Overexpression of BCL-2 Does Not Inhibit Autophagy in Human Follicular and Diffuse Large B-Cell Lymphomas

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3007-3007
Author(s):  
Aine McCarthy ◽  
Andrew James Clear ◽  
Jacek Marzec ◽  
Rita Coutinho ◽  
Robert D. Petty ◽  
...  
Keyword(s):  
B Cells ◽  
Protein Expression ◽  
B Cell ◽  
Prognostic Significance ◽  
B Cell Lymphoma ◽  
Beclin 1 ◽  
Pcr Array ◽  
B Cell Lymphomas ◽  
Autophagy Activity ◽  
Large B Cell

Abstract Background: Human follicular lymphoma (FL) and diffuse large B-cell lymphoma (DLBCL) are the most common forms of indolent and aggressive NHL, respectively. The t(14;18) translocation characterizes approximately 85% of FL and 20% of DLBCL and results in constitutive overexpression of the anti-apoptotic protein BCL-2. It was previously reported that BCL-2 plays dual roles in preventing apoptosis and autophagy. Autophagy is a physical and pathological process whereby cells sequester portions of cytoplasm including organelles to form autophagosomes where they are degraded and recycled. Growing evidence demonstrates that autophagy plays important roles in tumorigenesis, tumor progression, and resistance to chemotherapy. Aims: The autophagy status in human B-cell lymphomas is unknown. We hypothesized that overexpression of BCL-2 could change autophagy status and aimed to determined expression of autophagy-related genes and proteins in FL and DLBCL primary samples by PCR array and tissue microarray. We aimed to evaluate whether expression of the autophagy-related proteins p62, Beclin-1 and LC3 individually and in combination with BCL-2 protein expression could risk-stratify FL and DLBCL patients at diagnosis. Patients and methods: Using PCR array, the autophagy-related gene expression profiles were determined in purified and unpurified reactive and malignant human lymph node tissue biopsies. Diagnostic tissues from FL (n=117) and DLBCL (n=109) patients were microarrayed and autophagy protein expression was evaluated using immunohistochemistry. Univariate and multivariate analyses on both continuous and categorical variables were conducted to measure overall survival (OS), disease specific survival (DSS), and progression-free survival (PFS). Results: Seven autophagy machinery genes were up-regulated in purified FL B-cells, namely ATG9A, ATG16L1, MAP1LC3A, GABARAPL1, ULK1, LAMP1 and HDAC6 compared with reactive B-cells. Two autophagy machinery genes, MAP1LC3A and DRAM1, were up-regulated in DLBCL B-cells. In unpurified tissue biopsies, 20 of 46 genes in FL and 2 of 5 genes in DLBCL with increased expression were autophagy machinery genes. CTSD (cathepsin D) and TGM2 (transglutaminase 2) genes and proteins were mainly up-regulated in DLBCL tumor-infiltrating macrophages. These results demonstrate that FL and DLBCL showed increased expression of autophagy-related genes, regardless of the heterogeneity of these diseases. p62, a selective autophagy substrate; LC3, an autophagosome membrane protein; and Beclin-1, an essential autophagy effector, are often used to evaluate autophagy activity in the cell. 91% FL samples were BCL-2 positive but significantly decreased expression of p62, LC3 and Beclin-1 in FL samples was observed in both intra-follicular and non-malignant inter-follicular areas. This suggests that increased basal autophagy activity in both malignant FL cells and surrounding tumor infiltrating cells, indicating that BCL-2 does not inhibit basal autophagy activity. DLBCL samples displayed heterogeneous expression patterns of BCL-2, p62, LC3 and Beclin-1. We found that decreased p62 expression confers worse OS (continuous P=0.015; and categorical P=0.003), DSS (continuous P=0.037; categorical P=0.014) and PFS (categorical P=0.002) in DLBCL patients. Decreased expression of Beclin-1 was also confers poor prognosis in both FL and DLBCL as conducted by categorical analysis, OS (DLBCL, P=0.015; FL, P=0.004), DSS (FL, P=0.006), and PFS (DLBCL, P=0.029). p62 retains prognostic significance after adjustment for the International Prognostic Index (IPI) score and levels of BCL-2, Beclin-1 and LC3 in multivariate analysis. Beclin-1 retains its prognostic significance in FL after adjusting for FLIPI scores. Low p62 plus high BCL-2 expression in DLBCL confers the worst OS (P<0.0001) and DSS (P=0.001) compared with other combinations. Conclusions: These results demonstrate that FL has increased basal autophagy activity, while it varies in DLBCL. p62 is a novel, independent prognostic biomarker for DLBCL but not for FL. Combining p62 with BCL-2 provides a more robust and reliable method to risk-stratify DLBCL patients at diagnosis. Importantly, we report for the first time that overexpression of BCL-2 in human NHL does not inhibit basal autophagy activity. We propose that increased autophagy activity could be a therapeutic target for treatment of NHL. Disclosures Gribben: Celgene: Research Funding; Pharmacyclics: Honoraria; Roche: Honoraria.

Effect of siRNA interference of ubiquitin-specific protease 9X on apoptosis and growth of diffuse large B cell lymphoma cell line

2020 ◽  
Vol 10 (3) ◽  
pp. 446-453
Author(s):  
Wei Peng ◽  
Meizuo Zhong ◽  
Youhong Tang
Keyword(s):  
B Cells ◽  
Protein Expression ◽  
B Cell ◽  
Cell Apoptosis ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Large B Cell Lymphoma ◽  
Specific Protease ◽  
Sirna Interference ◽  
Large B Cell

Ubiquitin-specific protease 9X (USP9X) is crucial in the diagnosis and treatment of many tumor types, but its role in Diffuse Large B Cell Lymphoma (DLBCL) has not been determined. The current study aimed to examine the effects of RNA interference on USP9X expression, and subsequently on the bioactivity of DLBCL Farage and Pfeiffer cells. There were two groups in the study: USP9X-siRNA and NC. USP9X siRNA was transiently transferred into DLBCL cells by Cationic liposome. The total RNA was extracted using Fe2O3 and was retrieved into the DNA using the MagBeads Total RNA Extraction Kit. The protein expression of USP9X in Farage, Pfeiffer, and normal human B cell line at the cellular level was observed by Western blot. The Farage and Pfeiffer cells were infected with USP9X-siRNA. Cell apoptosis and cell growth viability were analyzed by flow cytometry and CCK8, Mcl-1 protein, a potential target of USP9X, and apoptosis factor proteins (such as Bak, Cytochrome C, Caspase 3, Caspase 8, PARP) were detected by Western blot after siRNA interference. The results showed that the protein expression of USP9X in malignant B cells was four times higher than that of the normal B cells. Inhibition of USP9X reduced the Mcl-1 activity, and increased the caspase-3, Bak and Cytochrome C activity. In the malignant B cells, Mcl-1 and Bak were binding in vivo; Bak was a new partner of Mcl-1. Inhibition of USP9X reduced cell proliferation and increased apoptosis. The expression of USP9X is upregulated in Diffuse large B cell lymphoma cells, Farage, and Pfeiffer. Inhibition expression of USP9X may induce cell apoptosis, inhibit cell growth, and downregulate Mcl-1 protein expression in Diffuse large B cell lymphoma cells, Farage, and Pfeiffer. USP9X has the ability in regulating cell apoptosis.

Constitutively Activated STAT3 Promotes Cell Proliferation and Survival in the Activated B Cell Subtype of Diffuse Large B Cell Lymphomas.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1621-1621
Author(s):  
Bihui Hilda Ye ◽  
Beibei Belinda Ding ◽  
Jian Jessica Yu ◽  
Raymond Y.-L. Yu ◽  
Lourdes M. Mendez ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
B Cells ◽  
B Cell ◽  
Germinal Center ◽  
B Cell Lymphoma ◽  
Cell Development ◽  
B Cell Development ◽  
Reciprocal Relationship ◽  
B Cell Lymphomas ◽  
Large B Cell

Abstract During B cell development, cell proliferation and survival are regulated by stage-specific transcription factors. Accordingly, distinct oncogenic pathways are employed by B cell lymphomas representing different stages of B cell development. Diffuse large B cell lymphoma (DLBCL) contains at least two main phenotypic subtypes, i.e. the germinal center B cell-like (GCB-DLBCL) and the activated B cell-like (ABC-DLBCL) groups. It has been shown that GCB-DLBCL responds favorably to chemotherapy and expresses high levels of BCL6, a transcription repressor known to play a causative role in lymphomagenesis. In comparison, ABC-DLBCL has lower levels of BCL6, constitutively activated NF-kappaB and tends to be refractory to chemotherapy. In this study, we investigated the relationship between BCL6 and STAT3 expression/activation in DLBCL and normal GC B cells. Our results demonstrate that BCL6 directly inhibits transcription of the STAT3 gene by binding to two BCL6 sites in its 5′ regulatory region. As a result, high level STAT3 expression and activation are preferentially detected in ABC-DLBCL and BCL6-negative normal germinal center B cells. Specifically, in tonsillar GCs, STAT3 expression and activation is restricted to a previously uncharacterized subset of BCL6−Blimp-1− B cells in the apical light zone. The location and phenotype of these cells suggest that they are in the process of exiting the BCL6-directed GC program and transitioning to a plasma cell differentiation process governed by Blimp-1. The reciprocal relationship between BCL6 and STAT3 is also conserved in DLBCL such that STAT3 expression and activation is preferentially associated with the BCL6-low, ABC subtype. Most importantly, inactivating STAT3 by either AG490 or small interference RNA in ABC-DLBCL cells inhibits cell proliferation and triggers apoptosis. These phenotypes are accompanied by decreased expression of several known STAT3 target genes, including c-Myc, JunB and Mcl-1, and increased expression of the cell cycle inhibitor p27. In addition to identifying STAT3 as a novel BCL6 target gene, our results define STAT3 activation as a second oncogenic pathway operating in ABC-DLBCL and suggest that blocking STAT3 may be potentially therapeutic in treatment of these aggressive lymphomas.

scholarly journals Cell-of-origin classification using the Hans and Lymph2Cx algorithms in primary cutaneous large B-cell lymphomas

2022 ◽  
Author(s):  
Anne M. R. Schrader ◽  
Ruben A. L. de Groen ◽  
Rein Willemze ◽  
Patty M. Jansen ◽  
Koen D. Quint ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
Prognostic Significance ◽  
B Cell Lymphoma ◽  
Large Cell ◽  
Cell Of Origin ◽  
B Cell Lymphomas ◽  
Large B Cell Lymphoma ◽  
And Behavior ◽  
Large B Cell

Abstract Primary cutaneous diffuse large B-cell lymphoma, leg type (PCDLBCL-LT) and primary cutaneous follicle center lymphoma with a diffuse population of large cells (PCFCL-LC) are both primary cutaneous B-cell lymphomas with large-cell morphology (CLBCL) but with different clinical characteristics and behavior. In systemic diffuse large B-cell lymphoma, not otherwise specified (DLBCL-NOS), gene-expression profiling (GEP) revealed two molecular subgroups based on their cell-of-origin (COO) with prognostic significance: the germinal center B-cell-like (GCB) subtype and the activated B-cell-like (ABC) subtype. This study investigated whether COO classification is a useful tool for classification of CLBCL. For this retrospective study, 51 patients with PCDLBCL-LT and 15 patients with PCFCL-LC were analyzed for their COO according to the immunohistochemistry-based Hans algorithm and the NanoString GEP-based Lymph2Cx algorithm. In PCFCL-LC, all cases (100%) classified as GCB by both Hans and Lymph2Cx. In contrast, COO classification in PCDLBCL-LT was heterogeneous. Using Hans, 75% of the PCDLBCL-LT patients classified as non-GCB and 25% as GCB, while Lymph2Cx classified only 18% as ABC, 43% as unclassified/intermediate, and 39% as GCB. These COO subgroups did not differ in the expression of BCL2 and IgM, mutations in MYD88 and/or CD79B, loss of CDKN2A, or survival. In conclusion, PCFCL-LC uniformly classified as GCB, while PCDLBCL-LT classified along the COO spectrum of DLBCL-NOS using the Hans and Lymph2Cx algorithms. In contrast to DLBCL-NOS, the clinical relevance of COO classification in CLBCL using these algorithms has limitations and cannot be used as an alternative for the current multiparameter approach in differentiation of PCDLBCL-LT and PCFCL-LC.

scholarly journals Clinical Interest of LMO2 Testing in Aggressive Large B-Cell Lymphomas

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2899-2899
Author(s):  
Luis Colomo ◽  
Ivonne Vazquez ◽  
Natalia Papaleo ◽  
Marta Salido ◽  
Anna Puiggros ◽  
...  
Keyword(s):  
Protein Expression ◽  
B Cell ◽  
Board Of Directors ◽  
B Cell Lymphoma ◽  
Clinical Impact ◽  
Low Grade ◽  
B Cell Lymphomas ◽  
Advisory Committees ◽  
Diagnostic Categories ◽  
Large B Cell

Introduction: MYC rearrangements (MYCr) occur in 5 to 15% of diffuse large B-cell lymphomas (DLBCL) and 20 to 35% of high-grade B-cell lymphoma, NOS (HGBL-NOS), are a defining criterion of the category HGBL with rearrangements of MYC and/or BCL2/BCL6 (HGBL, with MYCr and BCL2/BCL6), and may be present in 90% of Burkitt lymphoma. The current WHO classification considers cytogenetic techniques as the appropriate tool to detect MYCr but does not define how to approach to the identification of such alteration. As the global incidence of MYCr in large B-cell lymphomas (LBCL) is low, it is necessary to clarity whether FISH or other cytogenetic methods have to be applied to all LBCL or only in selected cases. We previously identified LMO2 as a potential surrogate marker of MYCr in LBCL (Colomo L, Am J Surg Pathol 2017). Our aim with this study is to confirm this observation and evaluate the clinical impact of this marker in the survival of patients with LBCL. Methods: We have prospectively studied between September 2014 and July 2019 a new series of 180 LBCL including patients with DLBCL, HGBL, with HGBL, with MYCr and BCL2/BCL6, HGBL-NOS and transformed low-grade lymphomas into DLBCL (tDLBCL) diagnosed according to WHO criteria. LMO2 (clone 1A9-1), MYC (clone Y69) and a common immunohistochemistry (IHC) panel of B and T-cell markers have been used for the histological categorization of the cases, using whole tissue sections. The cutoff for LMO2 and MYC were 30% and 40%, respectively. MYC and BCL6 genes were studied using break apart probes, and BCL2 gene using dual-color dual-fusion probes (IGH/BCL2), all from Vysis-Abbott. We have statistically correlated the loss of expression of LMO2 and the overexpression of MYC with the presence or absence of MYCr. Moreover, we performed survival analyses assessing the clinical impact of LMO2 in a series of 162 LBCL patients (112 DLBCL, 20 HGBL, with MYCr and BCL2/BCL6, 4 HGBL-NOS and 26 tDLBCL). The survival series included cases diagnosed before 2014 with IHC and FISH data. Results: The prospective series included 132 patients with DLBCL (78M/52F; median age 67 years, range 35-95), 9 HGBL, with MYCr and BCL2/BCL6 (5M/4F; median age 67 years, range 42-85), 4 HGBL-NOS (2M/2F; median age 58 years, range 42-89), and 35 tDLBCL (31 transformed follicular lymphomas, 3 marginal zone lymphoma and 1 lymphoplasmacytic lymphoma; 23M/20F; median age 64 years, range 40-82). LMO2 and MYC were expressed as follows, respectively: 84/130 (65%) and 46/132 (35%) in DLBCL; 1/9 (11%) and 8/9 (89%) in HGBL, with MYCr and BCL2/BCL6; 0/4 and 3/4 (75%) HGBL-NOS; 25/34 (73%) and 7/33 (21%) tDLBCL. MYCr were identified in 9/132 (7%) DLBCL; all HGBL, with MYCr and BCL2/BCL6; 4/4 HGBL-NOS; 7/35 (20%) tDLBCL. The table shows the comparisons between LMO2 and MYC protein expression for the identification of the presence of MYCr in the series of LBCL. Whereas in the whole series LMO2 and MYC had similar results, among CD10-positive cases, LMO2 had better results than MYC and identified better the presence of MYCr than MYC protein expression. The 5-year progression-free survival (PFS) according the diagnostic categories was 59% for DLBCL, 28% for HGBL, with MYCr and BCL2/BCL6, 25% for HGBL-NOS and 22% for tDLBCL (P=0.015). In addition, PFS was significantly lower for the presence of MYCr (26% vs 53%, P=0.02) and MYC IHC expression (35% vs 53%, P=0.005), and showed a positive trend for LMO2 loss of expression (39% vs 52%, P=0.1). The 5-year overall survival (OS) according the diagnostic categories was 67% for DLBCL, 23% for HGBL, with MYCr and BCL2/BCL6, 50% for HGBL-NOS and 77% for tDLBCL (P<0.001). In addition, OS was significantly shorter for the presence of MYCr (37% vs 71%, P=0.002), MYC protein expression (46% vs 75%, P=0.001), and for LMO2 loss of expression (46% vs 74%, P=0.005). In a Cox regression survival analysis including IPI and LMO2 for the 68 CD10-positive cases, IPI (HR: 1.61 P=0.03) was the most important variable for predicting OS, and LMO2 showed a significant trend (HR: 0.44 P=0.06). However, the addition of MYC IHC and MYCr did not add predictive accuracy to IPI score (HR: 1.6 P=0.31; HR: 1.8 P=0.19, respectively). Conclusions: LMO2 detection by IHC is a useful tool to detect MYCr in aggressive LBCL, particularly in CD10-positive cases. Moreover, LMO2 protein expression captures the prognostic significance of the different diagnostic histological categories and the presence of MYCr in this group of lymphomas. Disclosures Sanchez-Gonzalez: Alexion: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Shire: Consultancy, Honoraria; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Salar:Celgene: Consultancy; Gilead: Consultancy, Speakers Bureau; Janssen: Consultancy, Speakers Bureau.

Inactivation Of BANK1 By a Novel IGH-Associated Translocation and 5’ Hypermethylation In B-Cell Lymphomas

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2497-2497 ◽  
Author(s):  
Kui Nie ◽  
Taotao Zhang ◽  
Jiong Yan ◽  
Leonardo Boiocchi ◽  
Shuhua Cheng ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Line ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
Methylation Status ◽  
B Cell Lymphoma ◽  
B Cell Lymphomas ◽  
Intron 1 ◽  
Large B Cell

Abstract A novel IGH-associated reciprocal translocation, t(4;14)(q24;q32), was identified, along with trisomy 9, in 20 of 20 metaphases by conventional karyotyping in a case of malignant gastric post-transplant lymphoproliferative disorder (PTLD). Cloning of the translocation site by inverse PCR identified BANK1 (B-cell scaffold protein with ankyrin repeats 1), a B-cell-specific adaptor protein with putative functions in B-cell receptor and CD40 signaling, as a novel IGH translocation partner. The breakpoints were located at the Sα region of IGH and intron 1 of BANK1. The translocation juxtaposed the two genes in opposite orientations, and surprisingly, resulted in transcriptional inactivation of BANK1 as a result of dissociation of the major BANK1 promoter. While BANK1 isoforms were expressed in all tonsillar B-cells, with lower levels (∼ 5 fold) in the germinal centers (GC) compared to naïve and memory B-cells, transcription from the major promoter in the tumor was absent and transcription from the minor promoter was reduced 50% relative to GC B-cells, suggesting that the non-translocated BANK1 allele was also inactivated. The total BANK1 expression was very low (∼10% of normal GC B cells) and crytic promoter activation was not identified. Several genes (PPP3CA, MIR1255A, FLJ20021 and SLC39A8), located 180 to 440 kb away from BANK1, were analyzed for mRNA expression; there is no significant activation in any of these genes, further supporting that BANK1is indeed the target gene affected by the translocation. Interphase FISH using break-apart BANK1 probes confirmed breakpoint in the index case but did not identify translocations in additional 15 PTLDs and 68 diffuse large B-cell lymphomas (DLBCL), implying that BANK1 translocation may be a rare event. To determine if BANK1 inactivation may occur in B-cell lymphomas by other mechanisms, 23 B-cell lymphoma cell lines, including 8 Burkitt lymphoma (BL), 9 diffuse large B cell lymphoma (DLBCL), 3 primary effusion lymphoma (PEL), and 3 classical Hodgkin lymphoma (cHL) were bisulfite sequenced to assess the methylation status of 37 CpG dinucleotides in a 436 base-pair region at the 5’ end of BANK1, which extends across exon 1 into the 5’ portion of intron 1. High level of methylation (>60% methylation on average among all CpGs) was seen in all 3 cHL and 2 of 3 PEL cell lines. Regional methylation was seen in 3 of 8 BL lines and 1 of 3 PEL lines. No hypermemethylation was identified in the DLBCL lines or in normal tonsils. Hypermethylation was associated with almost complete silencing of BANK1 transcription. In the DLBCL lines and BL lines without BANK1 hypermethylation, BANK1mRNA expressions were variable, ranging from <5% to 130% of GCB cells. To confirm that BANK1 hypermethylation is present in primary lymphoma cases, methylation status of 17 of the 37 CpGs were assessed in 23 cHL cases using en bloc formalin-fixed, paraffin-embedded materials and also laser-capture micro-issected Hodgkin/Reed-Sternberg (HRS) cells. There was evidence of BANK1 hypermethylation in the tumor cells in 9 of 23 cHL. Tumor cell specificity of BANK1 hypermethylation was further confirmed in 4 cHL cases using micro-dissected HRS cells. HRS cells were negative for BANK1 in 28 of 29 cHL cases examined by immunohistochemistry, suggesting that other mechanisms other than DNA methylation may be responsible for silencing BANK1expression. To investigate whether BANK1 has biological effects on B-cells related to lymphoma development, exogenous BANK1 was re-introduced to BC3, a PEL cell line showing marked BANK1 hypermethylation with absence of BANK1 expression. We established a stable doxycycline-inducible BC3 cell line expressing BANK1. Inhibition of cell growth was observed 2 to 3 days after doxycyline induction, and the number of viable cells with transfected BANK1 was only 25% compared to BC3 cells carry vehicle alone at day 6. An analysis of 5-bromo-2’ deoxyuridine (BrdU) incorporation after 48 hours of doxycline induction revealed that the fraction of cells in S-phase was reduced by 50% in the BANK1 transfectants, suggesting that BANK1has a negative effect on cell proliferation in these B cells. In summary, we have identified a novel IGH translocation partner and provide an example of an unusual consequence (gene inactivation) of IGH-associated translocation. We provide for the first time evidence of a potential role of BANK1 down-regulation in the development of B-cell lymphomas. Disclosures: No relevant conflicts of interest to declare.

scholarly journals C-X-C Chemokine Receptor 4 in Diffuse Large B Cell Lymphoma: Achievements and Challenges

2019 ◽  
Vol 142 (2) ◽  
pp. 64-70 ◽  
Author(s):  
Hui Du ◽  
Lei Gao ◽  
Jing Luan ◽  
Hangfan Zhang ◽  
Taiwu Xiao
Keyword(s):  
B Cells ◽  
B Cell ◽  
Chemokine Receptor ◽  
Cell Lymphoma ◽  
Bone Marrow Involvement ◽  
Prognostic Significance ◽  
B Cell Lymphoma ◽  
Large B Cell Lymphoma ◽  
Chemokine Receptor 4 ◽  
Large B Cell

Diffuse large B cell lymphoma (DLBCL), an aggressive cancer of the B cells, is the most common subtype of non-Hodgkin lymphoma (NHL) worldwide. In China, the cases of DLBCL increase yearly. C-X-C chemokine receptor 4 (CXCR4) has been implicated in the migration and trafficking of malignant B cells in several hematological malignancies, and only a few reports have been published on the role of CXCR4 in the metastasis of DLBCL. This review summarizes the relevant perspectives on the functional mechanism, prognostic significance, and therapeutic applications of the CXCL12/CXCR4 axis in DLBCL, in particular DLBCL with bone marrow involvement.

The Oncoprotein LMO2 Is Expressed in a Germinal Center B-Cell-Associated Pattern and Predicts Survival in Patients with Diffuse Large B-Cell Lymphoma.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 810-810
Author(s):  
Yasodha Natkunam ◽  
Eric D. Hsi ◽  
Christine Hans ◽  
Shuchun Zhao ◽  
Behnaz Taidi ◽  
...  
Keyword(s):  
Protein Expression ◽  
B Cell ◽  
Germinal Center ◽  
Cell Lymphoma ◽  
Prognostic Significance ◽  
B Cell Lymphoma ◽  
Rna Expression ◽  
Large B Cell Lymphoma ◽  
Significant Difference ◽  
Large B Cell

Abstract Diffuse large B-cell lymphoma (DLBCL) is clinically and molecularly heterogeneous and portends a poor prognosis in more than half the affected patients. We developed a multivariate model based on the RNA expression of six genes – LMO2, BCL6, FN1, CCND2, SCYA3 and BCL2 – that independently predicts survival in DLBCL patients treated with anthracycline-containing regimens (Lossos et al, NEJM 2004). Since the transcription factor LMO2 emerged as the strongest predictor of superior outcome, we generated a monoclonal anti-LMO2 antibody in order to document the tissue expression pattern of LMO2 protein and to establish its prognostic significance. Immunohistological analysis of 1200 normal tissues and hematolymphoid neoplasms showed that LMO2 protein is expressed as a nuclear marker in normal germinal center (GC) B-cells and in a subset of B-cell lymphomas. It is rarely expressed in mature T, NK and plasma cell neoplasms. Immature precursors of all bone marrow hematopoietic lineages and a significant proportion of acute lymphomphoblastic and myeloid leukemias express LMO2 protein. Apart from endothelial cells, no other non-hematolymphoid tissues we tested showed LMO2 protein expression. Hierarchical cluster analysis of immunohistologic data in DLBCL demonstrated that the expression profile of LMO2 protein is similar to that of other GC-associated proteins (HGAL, BCL6 and CD10) but different from that of non-GC proteins (MUM1/IRF4 and BCL2). LMO2 protein expression paralleled its RNA expression in B-cell lymphoma cell lines and was found in GC B, but not in non-GC, B-cell lines. To test the prognostic significance of LMO2 protein we analyzed an independent cohort of 203 DLBCL patients (mean age 63, range 18–93), uniformly treated with anthracycline-containing chemotherapy not containing rituximab, from four medical centers. No significant difference in response to therapy (CR and CRu) was observed between patients with LMO2-positive and LMO2-negative lymphomas (77% and 59%, respectively, p 0.05). However, Kaplan- Meier curves demonstrated a statistically significant difference in overall survival (OS) between LMO2-positive and LMO2-negative cases (p 0.035; median OS of 74 and 20 months, respectively). Similarly, event-free survival (EFS) was also significantly longer in patients with LMO2-positive compared to LMO2-negative lymphomas (p 0.01, median EFS of 48 and 12 months, respectively). The predictive power of LMO2 expression was IPI-independent. Multivariate analyses with protein expression profiles of HGAL, BCL6, CD10, JAW1, MUM1/IRF4 and BCL2 on this cohort of patients are underway to construct a clinically applicable immunohistologic algorithm for predicting survival. The capacity of LMO2 protein to identify DLBCL patients with improved outcome in an IPI-independent manner indicates its important role in risk stratification in this disease.

MYC Protein Expression Correlates with C-MYC Transcriptional Activity in Absence of Gene Rearrangement in Diffuse Large B-Cell Lymphoma

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4761-4761
Author(s):  
Anna Misyurina ◽  
Vsevolod Andreevich Misyurin ◽  
Andrey Vitalievich Misyurin ◽  
Sergey Kirillovich Kravchenko ◽  
Alla M. Kovrigina ◽  
...  
Keyword(s):  
B Cells ◽  
Protein Expression ◽  
B Cell ◽  
Transcriptional Activity ◽  
Cell Lymphoma ◽  
Mrna Level ◽  
B Cell Lymphoma ◽  
Expression Level ◽  
Large B Cell Lymphoma ◽  
Large B Cell

Abstract Introduction. MYC increases proliferative capacity of malignant B-cells, independently from mechanisms led to increased protein expression. Tumors with solely MYC expression are highly effective curable with doses intensification. Ability of cells to escape apoptosis by several mechanisms like BCL2 or P53 соехpression promotes malignant B-cells growth and survival and represents a big therapeutic problem. Aim. To investigate mechanism of MYC hyperexpression in absence of c-MYC -rearrangement in DLBCL (diffuse large B-cell lymphoma). To analyze efficacy of intensive chemotherapy in pts with DLBCL who underwent modified NHL-BFM-90 (m-NHL-BFM-90) plus rituximab (R) in correspondence with MYC and BCL2 protein expression. Patients and methods. Data of 62 DLBCL pts (35 males and 27 females) who underwent m-NHL-BFM-90+R in National Research Center for Hematology (Moscow) between 2004 and 2013 years were analyzed. Tumor samples were stained with antibody to BCL2 (clone 124, Dako) and MYC (clone Y69, Epitomics). We used a previously reported cut off for MYC expression ≥40% and BCL2 ≥50% (N. Johnson et al., 2012). G-banding data were available in 19 pts. In one case was revealed c-MYC rearrangement into heavy chain locus t(8;14)(q24;q32). In all other cases FISH didn't reveal c-MYC orBCL2 rearrangements with DNA probes Vysis LSI MYC Dual color, Break Apart Rearrangement Probe, Vysis LSI BCL2 Dual color, Break Apart Rearrangement Probe. RQ-PCR was performed in 17 cases to estimate mRNA level of c-MYC expression relatively to ABL. To assess relation between mRNA of c-MYC and MYC protein expression was used correlation analysis (R²). To estimate treatment results were performed Kaplan-Meyer and Cox regression analyses (SAS 9.3). Results: Majority of pts - 48/62 (78%) attended to a high-risk group according IPI (3-5). 9/62 (14,5%) patients had MYC+/BCL2- tumors, 15 (24%) - "double-expressor" (DE) MYC+/BCL2+, 21 (34%) - MYC-/BCL2+, 17 (27,5%) - MYC-/BCL2-. Median age of DE pts was statistically higher than others (61 (25-73) vs 47 (15-73) years old, P<0,05). In case with t(8;14)(q24;q32) MYC protein expression was ≥40%. In 18/50 (36 %) cases in absence of c-MYC -rearrangement MYC protein expression was ≥40%. In 27/43 (63 %) cases in absence of BCL2 rearrangement BCL2 expression was ≥50%. Median level c-MYC mRNA-expression was 1748 % (492 % - 5408 %). There was a tendency to increase MYC expression level with rising quantity of c-MYC mRNA (R2 = 0,13, P = 0,08). In case with t(8;14)(q24;q32) c-MYC mRNA level was higher than median (3940 %). 45 (78%) of pts achieved a complete remission, 4 from them had second remission. Relapses and progression of DLBCL developed in 9 (15%) and 7 (11%) pts. 5 (8%) pts died because of other reasons. DE DLBCL pts had the highest risk of relapse or progression within 4 years: MYC+/BCL2- - 14%, MYC-/BCL2 - 14%, MYC+/BCL2+ - 65%, MYC-/BCL2+ - 24% (P=0,02). In multivariate analysis MYC/BCL2 double expression had an independent prognostic power (HR 4,717, P=0, 0024) from IPI. Conclusion. We illustrated that MYC protein expression level correlates with c-MYC transcriptional activity in absence of c-MYC rearrangement. MYC hyperexpression alone didn't influence on prognosis, only coexpression of MYC and BCL2 had a crucial role increasing probability of relapse or progression in DLBCL patients. Disclosures No relevant conflicts of interest to declare.

scholarly journals Prognostic Significance of MYC Single, Double, Triple Hit and MYC-Translocation Partner Status in Diffuse Large B-Cell Lymphoma - a Study By the Lunenburg Lymphoma Biomarker Consortium (LLBC)

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 344-344 ◽  
Author(s):  
Andreas Rosenwald ◽  
Laurie H Sehn ◽  
Delphine Maucort-Boulch
Keyword(s):  
Clinical Trials ◽  
B Cell ◽  
Prognostic Significance ◽  
B Cell Lymphoma ◽  
Prognostic Impact ◽  
B Cell Lymphomas ◽  
Cox Models ◽  
Partner Gene ◽  
Post Therapy ◽  
Large B Cell

Abstract Introduction MYC-rearrangement (MYC-R) occurs in approximately 10-15% of diffuse large B-cell lymphomas (DLBCL) and several studies suggest an inferior progression free (PFS) as well as overall survival (OS) compared to DLBCL without MYC-R. However, the prognostic significance of MYC single-hit (MYC-SH), MYC double/triple-hit (MYC and BCL2 and/or BCL6 translocation; MYC-DH/TH) in the context of the MYC translocation partner (MYC-IG versus MYC-non-IG) is less clear due to relatively small sample sizes in prior studies. The Lunenburg Lymphoma Biomarker Consortium (LLBC) set out to address these questions in a large number of DLBCL treated uniformly with rituximab (R)-CHOP- or R-CHOP-like chemotherapy. Methods Relevant clinical data (IPI factors, PFS, OS) from patients with aggressive B-cell lymphomas with confirmed DLBCL morphology derived from registry cohorts (Canada: British Columbia Cancer, UK: Leeds/HMRN, Barts, USA: Stanford) and prospective clinical trials (Germany: RICOVER, MegaCHOEP, France: LNH01-5B and LNH03-B, HOVON: HO 46 and 84) were pooled in the LLBC database (n=5118). Interphase FISH analysis (mostly on tissue microarrays) was used to determine the MYC-, BCL2- and BCL6-rearrangement status. All DLBCL with available tissue underwent break-apart testing for MYC (Vysis, LSI, Abbott). Identified MYC-R cases were subjected to FISH break-apart testing for the BCL2- and BCL6 loci as well as to MYC/IGH fusion testing and, if negative, to MYC/IGK and MYC/IGL testing to allow designation of the final MYC-R status (MYC-IG versus MYC-non-IG). Information on the cell of origin (COO) was generated using immunohistochemistry (Hans classifier) and/or gene expression based methods. Survival probabilities were estimated using the Kaplan Meier method and survival curves were compared with the log-rank test. The effects of variables of interest were estimated using univariate and multivariate Cox models stratified on the variable 'cohort' and 'trial'. Results 2380 DLBCL patients with full data on PFS, OS, IPI factors and MYC FISH results were available for analysis. 263 DLBCL cases (11%) had MYC-R which was associated with inferior PFS and OS compared to DLBCL patients without MYC-R (59% versus 72% OS at five years; log-rank <0.001; 56% versus 64% PFS at five years; log-rank <0.002). This effect was observed in both COO subgroups (GCB and non GCB). Out of 163 MYC-R patients with complete FISH data, 40 were MYC-SH with an IG partner (24.5%) and 17 (10.5%) with a non-IG partner. 53 patients had a MYC DH/TH constellation with an IG partner (32.5%) and 53 patients (32.5%) with a non-IG partner. The MYC-DH/TH group with an IG partner had the worst OS at 24 months: (50.9% compared to 76.4% for the remaining MYC-R DLBCL patients and to 82.4% for patients without MYC-R) (fig1). Similar results were obtained for PFS. In the MYC-DH group, there were no differences in OS and PFS between MYC/BCL2 and MYC/BCL6 'double hits'. Multivariate Cox models adjusting for the IPI and including a time-dependent effect showed, that the impact on outcomes was mainly seen in the first 24 months post therapy (hazard ratio (HR) of 2 [1.59-2.53] for MYC-R DLBCL patients compared to patients without MYC-R and a HR of 3.12 [2.09-4.64] for MYC-DH/TH tumors in which MYC is translocated to IG). In contrast, in MYC-R patients including MYC-SH and MYC-DH/TH without an IG partner, the HR was lower at 1.5 [1.01-2.24]. Conclusion This study by the LLBC in a very large cohort of DLBCL patients treated with R-CHOP or R-CHOP-like therapy confirms previous reports on the negative prognostic impact of an underlying MYC-translocation for both PFS and OS. This impact is predominantly observed in the first two years post therapy. Further, the large sample size extends previous observations that the partner gene of MYC (IG versus non-IG) also has prognostic impact. MYC-DH/TH DLBCL with an IG partner gene have the worst OS and PFS, while impact is moderate in other constellations (MYC-SH and MYC-DH/TH with a non-IG partner). Additionally, no differences between MYC/BCL2 and MYC/BCL6 'double hits' are seen in PFS and OS. Our results suggest that along with MYC testing in routine clinical practice, identification of the MYC partner gene (IG versus non-IG) is also warranted to identify further DLBCL subsets with poor outcomes which may have implications in the design and interpretation of future clinical trials. Figure 1. Figure 1. Disclosures Sehn: Merck: Consultancy, Honoraria; TG Therapeutics: Consultancy, Honoraria; Roche/Genentech: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Seattle Genetics: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Morphosys: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria; Lundbeck: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria.

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