Obinutuzumab (GA101) More Potently Engages Phagocytic-Lineage Cells Resulting In Enhanced Monocyte and Macrophage Activity When Compared To Rituximab and Ofatumumab

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5136-5136 ◽  
Author(s):  
Sylvia Herter ◽  
Christian Klein ◽  
Pablo Umana ◽  
Marina Bacac
Keyword(s):  
Cell Death ◽  
Tumor Cell ◽  
Nk Cell ◽  
Antibody Activity ◽  
Type I ◽  
Type Ii ◽  
Therapeutic Antibodies ◽  
Tumor Cell Death ◽  
Macrophage Activity ◽  
Anti Cd20

Abstract Therapeutic antibodies possess several clinically relevant mechanisms of action including cell death induction, perturbation of tumor cell signaling, activation of complement dependent cytotoxicity (CDC), antibody dependent cellular cytotoxicity (ADCC), antibody dependent cellular phagocytosis (ADCP) and induction of adaptive immunity. Obinutuzumab (GA101) is a novel humanized, glycoengineered Type II anti-CD20 monoclonal antibody engineered for displaying enhanced FcγRIIIa (CD16) binding affinity and characterized by stronger induction of ADCC and direct tumor cell death when compared to wild-type, Type I anti-CD20 antibodies rituximab and ofatumumab. In light of the important role of phagocytic lineage cells in the mechanism of action of therapeutic antibodies, we compared GA101, rituximab and ofatumumab for their ability to trigger FcγR-dependent monocyte and macrophage effector functions. We show that, due to glycoengineering, GA101 displays superior CD16-dependent binding to monocytes, M1 and M2c macrophages in presence of nonspecific, competing, human endogenous IgGs, a situation that more closely mimics physiological conditions. Subsequently, GA101 more strongly engages monocytes and macrophages and leads to significantly higher elimination of CD20-expressing tumor cells as shown by assays detecting total antibody activity (ADCP, ADCC and direct effects). In support of the stronger GA101 activity, higher nitric-oxide (NO) levels are also detected in supernatants of tumor/macrophage co-cultures treated with antibody. Taken together, our data show that in addition to stronger NK-cell mediated ADCC and direct cell death induction due to Type II CD20 binding, GA101 more potently engages phagocytic-lineage cells resulting in enhanced monocyte and macrophage activity under conditions that more closely resemble physiological settings. Disclosures: Herter: Roche: Employment. Klein:Roche Glycart AG: Employment. Umana:Roche: Employment, Equity Ownership. Bacac:Roche: Employment.

scholarly journals Ionizing radiation improves RIG-I mediated immunotherapy through enhanced p53 activation in malignant melanoma

2021 ◽  
Author(s):  
Silke Lambing ◽  
Stefan Holdenrieder ◽  
Patrick Müller ◽  
Christian Hagen ◽  
Stephan Garbe ◽  
...  
Keyword(s):  
Cell Death ◽  
Malignant Melanoma ◽  
Tumor Cell ◽  
Low Dose ◽  
Immune Cell ◽  
Cytotoxic T Cell ◽  
Great Promise ◽  
Type I ◽  
Tumor Cell Death

The activation of the innate immune receptor RIG-I is a promising approach in immunooncology and currently under investigation in clinical trials. RIG-I agonists elicit a strong immune activation in both tumor and immune cells and induce both direct and indirect immune cell-mediated tumor cell death which involves tumor-specific cytotoxic T-cell response and type I interferon-driven innate cytotoxic immunity. Besides RIG-I, irradiation is known to induce cytotoxic DNA damage resulting in tumor debulking followed by the induction of tumor-specific immunity. To date, it is unclear whether the molecular antitumor effects of RIG-I and irradiation are additive or even synergize. Here, we investigated the combination of RIG-I activation with radiotherapy in melanoma. We found that low dose x-ray irradiation enhanced the extent and immunogenicity of RIG-I mediated tumor cell death in human and murine melanoma cell lines and in the murine B16 melanoma model in vivo. Pathway analysis of transcriptomic data revealed a central role for p53 downstream of the combined treatment, which was corroborated using p53-/- B16 cells. In vivo, the additional effect of irradiation on immune cell activation and inhibition of tumor growth was lost in mice carrying p53-knockout B16 tumors, while the response to RIG-I stimulation in those mice was maintained. Thus, our results identify p53 as pivotal for the synergy of RIG-I with irradiation, resulting in potent induction of immunogenic tumor cell death. Consequently, low dose radiotherapy holds great promise to further improve the efficacy or RIG-I ligands especially in patients with malignant melanoma or other tumors exhibiting a functional p53 pathway.

scholarly journals Novel chemical compound SINCRO with dual function in STING ‐type I interferon and tumor cell death pathways

2018 ◽  
Vol 109 (9) ◽  
pp. 2687-2696 ◽  
Author(s):  
Yoshitaka Kimura ◽  
Hideo Negishi ◽  
Atsushi Matsuda ◽  
Nobuyasu Endo ◽  
Sho Hangai ◽  
...  
Keyword(s):  
Cell Death ◽  
Tumor Cell ◽  
Chemical Compound ◽  
Type I Interferon ◽  
Dual Function ◽  
Type I ◽  
Tumor Cell Death ◽  
Cell Death Pathways

scholarly journals Novel type II anti-CD20 monoclonal antibody (GA101) evokes homotypic adhesion and actin-dependent, lysosome-mediated cell death in B-cell malignancies

Blood ◽  
2011 ◽  
Vol 117 (17) ◽  
pp. 4519-4529 ◽  
Author(s):  
Waleed Alduaij ◽  
Andrei Ivanov ◽  
Jamie Honeychurch ◽  
Eleanor J. Cheadle ◽  
Sandeep Potluri ◽  
...  
Keyword(s):  
Cell Death ◽  
B Cell ◽  
Actin Polymerization ◽  
Type I ◽  
Type Ii ◽  
B Cell Malignancies ◽  
Therapeutic Success ◽  
Wide Range ◽  
Anti Cd20

Abstract The anti-CD20 mAb rituximab has substantially improved the clinical outcome of patients with a wide range of B-cell malignancies. However, many patients relapse or fail to respond to rituximab, and thus there is intense investigation into the development of novel anti-CD20 mAbs with improved therapeutic efficacy. Although Fc-FcγR interactions appear to underlie much of the therapeutic success with rituximab, certain type II anti-CD20 mAbs efficiently induce programmed cell death (PCD), whereas rituximab-like type I anti-CD20 mAbs do not. Here, we show that the humanized, glycoengineered anti-CD20 mAb GA101 and derivatives harboring non-glycoengineered Fc regions are type II mAb that trigger nonapoptotic PCD in a range of B-lymphoma cell lines and primary B-cell malignancies. We demonstrate that GA101-induced cell death is dependent on actin reorganization, can be abrogated by inhibitors of actin polymerization, and is independent of BCL-2 overexpression and caspase activation. GA101-induced PCD is executed by lysosomes which disperse their contents into the cytoplasm and surrounding environment. Taken together, these findings reveal that GA101 is able to potently elicit actin-dependent, lysosomal cell death, which may potentially lead to improved clearance of B-cell malignancies in vivo.

Enhanced Killing of Human B Lymphoma Targets by Combined Use of Cytokine Induced Killer (CIK) Cultures and Anti-CD20 Antibodies

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4285-4285
Author(s):  
Alice Pievani ◽  
Camilla Belussi ◽  
Christian Klein ◽  
Alessandro Rambaldi ◽  
Josée Golay ◽  
...  
Keyword(s):  
Cytotoxic Activity ◽  
Nk Cell ◽  
Type I ◽  
Type Ii ◽  
B Lymphoma ◽  
Cik Cells ◽  
High Concentrations ◽  
Anti Cd20 ◽  
Cd20 Antibodies

Abstract Abstract 4285 Cytokine induced killer (CIK) cells are immune-effector cells that can be expanded in vitro in presence of rhIL-2, starting from peripheral blood mononuclear cells stimulated by interferon-γ and anti-CD3 antibody. CIK cultures at the end of in vitro expansion contain a mean of 40–75% CD3+CD56+ CIK cells, 20–60% CD3+CD56- T cells and 1–10% CD3-CD56+ NK cells. They show MHC-unrestricted cytotoxicity towards neoplastic but not normal targets. Their ease of production in vitro and anti-tumor potential have made them suitable candidates for cell therapy programs in solid and hematopoietic tumour treatment. CIK cells have shown cytotoxic activity in vitro against hematopoietic neoplastic cells, including B Non-Hodgkin's lymphoma (B-NHL). Other biological treatments available for B-NHL are the anti-CD20 antibodies such as type I Rituximab and a new generation glycoengineered type II GA101 antibody. These antibodies are thought to act mostly through immune mediated mechanisms including phagocytosis, complement mediated cytotoxicity (CDC) and antibody dependent cellular cytotoxicity (ADCC). GA101 has reduced CDC activity compared to type I anti-CD20 antibodies such as Rituximab. In addition, GA101 was defucosylated in its Fc portion to mediate increased ADCC. We have investigated the possibility of combining adoptive immunotherapy by Cytokine Induced Killer (CIK) cells with anti-CD20 type I Rituximab and type II GA101mAb, to optimize B-NHL therapy. CIK cultures alone demonstrated significant cytotoxic activity against a panel of B-NHL cell lines or freshly isolated samples, in either an autologous or allogeneic combination (26-27% killing at 30:1 ET ratio). This natural cytotoxic activity was mainly due to the predominating CD3+CD56+ CIK population (40-75%) present in the cultures. The addition of anti-CD20 mAbs increased CIK mediated cytotoxicity versus B lymphoma target cells and major enhancement was observed with GA101 compared to Rituximab (respectively 34% versus 16% increased lysis at 10:1 E:T ratio). This enhancement was mainly due to ADCC mediated by the small NK cell fraction (1-10%) present in CIK cultures because NK depletion by CD5 immunoselection at the end of expansion did not abolish the basal natural cytotoxicity of CIK cultures but abolished the enhancement observed in presence of anti-CD20 antibodies. The activation of NK cells in CIK cultures, evaluated by CD107a mobilization, was much more effective using GA101 rather than Rituximab (respectively 28% versus 19% CD107a+, p<0.005). Furthermore, in presence of human serum, Rituximab-mediated NK cell activation was inhibited to 70%, whereas the GA101-mediated was fully maintained. This inhibition was presumably due to complement C3 deposition, since it was not observed with either heat inactivated serum or high concentrations of human immunoglobulins. Inhibition was however observed with serum plus anti-C5 antibody eculizumab, which blocks the complement cascade downstream from C3. Lack of inhibition by serum of GA101-mediated NK cell degranulation was probably due to the higher affinity binding of this mAb to CD16 and not to a lower C3 activation, because at high concentrations, GA101 is nearly as efficient as Rituximab at activating complement and C3 deposition. Finally, addition of serum to CIK and mAbs did not modify overall the target cell killing by either antibody. More importantly, the use of a partially complement and CIK resistant lymphoma cell line such as WSU-NHL showed that resistance could be reverted by combined exposure to CIK cultures and monoclonal anti-CD20 antibodies. Indeed overall lysis of WSU-NHL, even in presence of serum, was significantly increasedby both anti-CD20 antibodies, and most effectively by GA101, compared to CIK cells alone. This was due to the combined action of CDC, ADCC and CIK mediated natural killing. In conclusion, CIK cultures could be used to treat B-NHL patients in both an autologous or allogeneic setting. Furthermore Rituximab but even more so GA101, could be used in vivo to enhance CIK therapeutic activity in B-NHL. These data may open the way to possible therapeutic exploitation of combined strategies of cell mediated and antibody mediated immunotherapy protocols which make use of different mechanisms of action to try and overcome resistance. Disclosures: Klein: Roche: Employment, Equity Ownership, Patents & Royalties. Rambaldi:Roche: Honoraria. Golay:Roche: Honoraria. Introna:Roche: Honoraria.

scholarly journals The induction of immunogenic cell death by type II anti-CD20 monoclonal antibodies has mechanistic differences compared with type I rituximab

2013 ◽  
Vol 162 (6) ◽  
pp. 842-845 ◽  
Author(s):  
Eleanor J. Cheadle ◽  
Lauren Sidon ◽  
Simon J. Dovedi ◽  
Monique H.M. Melis ◽  
Waleed Alduaij ◽  
...  
Keyword(s):  
Cell Death ◽  
Monoclonal Antibodies ◽  
Immunogenic Cell Death ◽  
Type I ◽  
Type Ii ◽  
Anti Cd20

scholarly journals STEM-09. INTRINSIC INTERFERON SIGNALING REGULATES THE CELL DEATH OF GLIOBLASTOMA STEM CELLS

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii198-ii198
Author(s):  
Sabbi Khan Khan ◽  
Emmanuel Martinez-Ledesma ◽  
Sandeep Mittal ◽  
Kaitlin Gandy ◽  
Kristin Alfaro-Munoz ◽  
...  
Keyword(s):  
Cell Death ◽  
Treatment Resistance ◽  
Primary Brain Tumor ◽  
The Cancer Genome Atlas ◽  
Cytokine Signaling ◽  
Type I ◽  
Type Ii ◽  
Mechanistic Investigation ◽  
Differential Cell ◽  
Stat1 Signaling

Abstract Glioblastoma (GBM) is the most common, highly aggressive, and lethal primary brain tumor in adults. Interferon (IFN)-mediated signal transducer and activator of transcription 1 (STAT1) signaling contributes to various aspects of stemness, cell death, cytokine signaling in immune and non-immune cells. However, the role of IFN/STAT1 signaling in stemness, cell death and treatment resistance in GBM is unclear. This study aimed to investigate the cancer cell-intrinsic IFN/STAT1 signaling and its role in cell proliferation, stemness, and apoptosis. By using the metagene scores for type I and type II IFN-responsive genes, we evaluated basal IFN/STAT1 signaling in The Cancer Genome Atlas (TCGA) and in patient-derived cohorts of stem-like cells (GSCs) RNA expression datasets. In-silico analyses were further validated for the constitutive IFN signaling in a subset of GSCs using qPCR, WB and ELISA assays. We employed pharmacological activators and/or inhibitors of IFN/STAT1 signaling in GSCs to study its role in stemness and cell death. We found differential cell-intrinsic type I and type II IFN-signaling markers in GSCs and GBM tumors. High IFN-signaling is associated with mesenchymal phenotype and poor survival outcomes. Acute and chronic GSC exposure to recombinant IFNs reversibly activated both type I and II IFN-signaling in GSCs. IFN-β exposure induced apoptosis in intrinsically high IFN/STAT1-signaling GSCs, but not in the low IFN/STAT1-signaling GSCs. In summary, our findings demonstrate that GBM exhibit differential cell-intrinsic IFN-signaling, and basal IFN/STAT1 is a key factor for IFN-β-mediated cell death in GSCs. However, further mechanistic investigation of intrinsic IFN signaling in GBM, particularly in the stem cell compartment is needed.

scholarly journals Cellular cytotoxicity is a form of immunogenic cell death

2020 ◽  
Vol 8 (1) ◽  
pp. e000325 ◽  
Author(s):  
Luna Minute ◽  
Alvaro Teijeira ◽  
Alfonso R Sanchez-Paulete ◽  
Maria C Ochoa ◽  
Maite Alvarez ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Death ◽  
Dendritic Cells ◽  
Tumor Cell ◽  
Tumor Cells ◽  
Cd8 T Cells ◽  
Immunogenic Cell Death ◽  
Cell Debris ◽  
Tumor Cell Death ◽  
Mhc Multimer

BackgroundThe immune response to cancer is often conceptualized with the cancer immunity cycle. An essential step in this interpretation is that antigens released by dying tumors are presented by dendritic cells to naive or memory T cells in the tumor-draining lymph nodes. Whether tumor cell death resulting from cytotoxicity, as mediated by T cells or natural killer (NK) lymphocytes, is actually immunogenic currently remains unknown.MethodsIn this study, tumor cells were killed by antigen-specific T-cell receptor (TCR) transgenic CD8 T cells or activated NK cells. Immunogenic cell death was studied analyzing the membrane exposure of calreticulin and the release of high mobility group box 1 (HMGB1) by the dying tumor cells. Furthermore, the potential immunogenicity of the tumor cell debris was evaluated in immunocompetent mice challenged with an unrelated tumor sharing only one tumor-associated antigen and by class I major histocompatibility complex (MHC)-multimer stainings. Mice deficient inBatf3,Ifnar1andSting1were used to study mechanistic requirements.ResultsWe observe in cocultures of tumor cells and effector cytotoxic cells, the presence of markers of immunogenic cell death such as calreticulin exposure and soluble HMGB1 protein. Ovalbumin (OVA)-transfected MC38 colon cancer cells, exogenously pulsed to present the gp100 epitope are killed in culture by mouse gp100-specific TCR transgenic CD8 T cells. Immunization of mice with the resulting destroyed cells induces epitope spreading as observed by detection of OVA-specific T cells by MHC multimer staining and rejection of OVA+EG7 lymphoma cells. Similar results were observed in mice immunized with cell debris generated by NK-cell mediated cytotoxicity. Mice deficient inBatf3-dependent dendritic cells (conventional dendritic cells type 1, cDC1) fail to develop an anti-OVA response when immunized with tumor cells killed by cytotoxic lymphocytes. In line with this, cultured cDC1 dendritic cells uptake and can readily cross-present antigen from cytotoxicity-killed tumor cells to cognate CD8+T lymphocytes.ConclusionThese results support that an ongoing cytotoxic antitumor immune response can lead to immunogenic tumor cell death.

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