Homo sapiens
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PROVIDER: PRJNA104067 | ENA |
REPOSITORIES: ENA
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Project description:Transcriptional responses by alpha-emitting radioimmunoconjugate Bi-213-CD20-rituximab
| PRJNA98035 | ENA
Project description:Transcriptional responses in B-cell lymphomas by the alpha-emitting radioimmunoconjugate Bi-213-CD20-rituximab
| PRJNA104065 | ENA
Project description:The possibility to deliver radioisotopes directly to tumor cells by using monoclonal antibodies has become a promising concept for the elimination of small tumor nodules or single disseminated tumor cells. The alpha-particle emitter Bismut-213 which has a high linear energy transfer and a very short path length appears to be able to kill cells by only few nuclear hits. Bismut-213 based alpha-immunotherapy treatment is currently proved in clinical trials for AML, NHL and preclinically for CLL and multiple myeloma. Because little is known about the biological effects of alpha radiation we examined the molecular effects of the alpha emitter Bismut-213 in malignant and normal B-cells. In this study, we used the cell line Karpas 422 (K422) which had been derived from a malignant B-cell Non-Hodgkin’s lymphoma as well as normal primary CD19+ B-cells which had been selected immunomagnetically from peripheral blood. For the delivery and binding of the alpha emitter we used the monoclonal antibody rituximab which targets the CD20 receptor of malignant and normal B-cells. The lymphoma cell line K422 was incubated in vitro with Bismut-213-CD20 with 100 µCi and 200 µCi respectively, for 46 min (1 half life time of the alpha emitter) or 24 h. Immunomagnetically isolated CD19+ cells from healthy donors were incubated in vitro with Bismut-213-CD20 with 100 µCi or 200 µCi, for 46 min. Gene expression profiles of irradiated cells were analysed using cDNA oligonucleotide arrays (Affymetrix Human Genome Focus Arrays) comprising 8,793 genes and compared with those from untreated control cells. Following normalization significantly altered genes were identified by using the variance stabilization normalization (VSN) method and the significance analysis of microarrays (SAM) algorithm. Differentially expressed genes were defined to exhibit a fold change of 1,4 and higher or 0,7 and lower and a q-value of 5% or lower compared to the genes of the untreated control cells. Irradiated K422 cells with 100 µCi showed 42 differential expressed genes after 46 minutes (one half life time) and 451 deregulated genes after exposure of 24 hours. A similar response could be observed at an exposure of 200 µCi (50 differentially expressed genes after 46 min; 485 genes after 24 h). Hence, a severe radiation induced response could not be detected before 24 hours. 200 µCi irradiated CD19+ cells showed 67 differentially expressed genes after 46 minutes compared to the untreated CD19+ cells (for 100 µCi: 42 differentially expressed genes). By performing cluster analyses the irradiated cells exhibited a distinct homogenous molecular phenotype in comparison to not irradiated cells. Alpha irradiated cells showed transcriptional activation of DNA repair genes. Furthermore, both examined cell types showed up-regulated genes which are known to play a role in general and oxidative stress response. Surprisingly, up-regulation of interferon-stimulated genes in K422 cells could be observed. Because of the transcriptional activation of genes regulating the G1-S- cell cycle transition the supposition came up that the irradiated K422 do progress into the S-phase of the cell cycle and arrest in the S- or G2/M-phase. In contrast to the K422 cells the primary CD19+ B-cells revealed transcriptional activation of cell cycle inhibitors which is an already known reaction following ionizing radiation but no apoptosis associated genes could be observed. In spite of the massive cytotoxic damages which are assumed referring to the gene expression data the K422 cells appear to be able to activate several defense mechanisms in a precise manner. Hence, the malignant K422 cells exhibit a larger amount as well as more various responses than the CD19+ cells. This might be due to the tumorigenic transformation of the malignant cells that exhibit probably more effective defense strategies than the CD19+ cells. The balance between the therapeutic effect of alpha radiation on malignant cells and the damaging effect on normal cells is an essential issue and it is of important clinical relevance concerning radioimmunotherapy with alpha radiation. Keywords: time course and dose dependency
2007-07-01 | GSE7292 | GEO
Project description:The possibility to deliver radioisotopes directly to tumor cells by using monoclonal antibodies has become a promising concept for the elimination of small tumor nodules or single disseminated tumor cells. The alpha-particle emitter Bismut-213 which has a high linear energy transfer and a very short path length appears to be able to kill cells by only few nuclear hits. Bismut-213 based alpha-immunotherapy treatment is currently proved in clinical trials for AML, NHL and preclinically for CLL and multiple myeloma. Because little is known about the biological effects of alpha radiation we examined the molecular effects of the alpha emitter Bismut-213 in malignant and normal B-cells. In this study, we used the cell line Karpas 422 (K422) which had been derived from a malignant B-cell Non-Hodgkin’s lymphoma as well as normal primary CD19+ B-cells which had been selected immunomagnetically from peripheral blood. For the delivery and binding of the alpha emitter we used the monoclonal antibody rituximab which targets the CD20 receptor of malignant and normal B-cells. The lymphoma cell line K422 was incubated in vitro with Bismut-213-CD20 with 100 µCi and 200 µCi respectively, for 46 min (1 half life time of the alpha emitter) or 24 h. Immunomagnetically isolated CD19+ cells from healthy donors were incubated in vitro with Bismut-213-CD20 with 100 µCi or 200 µCi, for 46 min. Gene expression profiles of irradiated cells were analysed using cDNA oligonucleotide arrays (Affymetrix Human Genome Focus Arrays) comprising 8,793 genes and compared with those from untreated control cells. Following normalization significantly altered genes were identified by using the variance stabilization normalization (VSN) method and the significance analysis of microarrays (SAM) algorithm. Differentially expressed genes were defined to exhibit a fold change of 1,4 and higher or 0,7 and lower and a q-value of 5% or lower compared to the genes of the untreated control cells. Irradiated K422 cells with 100 µCi showed 42 differential expressed genes after 46 minutes (one half life time) and 451 deregulated genes after exposure of 24 hours. A similar response could be observed at an exposure of 200 µCi (50 differentially expressed genes after 46 min; 485 genes after 24 h). Hence, a severe radiation induced response could not be detected before 24 hours. 200 µCi irradiated CD19+ cells showed 67 differentially expressed genes after 46 minutes compared to the untreated CD19+ cells (for 100 µCi: 42 differentially expressed genes). By performing cluster analyses the irradiated cells exhibited a distinct homogenous molecular phenotype in comparison to not irradiated cells. Alpha irradiated cells showed transcriptional activation of DNA repair genes. Furthermore, both examined cell types showed up-regulated genes which are known to play a role in general and oxidative stress response. Surprisingly, up-regulation of interferon-stimulated genes in K422 cells could be observed. Because of the transcriptional activation of genes regulating the G1-S- cell cycle transition the supposition came up that the irradiated K422 do progress into the S-phase of the cell cycle and arrest in the S- or G2/M-phase. In contrast to the K422 cells the primary CD19+ B-cells revealed transcriptional activation of cell cycle inhibitors which is an already known reaction following ionizing radiation but no apoptosis associated genes could be observed. In spite of the massive cytotoxic damages which are assumed referring to the gene expression data the K422 cells appear to be able to activate several defense mechanisms in a precise manner. Hence, the malignant K422 cells exhibit a larger amount as well as more various responses than the CD19+ cells. This might be due to the tumorigenic transformation of the malignant cells that exhibit probably more effective defense strategies than the CD19+ cells. The balance between the therapeutic effect of alpha radiation on malignant cells and the damaging effect on normal cells is an essential issue and it is of important clinical relevance concerning radioimmunotherapy with alpha radiation. Keywords: time course and dose dependency
2007-07-01 | GSE7291 | GEO
Project description:B cell identity and function is dependent on the proteins residing in the B cell surfaceome and their respective nanoscale organization. Here we show that CD20 is a gatekeeper for B cell identity and function due to its control of the functional nanoscale organization of receptors within the B surfaceome and the resting state of B lymphocytes. CRISPR/Cas-based ablation of CD20 in Ramos B cells revealed that IgM class B cell antigen receptor (IgM-BCR) and the co-receptor CD19 are functionally interlinked in the absence of CD20. The resulting IgM-BCR/CD19 signalling synapse leads to transient B cell activation and the loss of B cell identity. Re-expression of CD20 involving an aptamer-controlled riboswitch restores resting-state B cell nanoscale organization and function. Treatment of naive human B cells with the anti-CD20 antibody Rituximab leads to transient B cell activation and formation of transiently activated IgM-BCR/CD19 signalling synapses providing new insights into the molecular mode of action of Rituximab. The loss of B cell identity due to CD20-deficiency is accompanied by a PAX5 to BLIMP-1 transcriptional switch and increased plasma cell development. This cellular B cell differentiation towards plasma B cells is mechanistically accompanied by a metabolic shift towards oxidative phosphorylation.
2020-06-18 | MSV000085605 | MassIVE
Project description:NK-92 a continuously growing cell line bioengineered to express human anti-CD19 chimeric antigen receptor (CAR) CD19.TaNK recognizing CD19+ B cells represents as potential “off the shelf” therapy candidate for B cell malignancies. The goal of this study was to establish the mechanistic rationale for CD19.TaNK therapy in B-cell NHL (bNHL) and to determine the therapeutic potency in vitro against a host of bNHL cell lines, patient derived primary cell lines (including anti-CD20 antibody resistant cell lines), and in in vivo mouse models. We utilized bNHL cell lines SU-DHL10, SU-DHL4, SU-DHL2 (DLBCL), HF-1 (follicular) and Raji (Burkitt’s) and Rituximab- (RR) and obinutuzumab (OR)-resistant bNHL cell lines (SU-DHL2, SU-DHL4, SU-DHL10), and patient derived primary cells (EL-5 and KSC) were investigated the cytolytic activity of CD19.TaNK. We observed significantly increased CD19.TaNK mediated cytolytic activity at E:T ratios (1:1-10:1) via LDH release in all bNHL cell lines and CD20 resistant bHNL cells. Further, the dynamic efficacy of CD19.TaNK determined using droplet based single cell microfluidics analysis of cell interactions (1:1) between CD19.TaNK and anti-CD20 sensitive or resistant bNHL cell showed that vast majority of the cells were killed by single contact >80% (SU-DHL 4, SUD-HL 4 -OR), 40% (SU-DHL10-RR), >60% (SU-DHL10, SUD-HL-10-OR) and 40% (SU-DHL10-RR) within first 40 minutes, while the remainder were killed through events requiring multiple contacts. Thus, suggesting that CD19.TaNK indiscriminately kills both anti-CD20 sensitive and resistant cells. Global transcriptome analysis performed using flow sorted bNHL co-cultured with CD19.TaNK at 1:1 ratio for two hours, revealed conserved activation of IFNγ signaling, execution of apoptosis, ligand binding, immunoregulatory or chemokine signaling pathways in these bNHL cells. Using proximity extension assay based 92-plex cytokine panel we observed increased secretion of various cytokines, granzymes and decreased secretions of ADA, HO-1, CD5, CD28, CD70, CD244, IFN and TNF consistently with anti-CD20 sensitive and resistant cells. Altogether these results demonstrate that CD19.TaNK inflicts mechanistically conserved killing activity against different bNHL cell lines, including in anti-CD20 refractory bNHL. Finally, in SCID mice experiments we observed marked reduction in the volume of SU-DHL10 derived tumor xenografts with infusion of CD19.TaNK compared to control. Overall, we observed potent anti-lymphoma activity with CD19.TaNK involving biologically conserved mechanisms indicating that CD19.TaNK could be equally active under untreated or refractory bNHL in the clinical settings.
2018-12-01 | GSE120316 | GEO
Project description:NK-92 a continuously growing cell line bioengineered to express human anti-CD19 chimeric antigen receptor (CAR) CD19.TaNK recognizing CD19+ B cells represents as potential “off the shelf” therapy candidate for B cell malignancies. The goal of this study was to establish the mechanistic rationale for CD19.TaNK therapy in B-cell NHL (bNHL) and to determine the therapeutic potency in vitro against a host of bNHL cell lines, patient derived primary cell lines (including anti-CD20 antibody resistant cell lines), and in in vivo mouse models. We utilized bNHL cell lines SU-DHL10, SU-DHL4, SU-DHL2 (DLBCL), HF-1 (follicular) and Raji (Burkitt’s) and Rituximab- (RR) and obinutuzumab (OR)-resistant bNHL cell lines (SU-DHL2, SU-DHL4, SU-DHL10), and patient derived primary cells (EL-5 and KSC) were investigated the cytolytic activity of CD19.TaNK. We observed significantly increased CD19.TaNK mediated cytolytic activity at E:T ratios (1:1-10:1) via LDH release in all bNHL cell lines and CD20 resistant bHNL cells. Further, the dynamic efficacy of CD19.TaNK determined using droplet based single cell microfluidics analysis of cell interactions (1:1) between CD19.TaNK and anti-CD20 sensitive or resistant bNHL cell showed that vast majority of the cells were killed by single contact >80% (SU-DHL 4, SUD-HL 4 -OR), 40% (SU-DHL10-RR), >60% (SU-DHL10, SUD-HL-10-OR) and 40% (SU-DHL10-RR) within first 40 minutes, while the remainder were killed through events requiring multiple contacts. Thus, suggesting that CD19.TaNK indiscriminately kills both anti-CD20 sensitive and resistant cells. Global transcriptome analysis performed using flow sorted bNHL co-cultured with CD19.TaNK at 1:1 ratio for two hours, revealed conserved activation of IFNγ signaling, execution of apoptosis, ligand binding, immunoregulatory or chemokine signaling pathways in these bNHL cells. Using proximity extension assay based 92-plex cytokine panel we observed increased secretion of various cytokines, granzymes and decreased secretions of ADA, HO-1, CD5, CD28, CD70, CD244, IFN and TNF consistently with anti-CD20 sensitive and resistant cells. Altogether these results demonstrate that CD19.TaNK inflicts mechanistically conserved killing activity against different bNHL cell lines, including in anti-CD20 refractory bNHL. Finally, in SCID mice experiments we observed marked reduction in the volume of SU-DHL10 derived tumor xenografts with infusion of CD19.TaNK compared to control. Overall, we observed potent anti-lymphoma activity with CD19.TaNK involving biologically conserved mechanisms indicating that CD19.TaNK could be equally active under untreated or refractory bNHL in the clinical settings.
2018-12-01 | GSE120315 | GEO
Project description:Rituximab alone or in combination with chemotherapeutics is the first-line therapy for variety of lymphoproliferative disorders including low- and high grade non-Hodgkin’s lymphomas (NHL). Although the complete response rate is quite impressive, vast majority of patient presents recurrent disease. The association between CD20 expression and clinical outcome in patients strongly suggests that reduced CD20 expression leads to inferior response to RCHOP (rituximab, cyclophosphamide, vincristine, doxorubicin and prednisone). In order to understand how loss of CD20 leads to development of RCHOP resistance, we developed rituximab resistant DOHH2 model in vivo by chronic exposure to rituximab. Characterization of several resistant in vivo xenografts revealed one model that maintained resistance to an acute dose of rituximab and demonstrated loss of CD20. Further characterization of the model demonstrated a loss of CD20 is associated with over expression of BCL2 and BIM. In vivo efficacy studies showed resistant line is insensitive to acute dose of RCHOP and treatment with an inhibitor of BCL2 (ABT199) in combination with chemotherapy resulted in better efficacy than RCHOP alone. We have identified an in vivo model of DLBCL where loss of CD20 and over expression of anti-apoptotic protein BCL2 leads to RCHOP resistance. These data suggest the addition of BCL2 inhibitor to chemotherapy might be effective in treating CD20 negative lymphomas. mRNA profiles of parental and rituximab resistant DOHH2 xenograft were generated by deep sequencing using Illumina HiSeq
2014-09-01 | E-GEOD-54201 | biostudies-arrayexpress
Project description:Rituximab alone or in combination with chemotherapeutics is the first-line therapy for variety of lymphoproliferative disorders including low- and high grade non-Hodgkin’s lymphomas (NHL). Although the complete response rate is quite impressive, vast majority of patient presents recurrent disease. The association between CD20 expression and clinical outcome in patients strongly suggests that reduced CD20 expression leads to inferior response to RCHOP (rituximab, cyclophosphamide, vincristine, doxorubicin and prednisone). In order to understand how loss of CD20 leads to development of RCHOP resistance, we developed rituximab resistant DOHH2 model in vivo by chronic exposure to rituximab. Characterization of several resistant in vivo xenografts revealed one model that maintained resistance to an acute dose of rituximab and demonstrated loss of CD20. Further characterization of the model demonstrated a loss of CD20 is associated with over expression of BCL2 and BIM. In vivo efficacy studies showed resistant line is insensitive to acute dose of RCHOP and treatment with an inhibitor of BCL2 (ABT199) in combination with chemotherapy resulted in better efficacy than RCHOP alone. We have identified an in vivo model of DLBCL where loss of CD20 and over expression of anti-apoptotic protein BCL2 leads to RCHOP resistance. These data suggest the addition of BCL2 inhibitor to chemotherapy might be effective in treating CD20 negative lymphomas.
2014-09-01 | GSE54201 | GEO
Project description:We investigated the differential regulation patterns of type I anti-CD20 monoclonal antibody (mAb) rituximab and type II obinutuzumab on a transcriptional level. Using a panel of MCL cell lines, we determined the effects of obinutuzumab and rituximab as monotherapies as well as in combination on cell viability and proliferation. Obinutuzumab induced a higher reduction in cell proliferation in each mantle cell lymphoma cell line than rituximab did. Results indicate a common pattern of expression changes after binding of anti-CD20 mAbs, but also reveal a significant difference between type I and type II treatment. Combination treatment resulted in a rituximab-like expression pattern. Many deregulated genes were associated with stress signalling, cell death, immune response and other functional clusters. Our analyses identified different and antibody-specific downstream expression patterns of obinutuzumab and rituximab, which may represent the molecular basis of the superior effect of obinutuzumab in comparison to rituximab.
2014-12-31 | GSE54169 | GEO