Targeting Bcl-2 family proteins modulates the sensitivity of B-cell lymphoma to rituximab-induced apoptosis

Blood ◽  
2008 ◽  
Vol 112 (8) ◽  
pp. 3312-3321 ◽  
Author(s):  
Claudia Stolz ◽  
Georg Hess ◽  
Patricia S. Hähnel ◽  
Florian Grabellus ◽  
Sandra Hoffarth ◽  
...  
Keyword(s):  
B Cell ◽  
Kinase Inhibitors ◽  
B Cell Lymphoma ◽  
Resistance Pattern ◽  
Phosphatidylinositol 3 Kinase ◽  
Endogenous Expression ◽  
Induced Apoptosis ◽  
Phosphatidylinositol 3

Abstract The chimeric monoclonal antibody rituximab is the standard of care for patients with B-cell non-Hodgkin lymphoma (B-NHL). Rituximab mediates complement-dependent cytotoxicity and antibody-dependent cellular cytotoxicity of CD20-positive human B cells. In addition, rituximab sensitizes B-NHL cells to cytotoxic chemotherapy and has direct apoptotic and antiproliferative effects. Whereas expression of the CD20 antigen is a natural prerequisite for rituximab sensitivity, cell-autonomous factors determining the response of B-NHL to rituximab are less defined. To this end, we have studied rituximab-induced apoptosis in human B-NHL models. We find that rituximab directly triggers apoptosis via the mitochondrial pathway of caspase activation. Expression of antiapoptotic Bcl-xL confers resistance against rituximab-induced apoptosis in vitro and rituximab treatment of xenografted B-NHL in vivo. B-NHL cells insensitive to rituximab-induced apoptosis exhibit increased endogenous expression of multiple antiapoptotic Bcl-2 family proteins, or activation of phosphatidylinositol-3-kinase signaling resulting in up-regulation of Mcl-1. The former resistance pattern is overcome by treatment with the BH3-mimetic ABT-737, the latter by combining rituximab with pharmacologic phosphatidylinositol-3-kinase inhibitors. In conclusion, sensitivity of B-NHL cells to rituximab-induced apoptosis is determined at the level of mitochondria. Pharmacologic modulation of Bcl-2 family proteins or their upstream regulators is a promising strategy to overcome rituximab resistance.

scholarly journals Validation of epigenetic mechanisms regulating gene expression in canine B-cell lymphoma: An in vitro and in vivo approach

PLoS ONE ◽  
2018 ◽  
Vol 13 (12) ◽  
pp. e0208709 ◽  
Author(s):  
Silvia Da Ros ◽  
Luca Aresu ◽  
Serena Ferraresso ◽  
Eleonora Zorzan ◽  
Eugenio Gaudio ◽  
...  
Keyword(s):  
Gene Expression ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Epigenetic Mechanisms

scholarly journals Cernunnos influences human immunoglobulin class switch recombination and may be associated with B cell lymphomagenesis

2012 ◽  
Vol 209 (2) ◽  
pp. 291-305 ◽  
Author(s):  
Likun Du ◽  
Roujun Peng ◽  
Andrea Björkman ◽  
Noel Filipe de Miranda ◽  
Cornelia Rosner ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
Class Switch Recombination ◽  
B Cell Lymphoma ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
End Joining ◽  
Class Switch

Cernunnos is involved in the nonhomologous end-joining (NHEJ) process during DNA double-strand break (DSB) repair. Here, we studied immunoglobulin (Ig) class switch recombination (CSR), a physiological process which relies on proper repair of the DSBs, in B cells from Cernunnos-deficient patients. The pattern of in vivo generated CSR junctions is altered in these cells, with unusually long microhomologies and a lack of direct end-joining. The CSR junctions from Cernunnos-deficient patients largely resemble those from patients lacking DNA ligase IV, Artemis, or ATM, suggesting that these factors are involved in the same end-joining pathway during CSR. By screening 269 mature B cell lymphoma biopsies, we also identified a somatic missense Cernunnos mutation in a diffuse large B cell lymphoma sample. This mutation has a dominant-negative effect on joining of a subset of DNA ends in an in vitro NHEJ assay. Translocations involving both Ig heavy chain loci and clonal-like, dynamic IgA switching activities were observed in this tumor. Collectively, our results suggest a link between defects in the Cernunnos-dependent NHEJ pathway and aberrant CSR or switch translocations during the development of B cell malignancies.

scholarly journals Tyrosine 1101 of Tie2 Is the Major Site of Association of p85 and Is Required for Activation of Phosphatidylinositol 3-Kinase and Akt

1998 ◽  
Vol 18 (7) ◽  
pp. 4131-4140 ◽  
Author(s):  
Christopher D. Kontos ◽  
Thomas P. Stauffer ◽  
Wen-Pin Yang ◽  
John D. York ◽  
Liwen Huang ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Vascular Development ◽  
Pi3 Kinase ◽  
Pleckstrin Homology Domain ◽  
Phosphatidylinositol 3 Kinase ◽  
Lipid Kinase ◽  
Embryonic Vascular Development ◽  
Phosphatidylinositol 3

ABSTRACT Tie2 is an endothelium-specific receptor tyrosine kinase that is required for both normal embryonic vascular development and tumor angiogenesis and is thought to play a role in vascular maintenance. However, the signaling pathways responsible for the function of Tie2 remain unknown. In this report, we demonstrate that the p85 subunit of phosphatidylinositol 3-kinase (PI3-kinase) associates with Tie2 and that this association confers functional lipid kinase activity. Mutation of tyrosine 1101 of Tie2 abrogated p85 association both in vitro and in vivo in yeast. Tie2 was found to activate PI3-kinase in vivo as demonstrated by direct measurement of increases in cellular phosphatidylinositol 3-phosphate and phosphatidylinositol 3,4-bisphosphate, by plasma membrane translocation of a green fluorescent protein-Akt pleckstrin homology domain fusion protein, and by downstream activation of the Akt kinase. Activation of PI3-kinase was abrogated in these assays by mutation of Y1101 to phenylalanine, consistent with a requirement for this residue for p85 association with Tie2. These results suggest that activation of PI3-kinase and Akt may in part account for Tie2’s role in both embryonic vascular development and pathologic angiogenesis, and they are consistent with a role for Tie2 in endothelial cell survival.

Novel cyclophosphamide of natural products osalmide and pterostilbene induces cytotoxicity and cell cycle arrest in diffuse large B-cell lymphoma cells

2020 ◽  
Vol 52 (4) ◽  
pp. 401-410
Author(s):  
Mengyu Xi ◽  
Wan He ◽  
Bo Li ◽  
Jinfeng Zhou ◽  
Zhijian Xu ◽  
...  
Keyword(s):  
Natural Products ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Underlying Mechanism ◽  
Anticancer Activities ◽  
Large B Cell Lymphoma ◽  
Large B Cell

Abstract Diffuse large B-cell lymphoma (DLBCL) is the most common category and disease entity of non-Hodgkin lymphoma. Osalmide and pterostilbene are natural products with anticancer activities via different mechanism. In this study, using a new synthetic strategy for the two natural products, we obtained the compound DCZ0801, which was previously found to have anti-multiple myeloma activity. We performed both in vitro and in vivo assays to investigate its bioactivity and explore its underlying mechanism against DLBCL cells. The results showed that DCZ0801 treatment gave rise to a dose- and time-dependent inhibition of cell viability as determined by CCK-8 assay and flow cytometry assay. Western blot analysis results showed that the expression of caspase-3, caspase-8, caspase-9 and Bax was increased, while BCL-2 and BCL-XL levels were decreased, which suggested that DCZ0801 inhibited cell proliferation and promoted intrinsic apoptosis. In addition, DCZ0801 induced G0/G1 phase arrest by downregulating the protein expression levels of CDK4, CDK6 and cyclin D1. Furthermore, DCZ0801 exerted an anti-tumor effect by down-regulating the expressions of p-PI3K and p-AKT. There also existed a trend that the expression of p-JNK and p-P38 was restrained. Intraperitoneal injection of DCZ0801 suppressed tumor development in xenograft mouse models. The preliminary metabolic study showed that DCZ0801 displayed a rapid metabolism within 30 min. These results demonstrated that DCZ0801 may be a new potential anti-DLBCL agent in DLBCL therapy.

scholarly journals Repurposing dasatinib for diffuse large B cell lymphoma

2019 ◽  
Vol 116 (34) ◽  
pp. 16981-16986 ◽  
Author(s):  
Claudio Scuoppo ◽  
Jiguang Wang ◽  
Mirjana Persaud ◽  
Sandeep K. Mittan ◽  
Katia Basso ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lines ◽  
Kinase Inhibitor ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Multikinase Inhibitor ◽  
Large B Cell Lymphoma ◽  
Large B Cell

To repurpose compounds for diffuse large B cell lymphoma (DLBCL), we screened a library of drugs and other targeted compounds approved by the US Food and Drug Administration on 9 cell lines and validated the results on a panel of 32 genetically characterized DLBCL cell lines. Dasatinib, a multikinase inhibitor, was effective against 50% of DLBCL cell lines, as well as against in vivo xenografts. Dasatinib was more broadly active than the Bruton kinase inhibitor ibrutinib and overcame ibrutinib resistance. Tumors exhibiting dasatinib resistance were commonly characterized by activation of the PI3K pathway and loss of PTEN expression as a specific biomarker. PI3K suppression by mTORC2 inhibition synergized with dasatinib and abolished resistance in vitro and in vivo. These results provide a proof of concept for the repurposing approach in DLBCL, and point to dasatinib as an attractive strategy for further clinical development in lymphomas.

scholarly journals P09.01 Adoptive cell therapy of hematological malignancies using cytokine-induced killer cells retargeted with monoclonal antibodies

2020 ◽  
Vol 8 (Suppl 2) ◽  
pp. A51.2-A52
Author(s):  
A Dalla Pietà ◽  
E Cappuzzello ◽  
P Palmerini ◽  
R Sommaggio ◽  
G Astori ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Cell Therapy ◽  
B Cell ◽  
Cell Lymphoma ◽  
Adoptive Cell Therapy ◽  
B Cell Lymphoma ◽  
Clinical Grade ◽  
Cik Cells

BackgroundCytokine-Induced Killer (CIK) cells are a population of effector cells that represents a promising tool for adoptive cell therapy. They are easily expandable ex-vivo, safe, and exert cytotoxicity against a broad range of tumor histotypes.1 We recently reported that they have a relevant expression of FcγRIIIa (CD16a), which can be exploited in combination with clinical-grade monoclonal antibodies (mAbs) to redirect their cytotoxicity in an antigen-specific manner, to improve their antitumor activity.2 Indeed, the engagement of CD16a on CIK cells leads to a potent antibody-dependent cell-mediated cytotoxicity (ADCC) against ovarian cancer both in vitro and in vivo. Based on this observation, we investigated whether CIK cells can be specifically retargeted against B-cell malignancies by combination with anti-CD20 mAbs, namely Rituximab® (RTX) and Obinutuzumab® (OBI).Materials and MethodsCIK cells were obtained from peripheral blood mononuclear cells of healthy donors, and stimulated in vitro with IFN-γ, CD3 mAb and IL-2 for 14 days; fresh IL-2 was provided every 3–4 days. CIK cell phenotype was analyzed by multicolor flow cytometry; cytotoxic activity was assessed by calcein AM-release assay against B-cell lines, primary samples and patient-derived xenografts (PDX) obtained from B-cell lymphoma patients after written informed consent.ResultsThe combination with both RTX and OBI significantly increased specific CIK cells lysis against several CD20-expressing lymphoma B cell lines, primary tumors from B-cell lymphoma patients and an established PDX, compared to the combination with a control mAb (cetuximab, CTX). NK-depletion demonstrated that the mAb-mediated cytotoxicity is accountable to the CIK cells fraction within the bulk population since no difference in the lytic activity was detectd in the absence of NK cells. In addition, these results are further supported by in vivo preliminary experiments where the treatment with CIK cells in combination with OBI extensively reduced the growth of PDX and increased mice survival, compared to CIK cells or OBI administered alone.ConclusionsHere we proved that CIK cells can be retargeted with clinical-grade mAbs against CD20-expressing lymphomas. These data indicate that the combination of CIK cells with mAbs can represent a novel approach for the treatment of haematological malignancies.ReferencesFranceschetti M, Pievani A, Borleri G, Vago L, Fleischhauer K, Golay J, et al. Cytokine-induced killer cells are terminally differentiated activated CD8 cytotoxic T-EMRA lymphocytes. Exp Hematol 2009;37:616–28.Cappuzzello E, Tosi A, Zanovello P, Sommaggio R, Rosato A. Retargeting cytokine-induced killer cell activity by CD16 engagement with clinical-grade antibodies. Oncoimmunology 2016 Aug;5(8):e1199311.The research leading to these results has received funding from Fondazione AIRC under IG 2018 - ID. 21354 project - P.I. Rosato AntonioDisclosure InformationA. Dalla Pietà: None. E. Cappuzzello: None. P. Palmerini: None. R. Sommaggio: None. G. Astori: None. K. Chieregato: None. O. Perbellini: None. M. Tisi: None. C. Visco: None. M. Ruggeri: None. A. Rosato: None.

scholarly journals 11C-Labeled Pictilisib (GDC-0941) as a Molecular Tracer Targeting Phosphatidylinositol 3-Kinase (PI3K) for Breast Cancer Imaging

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Na Han ◽  
Yaqun Jiang ◽  
Yongkang Gai ◽  
Qingyao Liu ◽  
Lujie Yuan ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Pik3ca Mutation ◽  
Phosphatidylinositol 3 Kinase ◽  
Pet Tracer ◽  
Positron Emission ◽  
Pet Scans ◽  
Mcf 7 ◽  
Phosphatidylinositol 3

Pictilisib (GDC-0941) is an inhibitor of phosphatidylinositol 3-kinase (PI3K), part of a signaling cascade involved in breast cancer development. The purpose of this study was to evaluate the pharmacokinetics of pictilisib noninvasively by radiolabeling it with 11C and to assess the usability of the resulting [11C]-pictilisib as a positron-emission tomography (PET) tracer to screen for pictilisib-sensitive tumors. In this study, pictilisib was radiolabeled with [11C]-methyl iodide to obtain 11C-methylated pictilisib ([11C]-pictilisib) using an automated synthesis module with a high radiolabeling yield. Considerably higher uptake ratios were observed in MCF-7 (PIK3CA mutation, pictilisib-sensitive) cells than those in MDA-MB-231 (PIK3CA wild-type, pictilisib-insensitive) cells at all evaluated time points, indicating good in vitro binding of [11C]-pictilisib. Dynamic micro-PET scans in mice and biodistribution results showed that [11C]-pictilisib was mainly excreted via the hepatobiliary tract into the intestines. MCF-7 xenografts could be clearly visualized on the static micro-PET scans, while MDA-MB-231 tumors could not. Biodistribution results of two xenograft models showed significantly higher uptake and tumor-to-muscle ratios in the MCF-7 xenografts than those in MDA-MB-231 xenografts, exhibiting high in vivo targeting specificity. In conclusion, [11C]-pictilisib was first successfully prepared, and it exhibited good potential to identify pictilisib-sensitive tumors noninvasively, which may have a great impact in the treatment of cancers with an overactive PI3K/Akt/mTOR signal pathway. However, the high activity in hepatobiliary system and intestines needs to be addressed.

scholarly journals Establishment of B-Cell Lymphoma Cell Lines Persistently Infected with Hepatitis C Virus In Vivo and In Vitro: the Apoptotic Effects of Virus Infection

2003 ◽  
Vol 77 (3) ◽  
pp. 2134-2146 ◽  
Author(s):  
Vicky M.-H. Sung ◽  
Shigetaka Shimodaira ◽  
Alison L. Doughty ◽  
Gaston R. Picchio ◽  
Huong Can ◽  
...  
Keyword(s):  
Hepatitis C ◽  
B Cell ◽  
Cell Line ◽  
Cell Lines ◽  
Culture System ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Hcv Rna

ABSTRACT Hepatitis C virus (HCV) is a major cause of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. Studies of HCV replication and pathogenesis have so far been hampered by the lack of an efficient tissue culture system for propagating HCV in vitro. Although HCV is primarily a hepatotropic virus, an increasing body of evidence suggests that HCV also replicates in extrahepatic tissues in natural infection. In this study, we established a B-cell line (SB) from an HCV-infected non-Hodgkin's B-cell lymphoma. HCV RNA and proteins were detectable by RNase protection assay and immunoblotting. The cell line continuously produces infectious HCV virions in culture. The virus particles produced from the culture had a buoyant density of 1.13 to 1.15 g/ml in sucrose and could infect primary human hepatocytes, peripheral blood mononuclear cells (PBMCs), and an established B-cell line (Raji cells) in vitro. The virus from SB cells belongs to genotype 2b. Single-stranded conformational polymorphism and sequence analysis of the viral RNA quasispecies indicated that the virus present in SB cells most likely originated from the patient's spleen and had an HCV RNA quasispecies pattern distinct from that in the serum. The virus production from the infected primary hepatocytes showed cyclic variations. In addition, we have succeeded in establishing several Epstein-Barr virus-immortalized B-cell lines from PBMCs of HCV-positive patients. Two of these cell lines are positive for HCV RNA as detected by reverse transcriptase PCR and for the nonstructural protein NS3 by immunofluorescence staining. These observations unequivocally establish that HCV infects B cells in vivo and in vitro. HCV-infected cell lines show significantly enhanced apoptosis. These B-cell lines provide a reproducible cell culture system for studying the complete replication cycle and biology of HCV infections.

scholarly journals Antigen-stimulated Activation of Phospholipase D1b by Rac1, ARF6, and PKCα in RBL-2H3 Cells

2002 ◽  
Vol 13 (4) ◽  
pp. 1252-1262 ◽  
Author(s):  
Dale J. Powner ◽  
Matthew N. Hodgkin ◽  
Michael J.O. Wakelam
Keyword(s):  
Plasma Membrane ◽  
Cell Types ◽  
Enzyme Activation ◽  
Dominant Negative ◽  
Ph Domain ◽  
Phosphatidylinositol 3 Kinase ◽  
Stimulation Of ◽  
Phosphatidylinositol 3

Phospholipase D (PLD) activity can be detected in response to many agonists in most cell types; however, the pathway from receptor occupation to enzyme activation remains unclear. In vitro PLD1b activity is phosphatidylinositol 4,5-bisphosphate dependent via an N-terminal PH domain and is stimulated by Rho, ARF, and PKC family proteins, combinations of which cooperatively increase this activity. Here we provide the first evidence for the in vivo regulation of PLD1b at the molecular level. Antigen stimulation of RBL-2H3 cells induces the colocalization of PLD1b with Rac1, ARF6, and PKCα at the plasma membrane in actin-rich structures, simultaneously with cooperatively increasing PLD activity. Activation is both specific and direct because dominant negative mutants of Rac1 and ARF6 inhibit stimulated PLD activity, and surface plasmon resonance reveals that the regulatory proteins bind directly and independently to PLD1b. This also indicates that PLD1b can concurrently interact with a member from each regulator family. Our results show that in contrast to PLD1b's translocation to the plasma membrane, PLD activation is phosphatidylinositol 3-kinase dependent. Therefore, because inactive, dominant negative GTPases do not activate PLD1b, we propose that activation results from phosphatidylinositol 3-kinase–dependent stimulation of Rac1, ARF6, and PKCα.

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