene. These findings suggest that the SHM mechanism may continuously operate in certain subsets of CLL patients, particularly those cases expressing stereotyped IGHV4-34/IGKV2-30 BCRs typical of subset #4. In such cases, the observed ID patterns attest to the very precise targeting of the SHM process and may be considered as evidence for a “stereotyped response” to an active, ongoing interaction with Ag(s). Disclosures: No relevant conflicts of interest to declare.

Analysis of Stereotyped IGHV Distribution In a Series of 1133 Chronic Lymphocytic Leukemia Patients: The Experience of a Multicenter Italian Study Group

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2423-2423
Author(s):  
Francesco Maura ◽  
Giovanna Cutrona ◽  
Massimo Gentile ◽  
Serena Matis ◽  
Monica Colombo ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Heavy Chain ◽  
Conflicts Of Interest ◽  
Pairwise Alignment ◽  
Immunoglobulin Heavy Chain ◽  
Lymphocytic Leukemia ◽  
Study Group ◽  
Data Set ◽  
Italian Study ◽  
Mutational Status

Abstract Abstract 2423 Chronic lymphocytic leukemia (CLL) is characterized by an extremely variable clinical course. Mutational status of the immunoglobulin heavy-chain variable (IGHV) region defines two disease subsets with different prognosis. A fraction of CLL cases carries highly homologous B-cell receptors (BCR), i.e. characterized by non-random combinations of immunoglobulin heavy-chain variable (IGHV) genes and heavy-chain complementarity determining region-3 (HCDR3). We performed sequence analysis to characterize IGHV regions in a panel of 1133 CLL patients investigated by a multicenter Italian study group. A total of 1148 rearrangements were identified; the analysis of stereotyped subsets was performed based on previously reported criteria (Messmer et al, J Exp Med 2004; Stamatopuolos et al, Blood 2007). Specifically, we compared all our sequences with those found in three different publicly available data sets (Stamatopoulos et al, Blood 2007; Murray et al, Blood 2008 and Rossi et al, 2009 Clin Cancer Res). In addition, a pairwise alignment within all sequences was performed in order to discover novel potential subsets (HCDR3 identity > 60%). Based on the 2% cut-off used to discriminate between Mutated (M) and Unmutated (UM) cases, 777 sequences (67.59%) were classified as M, while 371 sequences (32.3%) as UM. The most represented IGHV genes within mutated cases were IGHV4-34 (104/118) and IGHV3-23 (85/96), whereas IGHV1-69 (97/112) was the most frequently used in the UM group. Interestingly, the IGHV3-21 gene, reported to be frequently expressed in CLL patients from Northern Europe, was present in only a small fraction of cases (24; 2.07%), confirming a previous finding reported by Ghia et al (Blood 2005) in a smaller panel. In our series, stereotyped HCDR3 sequences were found in 407/1148 (35.45%) patients, 177 of whom were M and 230 were UM cases. Overall, we observed that stereotyped sequences were significantly associated with UM IGHV status (Fisher's exact test, P<0.0001). Among the 407 stereotyped HCDR3 sequences, 345 belong to the clusters reported by Murray et al and 14 to those described by Rossi et al., 2009 Clin Cancer Res. The most frequent stereotyped subsets identified in our panel were #1 (35 cases), #7 (28 cases), #4 (24 cases), #3 and #9 (16 cases), #28 (13 cases), and #2 (12 cases), together with subsets #5, #8, #10, #12, #13, #16 and #22 (all ranging from 6 to 9 cases). Finally, we were able to identify by auto-matching analysis 48 sequences potentially specific for 23 novel putative stereotype subsets. In our series we identified 407/1148 (35.45%) stereotyped HCDR3 sequences. The percentage was higher than that reported by Stamatopoulos et al and Murray et al. This discrepancy may partially be due to the different approach used in our analysis, namely the matching to a general data set including all published stereotyped subsets instead of the auto-matching performed by those Authors. We demonstrated a significant association between IGHV status and stereotyped sequences and confirmed the finding that #1 is the most frequent subset identified so far. Finally, we were able to identify a series of 23 novel putative subsets that will require further confirmation. Disclosures: No relevant conflicts of interest to declare.

Microarray Gene Expression Profiling of B-Cell Chronic Lymphocytic Leukemia Subgroups Defined by Genomic Aberrations and VH Mutation Status

2004 ◽  
Vol 22 (19) ◽  
pp. 3937-3949 ◽  
Author(s):  
Christian Haslinger ◽  
Norbert Schweifer ◽  
Stephan Stilgenbauer ◽  
Hartmut Döhner ◽  
Peter Lichter ◽  
...  
Keyword(s):  
Gene Expression ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Differentially Expressed Genes ◽  
Expression Profiling ◽  
Lymphocytic Leukemia ◽  
Differentially Expressed ◽  
Mutational Status ◽  
Genomic Aberrations ◽  
Cell Chronic Lymphocytic Leukemia

Purpose Genomic aberrations and mutational status of the immunoglobulin variable heavy chain (VH) gene have been shown to be among the most important predictors for outcome in patients with B-cell chronic lymphocytic leukemia (B-CLL). In this study, we report on differential gene expression patterns that are characteristic for genetically defined B-CLL subtypes. Materials and Methods One hundred genetically well-characterized B-CLL samples, together with 11 healthy control samples, were analyzed using oligonucleotide arrays, which test for the expression of some 12,000 human genes. Results Aiming at microarray-based subclassification, class predictors were constructed using sets of differentially expressed genes, which yielded in zero or low misclassification rates. Furthermore, a significant number of the differentially expressed genes clustered in chromosomal regions affected by the respective genomic losses/gains. Deletions affecting chromosome bands 11q22-q23 and 17p13 led to a reduced expression of the corresponding genes, such as ATM and p53, while trisomy 12 resulted in the upregulation of genes mapping to chromosome arm 12q. Using an unsupervised analysis algorithm, expression profiling allowed partitioning into predominantly VH-mutated versus unmutated patient groups; however, association of the expression profile with the VH mutational status could only be detected in male patients. Conclusion The finding that the most significantly differentially expressed genes are located in the corresponding aberrant chromosomal regions indicates that a gene dosage effect may exert a pathogenic role in B-CLL. The significant difference in the partitioning of male and female B-CLL samples suggests that the genomic signature for the VH mutational status might be sex-related.

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