B Cell Lymphomas Are Resistant towards Bone Morphogenetic Protein (BMP) Induced Growth Inhibition.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2381-2381
Author(s):  
Kanutte Huse ◽  
Marianne B. Eide ◽  
Christian Kersten ◽  
Erlend B. Smeland ◽  
June H. Myklebust
Keyword(s):  
Growth Inhibition ◽  
B Cell ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cell ◽  
Lymphoma Cells ◽  
Bmp Receptors ◽  
Induced Apoptosis

Abstract Bone morphogenetic proteins (BMPs) belong to the TGF-β superfamily, and mediate their effects mainly through the Smad signalling pathway. Whereas TGF-β is well established as one of the most potent negative regulators in hematopoietic cells, the role of BMPs remains more elusive. We have previously shown that BMP-6 inhibits the growth of naïve and memory human B cells. As high BMP-6 mRNA expression is associated with poor outcome in diffuse large B cell lymphoma (DLBCL; Rosenwald et al, N Engl J Med 2002), we hypothesized that resistance towards BMP-induced growth inhibition is a possible mechanism for lymphomagenesis. In the current study, 7 B cell lymphoma cell lines (representing Burkitt lymphoma (BL) and DLBCL) and tumour material from lymphoma patients were investigated to unravel the role of BMPs in lymphomas. We analyzed the expression of BMP receptors by FACS analysis, and found variable expression of the BMP receptor type I (Alk2, Alk3 and Alk6) and type II (BMP RII, Activin RIIA and RIIB) among the cell lines and in primary lymphoma cells, suggesting variable binding of BMPs. We next investigated the effect of BMP-2, BMP-4, BMP-6 and BMP-7 on proliferation and survival of B lymphoma cell lines, and found 2 of 7 cell lines to be resistant towards BMP-2 and BMP-4 induced growth inhibition. In contrast, 4 of 7 and 7 of 7 cell lines were resistant to BMP-6 and BMP-7 induced growth inhibition, respectively. In Sudhl6 cells that were highly sensitive to BMP-2 and BMP-6 induced apoptosis and inhibition of proliferation, we demonstrated that the cytokines IL-10, CD40 Ligand and BLyS were able to counteract the negative effects induced by BMPs, while IL-2 and IL-4 were not. On the contrary, both BMP-2 and BMP-6 greatly increased anti-IgM activation induced apoptosis. In resistant lymphoma cells, the BMPs were not able to induce detectable levels or induced low levels of phosphorylated SMAD1/5/8 compared to sensitive cell lines. Low or no increase in phosphorylation of SMAD1/5/8 induced by BMPs could only partly be explained by low/ undetectable expression of BMP receptors. Hence, upregulation of inhibitory Smads (Smad6, Smad7) or mutations in receptors or Smads represent other possible mechanisms for resistance to BMPs in lymphomas, and this is currently under investigation. We also investigated if the lymphoma cells produced BMPs themselves and found that 5 of 7 cell lines and 3 of 5 primary lymphomas produced significant amounts of BMP-7. Some lymphoma cells also had detectable levels of BMP-4 and BMP-6. Our findings that lymphoma cells are resistant towards BMP-7 and to some degree BMP-6 induced growth inhibition, whereas they produce these cytokines, suggest that resistance towards BMP induced signalling in B cell lymphomas can contribute to increased tumour growth.

Anti-CD45 Mediated Cytolysis of Human T-Cell Lymphoma Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4637-4637
Author(s):  
Gerald G. Wulf ◽  
Anita Boehnke ◽  
Bertram Glass ◽  
Lorenz Truemper
Keyword(s):  
T Cell ◽  
B Cell ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Cytolytic Activity ◽  
T Cell Lymphoma ◽  
Lymphoma Cell ◽  
Lymphoma Cells ◽  
Human T Cell

Abstract Anti-CD45 mediated cytoreduction is an effective means for T-cell depletion in rodents and humans. In man, the CD45-specific rat monoclonal antibodies YTH24 and YTH54 are IgG2b subclass, exert a predominantly complement-dependent cytolytic activity against normal T-lymphocytes, and have been safely given to patients as part of conditioning therapies for allogeneic stem cell transplantation. The efficacy of such antibodies against human lymphoma is unknown. Therefore, we evaluated the cytolytic activity of YTH24 and YTH54 by complement-dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC), as well as by direct apoptotic and antiproliferative effects, against a panel of Hodgkin disease (HD) and non-Hodgkin lymphoma (NHL) cell lines, and against primary specimens. Significant CDC activity (>50% cytolysis) of the antibodies YTH54 and YTH24 was observed against three of five T-cell lymphoma lines, but against only one of nine B-cell lymphoma lines and none of four HD cell lines. The combination of YTH54 and YTH24 induced ADCC in all T-cell lymphoma cell lines and three primary leukemic T-cell lymphoma specimens, but were ineffective in B-cell lymphoma and HD cell lines.There were only minor effects of either antibody or the combination on lymphoma cell apoptosis or cell cycle arrest. In summary, anti-CD45 mediated CDC and ADCC via the antibodies YTH24 and YTH54 are primarily effective against lymphoma cells with T-cell phenotype, and may be an immunotherapeutic tool for the treatment of human T-cell lymphoma.

scholarly journals Identification of Anti-Lymphoma Biomarkers of Response to the Anti-CD37 Antibody Drug Conjugate (ADC) IMGN529

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4187-4187 ◽  
Author(s):  
Eugenio Gaudio ◽  
Chiara Tarantelli ◽  
Alberto Arribas ◽  
Luciano Cascione ◽  
Ivo Kwee ◽  
...  
Keyword(s):  
T Cell ◽  
B Cell ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
T Cell Lymphoma ◽  
Lymphoma Cell ◽  
Cell Of Origin ◽  
Antibody Drug Conjugate ◽  
Induced Apoptosis

Abstract Background IMGN529 is an antibody drug conjugate (ADC) consisting of an anti-CD37 antibody with direct anti-tumor activity conjugated via a thioether linker to the cytotoxic maytansinoid antimicrotubule agent DM1. IMGN529 has shown pre-clinical (Deckert et al, Blood 2013) and clinical activity in lymphoma (Stathis et al, ASH 2014; NCT01534715). Here, we assessed the anti-tumor activity of IMGN529 on a large panel of B cell and T cell human lymphomas to identify potential biomarkers of response. Methods Fifty-four lymphoma cell lines [diffuse large B cell lymphoma (DLBCL), n.=27; mantle cell lymphoma (MCL), n.=10; anaplastic large T-cell lymphoma, n.=5; marginal zone lymphomas, n=6, others, n=6] were exposed to increasing doses of IMGN529 or to the unconjugated DM1 for 72h. Cell proliferation was measured using the MTT. Apoptosis induction was defined by at least 1.5-fold increase in caspase 3/7 signal activation with respect to controls using the Promega ApoTox-Glo Triplex Assay. CD37 surface expression was assessed by cytofluorimetry. Gene expression profiling (GEP) was done with the Illumina HumanHT-12 Expression BeadChips on untreated cell lines followed by GSEA (NES > |2|, P<0.05, FDR<0.25) and limma t-test (FC> |1.2|; P< 0.05; top 200 up and top 200 down). Results. The IMGN529 median IC50 in the 54 cell lines was 780pM (95%C.I., 263pm-11.45nM). Activity was stronger (P<0.001) in B cell lymphoma cell lines (n= 46; median IC50=450pM; 95%C.I., 150-800pM) than in T cell lymphoma cell lines (n=8; median IC50=22.5nM; 95%C.I., 14-40nM). The median IC50 for DM1 was 30pM (C.I.95%, 20-40pM) with no differences between B and T cell lymphoma origin. IMGN529 induced apoptosis in 33/54 (61%) lymphoma cell lines. Surface CD37 expression was higher in cell lines derived from B than from T cells (P< 0.0001): IMGN529 IC50 values, but not of DM1, were negatively correlated with surface CD37 expression across all cell lines (R=-0.39; P= 0.018), but not within the individual B or T cell subgroups. Among B cell lines, DLBCL cell of origin, TP53 status or the presence of BCL2 translocation did not affect the sensitivity to IMGN529, while IC50s were higher in the presence of MYC translocation (P= 0.043). No association was seen between IMGN529-induced apoptosis or the sensitivity to DM1 with DLBCL cell of origin, TP53 status or the presence of BCL2 or MYC translocations. We then compared the baseline gene expression profiling of DLBCL cell lines that were highly sensitive to IMGN529 (IC50< 800pM; "S") versus less sensitive/resistant DLBCL cell lines (IC50>10nM, "R"), separately for germinal center B cell type (GCB) (S, n=11; R, n=8) and for activated B cell like (ABC) (S, n=4; R, n=3). In both DLBCL groups, MYC targets, genes involved in unfolded protein response, glycolysis and DNA repair were enriched in transcripts more expressed in R than S cell lines. Transcripts associated with low sensitivity included CD44, VIM, ANXA2, BCL2, ANXA2P1, HSP90B1, NFKBIZ, CDK6, BIRC5 in GCB and HSPA1B, HSP90AA1, CADM1, CD86, TUBB2A, TUBG1, NOTCH1 in ABC cell lines. HEBP1, PHB, PSME3, RNU6-15, RPL13 were more expressed in both GCB and ABC R. Genes involved in PI3K/AKT/mTOR, hypoxia, INF-gamma, TNFA signaling via NFKB and in complement were more expressed in S than in R cell lines. Genes associated with sensitivity to IMGN529 comprised: CD37 (IMGN529 target), CD79A, CHI3L2, FAM117B, LPAR5, NFATC1, PTPN22, RBM38, SGPP1, SLC6A16 in both GCB and ABC cell lines; BASP1, CXCR5, BIK, LY86, TLR10, CD86, LCK, CD22, PTPN22, BCL6, PIK3IP1, CDKN2A in GCB; AFF3, PIM1, MGMT, PDE4B, NFKBIE, SYK, FOXO1in ABC. Conclusions. IMGN529 showed a very strong anti-tumoral activity in pre-clinical lymphoma models. High expression of CD37 and mostly genes involved in BCR signalling were associated with sensitivity to IMGN529. Conversely, the presence of MYC translocation, a high expression of MYC targets and of genes known to be involved in drug resistance (BCL2, BIRC5, CDK6, heat-shock proteins, annexins, proteasome and tubulin components) appeared to negatively affect the response to the ADC but also represent therapeutic targets for novel combinations to be explored. Disclosures Rossi: Gilead: Honoraria, Research Funding; Abbvie: Honoraria; Janseen: Honoraria. Sloss:Immunogen Inc: Employment.

RAD001 (everolimus) down-regulates cyclin D3 and c-Myc and is particularly effective in the treatment of aggressive B-cell lymphomas

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 17573-17573
Author(s):  
S. H. Kuo ◽  
C. H. Hsu ◽  
P. Y. Yeh ◽  
H. C. Hsu ◽  
M. Gao ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
B Cell ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cell ◽  
Cyclin D3 ◽  
Akt Signaling Pathway ◽  
P70s6 Kinase ◽  
Aggressive B Cell Lymphoma

17573 Background: Aggressive B-cell lymphoma is recently found to be associated with constitutive activation of the PI3K/Akt pathway, and thereby is relatively resistant to apoptosis. Furthermore, activation of the PI3K/Akt signaling pathway can result in the upregulation of cyclin D3 and c-Myc through inhibition of the cap-independent RNA translation. Since mTOR is closely associated with the regulation of the translation process, and is a downstream mediator of the PI3K/Akt signalling pathway, this study aimed to examine if RAD001, an inhibitor of mTOR, can be effective in the treatment of aggressive B-cell lymphoma. Methods: Pfeiffer (diffuse large cell lymphoma), Ramos (EBV (−) Burkitt’s lymphoma), Raji (EBV (+) Burkitt’s lymphoma), and MC116 (EBV (−) undifferentiated lymphoma) cell lines were used in this study. Expression of pAkt, p70S6 kinase, pp70 S6 kinase, 4E-BP1, p4E-BP1, cyclin D3, and c-Myc was measured by immunoblotting. Results: Akt was constitutively activated in all four lymphoma cell lines. Downstream effectors of PI3/Akt signaling pathway, including p70S6 kinase and 4E-BP1, were also phosphorylated in these lymphoma cell lines. RAD001 downregulated p70S6 kinase and 4E-BP1, and the IC50s of growth suppression ranged from 5 to 50 nM, without significant difference between EBV (+) and EBV (−) cell lines. The IC50s of these lymphoma cell lines appeared to be much lower than those obtained from the carcinoma cell lines, suggesting that lymphomas may be particularly sensitive to growth inhibition by RAD001. RAD001 blocked cell cycle progression in G0/G1 phase in all four lymphoma cell lines while there was no significant increase in sub-G1 phases, suggesting that apoptosis was not the major mechanism of RAD001-induced growth inhibition. In addition, in parallel with the RAD001-induced growth inhibition, a dose-dependent down-regulation of the expression of cyclin D3 and c-Myc was demonstrated. Conclusions: The mechanism of action is at least partly due to downregulation of cyclin D3 and c-Myc, and subsequent G0/G1 block. Since overexpression of cyclin D3 and c-Myc is also closely associated with chemoresistance of aggressive B-cell lymphoma, RAD001 may be a useful adjunct in the treatment of this group of tumors. No significant financial relationships to disclose.

Role of phosphatidylinositol 3′-kinase/AKT pathway in diffuse large B-cell lymphoma survival

Blood ◽  
2006 ◽  
Vol 108 (13) ◽  
pp. 4178-4186 ◽  
Author(s):  
Shahab Uddin ◽  
Azhar R. Hussain ◽  
Abdul K. Siraj ◽  
Pulicat S. Manogaran ◽  
Naif A. Al-Jomah ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Akt Pathway ◽  
Phosphatidylinositol 3 Kinase ◽  
Large B Cell Lymphoma ◽  
Induced Apoptosis ◽  
Large B Cell

Abstract Phosphatidylinositol 3′-kinase (PI3K) is a key player in cell-growth signaling in a number of lymphoid malignancies, but its role in diffuse large B-cell lymphoma (DLBCL) has not been fully elucidated. Therefore, we investigated the role of the PI3K/AKT pathway in a panel of 5 DLBCL cell lines and 100 clinical samples. Inhibition of PI3K by a specific inhibitor, LY294002, induced apoptosis in SUDHL4, SUDHL5, and SUDHL10 (LY-sensitive) cells, whereas SUDHL8 and OCI-LY19 (LY-resistant) cells were refractory to LY294002-induced apoptosis. AKT was phosphorylated in 5 of 5 DLBCL cell lines and inhibition of PI3K caused dephosphorylation/inactivation of constitutively active AKT, FOXO transcription factor, and GSK3 in LY-sensitive cell lines. In addition, there was a decrease in the expression level of inhibitory apoptotic protein, XIAP, in the DLBCL cell lines sensitive to LY294002 after treatment. However, no effect was observed in XIAP protein levels in the resistant DLBCL cell lines following LY294002 treatment. Finally, using immunohistochemistry, p-AKT was detected in 52% of DLBCL tumors tested. Furthermore, in univariate analysis, high p-AKT expression was associated with short survival. In multivariate analysis, this correlation was no longer significant. Altogether, these results suggest that the PI3K/AKT pathway may be a potential target for therapeutic intervention in DLBCL.

scholarly journals Deletion of Murine Rhoh induces More Aggressive Diffuse Large B Cell Lymphoma (DLBCL) Via Interaction with Kaiso and Regulation of BCL-6 Expression

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1574-1574
Author(s):  
Hiroto Horiguchi ◽  
Marioara Felicia Ciuculescu ◽  
Anja Troeger ◽  
Haiming Xu ◽  
Christian Brendel ◽  
...  
Keyword(s):  
T Cell ◽  
B Cell ◽  
Cell Lines ◽  
Nuclear Localization ◽  
Germinal Center ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cell ◽  
Germinal Center Formation

Abstract RHOH encodes a GTPase-deficient, hematopoietic-specific small GTPase first identified as a hypermutable gene in DLBCL (Pasqualucci et al. 2001). RhoH is critical for T cell receptor signaling and Rhoh-deficient (RhohKO) mice have T cell lymphopenia (Gu et al., 2006) and loss of function mutations of RHOH are associated with Epidermodysplasia Verruciformis (Crequer et al., 2012). However, the role of RhoH in the biology of DLBCL is still unknown and its role in B lymphoid development is incompletely studied. We investigated the role of RhoH in normal germinal center formation and in a murine model of DLBCL by crossing RhohKO mice with Iµ-HABcl-6 transgenic (Bcl-6Tg) mice (Cattoretti G, et al., 2005). In young RhohKOmice, deficient development of CXCR5+ follicular T helper (Tfh) cells results in defective germinal center (GC) formation and impaired immunoglobulin switching in vivo. In spite of this defect in GC formation, RhohKO; Bcl-6Tg (KOTg) mouse demonstrated accelerated lymphoma progression associated with larger spleens and significantly earlier death (Log-rank test p<0.01, Figure 1). Immunohistochemistry data suggested increased expression of IRF-4 and enhanced expression of BCL-6 in KOTg mice, findings confirmed by immunoblot and consistent with an activated B-cell (ABC)-DLBCL phenotype. To analyze the mechanism underlying these results, B cell lymphoma cell lines from KOTg lymphoma mice were established. Multiple attempts to establish RhohWT lymphoma cell lines failed, although we also successfully established a lymphoma cell line from RhohKO; Bcl-6(ntg) (KONtg) mice. Re-expression of RhoH in these lines via retrovirus mediated gene transfer led to significantly decreased proliferation (5.9x106±9.6x105 cells vs 8.6x106±9.6x105 cells after 5-days culture; KOTg vs KOTg-RhoH, mean±SEM, p<0.05) that was associated with clear reduction in BCL-6 expression. These data suggest that BCL-6 is a direct or an indirect transcriptional target of RhoH. Our laboratory previously reported that KAISO, a dual-specific, Broad complex, Trantrak, Bric-a-brac/Pox virus, Zinc finger (POZ-ZF) transcription factor interacts and colocalizes with RhoH in the nucleus, whereas knockdown of RhoH inhibits the nuclear localization of KAISO in Jurkat cells (Mino A, et al., 2016). In addition, Kaiso has been shown to be a key regulator of spleen germinal center formation by repressing Bcl-6 expression in splenocytes (Koh D, et al., 2013). We hypothesized that the deletion of Rhoh may lead to the decreased nuclear localization of KAISO and result in increased the expression of Bcl-6. We first confirmed that RhoH bound KAISO in RhoH-transduced KO lymphoma cells by co-immunoprecipitation. Further immunoblot analysis and quantitative PCR (qPCR) demonstrated decreased BCL-6 expression in lymphoma cells in which RhoH was re-expressed (KOTg-RhoH and KONtg-RhoH) compared with empty vector-transduced lymphoma cell lines. Interestingly, p53 a BCL-6 target was increased in RhoH-transduced lymphoma cell lines. These data indicate that RhoH affects BCL-6 expression in B cell lymphoma cell lines and suggest that RhoH may be involved in DLBCL development by co-regulating BCL-6 expression affecting downstream targets via interaction with KAISO. Figure. Figure. Disclosures Williams: Bluebird Bio: Research Funding.

scholarly journals Targeting MYC-Driven B-Cell Lymphoma By Inhibition of the Histone Methyltransferase DOT1L

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2839-2839 ◽  
Author(s):  
Anagha Deshpande ◽  
Benson Chen ◽  
Parham Ramezani-Rad ◽  
Alessandro Pastore ◽  
Luyi Zhao ◽  
...  
Keyword(s):  
B Cell ◽  
Board Of Directors ◽  
Cell Lines ◽  
Target Genes ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cell ◽  
Advisory Committees ◽  
Genome Wide

Abstract Aberrant activation of the MYC proto-oncogene is a recurrent feature in human B-cell lymphomas of diverse sub-types, correlating with adverse prognosis and therapy resistance. Direct pharmacological MYC-targeting has proved difficult, but recent studies have shown that targeting chromatin regulators critical for MYC-driven oncogenesis may provide alternative avenues for therapeutic intervention. Recently, it has been demonstrated that MYC-driven oncogenesis in certain solid tumors is dependent on the histone 3 lysine 79 (H3K79) methyltransferase DOT1L. We hypothesized that B-cell lymphomas with hyperactive MYC-signaling might be responsive to DOT1L inhibition. In order to test this hypothesis, we tested the effect of the DOT1L inhibitor Pinometostat (EPZ-5676) on a panel of human B-cell lymphoma cell lines featuring elevated MYC. Pinometostat treatment reduced global H3K79 methylation levels, accompanied by a time and dose-dependent decrease in proliferation of several Burkitt's lymphoma cell lines including P493-6, Daudi and Raji. We observed that key MYC-target genes including CDK4, PPAT and NPM1 were significantly downregulated upon Pinometostat treatment, suggesting that DOT1L is required for the transcriptional activation of MYC-target genes in these cells. Pinometostat-treated B-lymphoma cells showed a significant decrease of cells in S-phase compared to controls as assessed by BrdU-labeling assays. Similar results were also obtained in a panel of B-cell lymphoma cell lines with MYC-rearrangements including mantle cell lymphoma (MCL) cell lines Jeko-1, JVM2, Mino-1 and Maver-1 and the diffused large B-cell lymphoma (DLBCL) cell line Karpas 422. Next, we sought to investigate whether the DOT1L-dependence of MYC-driven B-cell lymphoma could be reproduced in a well-defined model of MYC-driven B-cell lymphoma. Towards this end, we utilized a mouse model in which expression of the Cre recombinase from a B cell specific promoter leads to ectopic expression of a transgenic human MYC allele and concomitant deletion of the tumor suppressor Pten in B cells. Similar to our in vitro studies, Pinometostat treatment led to a significant reduction in proliferation of B-cell lymphoma cells from these mice with an IC50 of 0.5 µM. Furthermore, we sought to ascertain whether these findings reflected on-target effects related to DOT1L inhibition. Therefore, we deleted DOT1L using CRISPR/Cas9 in B-cell lymphoma cell lines and assessed the effect on proliferation using competitive-proliferation assays. We observed that DOT1L-deletion progressively diminished the relative growth of anti-DOT1L sgRNA-expressing P493-6 and Jeko1 cells compared to non-targeted cells invitro. In order to test the requirement for DOT1L in lymphoma propagation in vivo, we performed intravenous injections of equal number of Jeko-1 cells with either anti-DOT1L or anti-Renilla control sgRNAs into sub-lethally irradiated non-obese diabetic/severe combined immunodeficiency mice (NOD/SCID) mice. Mice injected with control anti-Renilla sgRNAs succumbed to disease with a median latency of 34 days while the latency of disease in the anti-DOT1L sgRNA cohort was 45 days. In summary, DOT1L depletion significantly delayed disease latency in this invivo disseminated model of B-cell lymphoma (P=0.02). We then performed transcriptomic analyses of Pinometostat-treated B-cell lymphoma cell lines compared to DMSO-treated counterparts using RNA-seq. Gene-set enrichment analysis (GSEA) of RNA-seq data from three different B-cell lymphoma cell lines demonstrated that Pinometostat treatment significantly decreased the expression of MYC-target genes. In order to investigate the intriguing role of DOT1L in regulating MYC-target gene expression, we used ChIP-seq to assess the genome-wide occupancy of MYC following DOT1L inhibitor treatment. Strikingly, our studies demonstrated that DOT1L inhibition significantly reduced the chromatin occupancy of MYC. Taken together, our experiments demonstrate the role of DOT1L in MYC-driven B-cell lymphoma pathogenesis invitro and invivo. Furthermore, our genome-wide studies demonstrate the importance of DOT1L for genomic MYC occupancy. Based on these findings, we propose that therapeutic DOT1L targeting may be a viable strategy in MYC-driven B-cell lymphoma. Disclosures Weigert: Roche: Research Funding; Novartis: Research Funding. Rickert:Pfizer: Employment. Ren:Elli Lilly: Consultancy, Membership on an entity's Board of Directors or advisory committees; Arima Genomics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Deshpande:Salgomed Therapeutics: Membership on an entity's Board of Directors or advisory committees; A2A Pharma: Membership on an entity's Board of Directors or advisory committees.

scholarly journals Endogenous Neurotrophins and Trk Signaling in Diffuse Large B Cell Lymphoma Cell Lines Are Involved in Sensitivity to Rituximab-Induced Apoptosis

PLoS ONE ◽  
2011 ◽  
Vol 6 (11) ◽  
pp. e27213 ◽  
Author(s):  
Cynthia Bellanger ◽  
Lydie Dubanet ◽  
Marie-Claude Lise ◽  
Anne-Laure Fauchais ◽  
Dominique Bordessoule ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cell ◽  
Large B Cell Lymphoma ◽  
Induced Apoptosis ◽  
Large B Cell

Combination of NVP-BEZ235 and Enzastaurin on B-Cell Lymphoma Cell Lines: An Effective Therapeutic Strategy

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2719-2719 ◽  
Author(s):  
Monica Civallero ◽  
Maria Cosenza ◽  
Samantha Pozzi ◽  
Stefano Sacchi
Keyword(s):  
Bone Marrow ◽  
B Cell ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoid Cells ◽  
Lymphoma Cell ◽  
Lymphoma Cells ◽  
Apoptosis Pathways ◽  
Extrinsic Apoptosis

Abstract Abstract 2719 Non-Hodgkin's lymphoma is the most common hematologic neoplasm in adults. Chemotherapy combined with CD-20 monoclonal antibodies has improved survival in both indolent and aggressive B-NHL. However, a substantial subset of patients does not achieve a cure or long disease remission. This has promoted the identification of new targeted treatments and new agents that have shown promising efficacy for future B-NHL therapies. The phosphatidylinositol 3-kinase (PI3K) mammalian target of rapamicin (mTOR) pathway mediates proliferation, survival and drug resistance in lymphoma cells. NVP-BEZ235 (BEZ235) is a new, orally bio available inhibitor of PI3K and mTOR and a representative of a new class of anti-tumour agents. In the current study, the efficacy of the combination of two orally available inhibitor to PI3K/mTOR (BEZ235) and PKCbeta/AKT (enzastaurin) was evaluated in B-cell lymphoma cell lines (RL, WSH-NHL, Jeko and Granta). First, we tested the anti-lymphoma activity of BEZ235 alone and in combination with enzastaurin, everolimus and perifosine. Results using MTT assay were expressed as fraction of cells killed by the individual drug or the combination in the drug-treated versus untreated cells. The interaction between drugs was analyzed by isobologram analysis using the STACorp8.2 software program based upon the Chou-Talalay method to determine if the combination were additive or synergistic. We found that enzastaurin, everolimus and perifosine enhanced the cytotoxicity triggered by BEZ235; a clear synergistic interaction (CI<1) appeared after 48 hours using low concentrations of the all compounds. We examined the functional effects of BEZ235 alone and in combination on apoptosis in lymphoma cells. We demonstrated that BEZ235 (20nM) alone after 24 hours induces an increase of 8–10% of apoptotic cells versus untreated, instead BEZ235 (20nM) in combination with enzastaurin (5microM) after 24 hours induces an increase of 25%. We next defined mechanisms whereby BEZ235 alone and in combination induce apoptosis in lymphoid cells. In particular, BEZ235 combined with enzastaurin induces both intrinsic and extrinsic apoptosis pathways with caspase 3, caspase 9, caspase 8 cleavage. We also showed that the combination of BEZ235 and enzastaurin decreases viability and induce apoptosis in B-cell lymphoma cell lines and peripheral blood mononuclear cells (PBMCs) from lymphoma patients. The combination has no effect on normal PBMCs and suppresses cell prolipheration of B-cell lymphoma cell lines (RL and Jeko) when co-cultured with bone marrow stromal cells in a system that mimics the bone marrow microenvironment. BEZ235, enzastaurin, everolimus and perifosine are inhibitors of intracellular pathways, thought we investigated effects of BEZ235 alone and in combinations with the other compounds in targeting p-AKT, p-mTOR, p-GSK3beta, p-p70, p-p90, p-MAPK, p-4EBP1 and cyclin D1 pathways by Western Blot. In addition, we demonstrated that BEZ235 plus enzastaurin resulted in increased expression of pro-apoptotic Bim, and in decrease expression of anti-apoptotic Bcl-2, which could not be abrogated by BEZ235 alone. In conclusion, our data suggest that in B cell lymphoma cell lines, BEZ235 in combination with enzastaurin elicits its antitumor effect better that combinated with perifosine and everolimus. Our data reveals that the drug combination targets phosphorilation of PI3K/Akt/mTOR pathways and induces both intrinsic and extrinsic apoptosis pathways. Furthermore, inhibition of Bcl-2 anti-apoptosis family members may, in part, explain the efficacy of signalling blockade in lymphoma cells and suggests an additional therapeutic targeting strategy. Therefore, these preclinical data support the potential use of BEZ235 in patients with NHL, and in particular provide rationale for combination with enzastaurin. Disclosures: No relevant conflicts of interest to declare.

Epstein-Barr virus-mediated protection against etoposide-induced apoptosis in BJA-B B cell lymphoma cells: role of Bcl-2 and caspase proteins

2004 ◽  
Vol 149 (2) ◽  
pp. 289-302 ◽  
Author(s):  
A. Blood ◽  
C. J. Edwards ◽  
H. H. Ishii ◽  
B. K. Pat ◽  
G. Bryson ◽  
...  
Keyword(s):  
B Cell ◽  
Epstein Barr Virus ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cells ◽  
Barr Virus ◽  
Epstein Barr ◽  
Induced Apoptosis

scholarly journals Prosurvival and Chemoprotective Effects of Tumor-Associated Macrophages Reversed By Anti-CSF-1R Monoclonal Antibody in B-Cell Lymphomas

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 498-498
Author(s):  
Anupama Gopisetty ◽  
Myriam Foglietta ◽  
Min Zhang ◽  
Zhiqiang Wang ◽  
Nathan Fowler ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Line ◽  
Tumor Cells ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cell ◽  
B Cell Lymphomas ◽  
Lymphoma Cells ◽  
Tumor Survival

Abstract The results of gene expression profiling (GEP) and immunohistochemical studies indicate that survival is worsened by macrophages (MΦ) in the tumor microenvironment of various B-cell lymphomas including follicular lymphoma (FL) and diffuse large B-cell lymphoma (DLBCL). Tumor-associated macrophages (TAMs) are known to be different from other types of MΦ, but the effects of TAMs that worsen prognosis in B-cell lymphoma are essentially unknown, as are the mechanisms of these effects. Here, we determined the phenotype and effects of TAMs on tumor survival, proliferation, and drug resistance in B-cell lymphomas and evaluated strategies to reverse their effects. As compared to peripheral blood monocytes (Mo) from normal donors (ND), Mo from FL patients were differentiated less into M1 MΦ (defined as CD68+CD163loCD206loCD86hi) by culture with CSF-1 for 5 days followed by IFN-g + LPS for 2 days more. In contrast, Mo from FL patients and ND were differentiated similarly into M2 MΦ (defined as CD68+CD163hiCD206hiCD86lo) by culture with CSF-1 followed by IL-4. Consistent with this, MΦ gene signatures from FL tumors were more similar to previously-described signatures of M2 rather than M1 MΦ (Martinez et al, J Immunol, 2006, 177(10):7303-11). In co-culture, primary FL tumor cells and lymphoma cell lines (including RL, a transformed FL cell line; Granta 519, a mantle cell lymphoma (MCL) cell line; and Raji, a Burkitt lymphoma cell line) induced differentiation of Mo into MΦ. Differentiation could be prevented by CS4 monoclonal antibody (mAb), a fully human IgG1 anti-human CSF-1R mAb (ImClone/Eli Lilly), but not isotype control Ab. Elevated levels of CSF-1 in culture supernatants after addition of CS4 mAb and real-time PCR of tumor cells suggested secretion of CSF-1 by lymphoma cells. Spontaneous apoptosis of primary FL and MCL tumor cells, determined by Annexin V and propidium iodide staining, was significantly reduced by co-culture with ND Mo (p<0.01), whether pre-differentiated into MΦ with CSF-1 or not, but this protection could be reversed by CS4 mAb. Mo and/or pre-differentiated MΦ protected primary FL and MCL tumor cells from cytotoxic effects of doxorubicin and/or bendamustine (p<0.01), but CS4 mAb reversed this effect. To assess effects of MΦ on proliferation, lymphoma cell lines (RL, Granta 519, and Raji) were CFSE-labeled prior to co-culture with Mo and doxorubicin, and proliferation assessed by CFSE dilution by flow cytometry in the presence or absence of CS4 or isotype control mAbs. MΦ promoted proliferation of all three cell lines, but this effect could be reversed by CS4 mAb. To further understand the mechanism by which MΦ promote tumor survival and growth, we performed phosflow analysis and found increased phosphorylation of STAT3 in co-cultured lymphoma cells. Consistent with this, we observed a correlation between an 11-gene STAT3 activation signature, described by Huang et al in DLBCL tumors (J Clin Oncol, 2013, 52.8414), and a MΦ gene signature in whole genome GEP studies of 191 FL tumors (Pearson correlation co-efficient=0.396, p<0.001). In conclusion, our results suggest that Mo from FL patients are predisposed to differentiate into an M2-like MΦ state. The interaction between lymphoma cells and Mo/MΦ is reciprocal: a change in Mo (MΦ differentiation) induced by interaction with lymphoma tumor cells leads to a change in the tumor cells (promotion of survival, proliferation, and chemoresistance). More importantly, our results demonstrate that targeting TAMs using CS4, an anti-CSF-1R mAb, can be an effective strategy to overcome the adverse effects of TAMs and reverse chemoresistance. Further studies are needed to determine whether STAT3 activation contributes to the protumor effects of TAMs. This may provide novel insights into the molecular mechanisms related to TAMs and lymphoma cells and offers additional targets for therapeutic development. In the long term, strategies targeting TAMs is especially appealing, as they should be able to be combined with existing therapies including chemotherapy, other immunotherapy, and targeted therapy, potentially improving their efficacy without increasing toxicity for FL, DLBCL, and other B-cell malignancies. Disclosures No relevant conflicts of interest to declare.

Sign in / Sign up

Export Citation Format

Share Document