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Pablo Umana - One of the best experts on this subject based on the ideXlab platform.
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modulation of therapeutic antibody effector functions by glycosylation engineering influence of golgi Enzyme Localization domain and co expression of heterologous β1 4 n acetylglucosaminyltransferase iii and golgi α mannosidase ii
Biotechnology and Bioengineering, 2006Co-Authors: Claudia Ferrara, Peter Brunker, Tobias Suter, Samuel Moser, Ursula Puntener, Pablo UmanaAbstract:The effector functions elicited by IgG antibodies strongly depend on the carbohydrate moiety linked to the Fc region of the protein. Therefore several approaches have been developed to rationally manipulate these glycans and improve the biological functions of the antibody. Overexpression of recombinant beta1,4-N-acetylglucosaminyltransferase III (GnT-III) in production cell lines leads to antibodies enriched in bisected oligosaccharides. Moreover, GnT-III overexpression leads to increases in non-fucosylated and hybrid oligosaccharides. Such antibody glycovariants have increased antibody-dependent cellular cytotoxicity (ADCC). To explore a further variable besides overexpression of GnT-III, we exchanged the Localization domain of GnT-III with that of other Golgi-resident Enzymes. Our results indicate that chimeric GnT-III can compete even more efficiently against the endogenous core alpha1,6-fucosyltransferase (alpha1,6-FucT) and Golgi alpha-mannosidase II (ManII) leading to higher proportions of bisected non-fucosylated hybrid glycans ("Glyco-1" antibody). The co-expression of GnT-III and ManII led to a similar degree of non-fucosylation as that obtained for Glyco-1, but the majority of the oligosaccharides linked to this antibody ("Glyco-2") are of the complex type. These glycovariants feature strongly increased ADCC activity compared to the unmodified antibody, while Glyco-1 (hybrid-rich) features reduced complement-dependent cytotoxicity (CDC) compared to Glyco-2 or unmodified antibody. We show that apart from GnT-III overexpression, engineering of GnT-III Localization is a versatile tool to modulate the biological activities of antibodies relevant for their therapeutic application.
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modulation of therapeutic antibody effector functions by glycosylation engineering influence of golgi Enzyme Localization domain and co expression of heterologous β1 4 n acetylglucosaminyltransferase iii and golgi α mannosidase ii
Biotechnology and Bioengineering, 2006Co-Authors: Claudia Ferrara, Peter Brunker, Tobias Suter, Samuel Moser, Ursula Puntener, Pablo UmanaAbstract:The effector functions elicited by IgG antibodies strongly depend on the carbohydrate moiety linked to the Fc region of the protein. Therefore several approaches have been developed to rationally manipulate these glycans and improve the biological functions of the antibody. Overexpression of recombinant β1,4-N-acetylglucosaminyltransferase III (GnT-III) in production cell lines leads to antibodies enriched in bisected oligosaccharides. Moreover, GnT-III overexpression leads to increases in non-fucosylated and hybrid oligosaccharides. Such antibody glycovariants have increased antibody-dependent cellular cytotoxicity (ADCC). To explore a further variable besides overexpression of GnT-III, we exchanged the Localization domain of GnT-III with that of other Golgi-resident Enzymes. Our results indicate that chimeric GnT-III can compete even more efficiently against the endogenous core α1,6-fucosyltransferase (α1,6-FucT) and Golgi α-mannosidase II (ManII) leading to higher proportions of bisected non-fucosylated hybrid glycans (“Glyco-1” antibody). The co-expression of GnT-III and ManII led to a similar degree of non-fucosylation as that obtained for Glyco-1, but the majority of the oligosaccharides linked to this antibody (“Glyco-2”) are of the complex type. These glycovariants feature strongly increased ADCC activity compared to the unmodified antibody, while Glyco-1 (hybrid-rich) features reduced complement-dependent cytotoxicity (CDC) compared to Glyco-2 or unmodified antibody. We show that apart from GnT-III overexpression, engineering of GnT-III Localization is a versatile tool to modulate the biological activities of antibodies relevant for their therapeutic application. © 2006 Wiley Periodicals, Inc.
Claudia Ferrara - One of the best experts on this subject based on the ideXlab platform.
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modulation of therapeutic antibody effector functions by glycosylation engineering influence of golgi Enzyme Localization domain and co expression of heterologous β1 4 n acetylglucosaminyltransferase iii and golgi α mannosidase ii
Biotechnology and Bioengineering, 2006Co-Authors: Claudia Ferrara, Peter Brunker, Tobias Suter, Samuel Moser, Ursula Puntener, Pablo UmanaAbstract:The effector functions elicited by IgG antibodies strongly depend on the carbohydrate moiety linked to the Fc region of the protein. Therefore several approaches have been developed to rationally manipulate these glycans and improve the biological functions of the antibody. Overexpression of recombinant beta1,4-N-acetylglucosaminyltransferase III (GnT-III) in production cell lines leads to antibodies enriched in bisected oligosaccharides. Moreover, GnT-III overexpression leads to increases in non-fucosylated and hybrid oligosaccharides. Such antibody glycovariants have increased antibody-dependent cellular cytotoxicity (ADCC). To explore a further variable besides overexpression of GnT-III, we exchanged the Localization domain of GnT-III with that of other Golgi-resident Enzymes. Our results indicate that chimeric GnT-III can compete even more efficiently against the endogenous core alpha1,6-fucosyltransferase (alpha1,6-FucT) and Golgi alpha-mannosidase II (ManII) leading to higher proportions of bisected non-fucosylated hybrid glycans ("Glyco-1" antibody). The co-expression of GnT-III and ManII led to a similar degree of non-fucosylation as that obtained for Glyco-1, but the majority of the oligosaccharides linked to this antibody ("Glyco-2") are of the complex type. These glycovariants feature strongly increased ADCC activity compared to the unmodified antibody, while Glyco-1 (hybrid-rich) features reduced complement-dependent cytotoxicity (CDC) compared to Glyco-2 or unmodified antibody. We show that apart from GnT-III overexpression, engineering of GnT-III Localization is a versatile tool to modulate the biological activities of antibodies relevant for their therapeutic application.
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modulation of therapeutic antibody effector functions by glycosylation engineering influence of golgi Enzyme Localization domain and co expression of heterologous β1 4 n acetylglucosaminyltransferase iii and golgi α mannosidase ii
Biotechnology and Bioengineering, 2006Co-Authors: Claudia Ferrara, Peter Brunker, Tobias Suter, Samuel Moser, Ursula Puntener, Pablo UmanaAbstract:The effector functions elicited by IgG antibodies strongly depend on the carbohydrate moiety linked to the Fc region of the protein. Therefore several approaches have been developed to rationally manipulate these glycans and improve the biological functions of the antibody. Overexpression of recombinant β1,4-N-acetylglucosaminyltransferase III (GnT-III) in production cell lines leads to antibodies enriched in bisected oligosaccharides. Moreover, GnT-III overexpression leads to increases in non-fucosylated and hybrid oligosaccharides. Such antibody glycovariants have increased antibody-dependent cellular cytotoxicity (ADCC). To explore a further variable besides overexpression of GnT-III, we exchanged the Localization domain of GnT-III with that of other Golgi-resident Enzymes. Our results indicate that chimeric GnT-III can compete even more efficiently against the endogenous core α1,6-fucosyltransferase (α1,6-FucT) and Golgi α-mannosidase II (ManII) leading to higher proportions of bisected non-fucosylated hybrid glycans (“Glyco-1” antibody). The co-expression of GnT-III and ManII led to a similar degree of non-fucosylation as that obtained for Glyco-1, but the majority of the oligosaccharides linked to this antibody (“Glyco-2”) are of the complex type. These glycovariants feature strongly increased ADCC activity compared to the unmodified antibody, while Glyco-1 (hybrid-rich) features reduced complement-dependent cytotoxicity (CDC) compared to Glyco-2 or unmodified antibody. We show that apart from GnT-III overexpression, engineering of GnT-III Localization is a versatile tool to modulate the biological activities of antibodies relevant for their therapeutic application. © 2006 Wiley Periodicals, Inc.
Patricia T Bozza - One of the best experts on this subject based on the ideXlab platform.
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leukocyte lipid bodies regulation and function contribution to allergy and host defense
Pharmacology & Therapeutics, 2007Co-Authors: Patricia T Bozza, Rossana C N Melo, Christianne BandeirameloAbstract:Lipid bodies are lipid-rich organelles found in the cytoplasm of a variety of cells, including leukocytes. Lipid body morphology, its ability to interact with other organelles and its functions are dictated by its lipid arrangement, as well as its protein composition. Both may vary according to the cell type and with the specific lipid body biogenic stimulatory pathways. Nascent lipid bodies, which are formed in vivo in the course of a variety of immunopathological conditions, are sites of Enzyme Localization, eicosanoid production, as well as, sites for cytokine storage in inflammatory leukocytes, suggesting that lipid bodies function as inducible intracellular platforms for spatial segregation and organization of signaling leading to inflammatory mediator secretion during inflammation. The emerging role of lipid bodies as inflammatory organelles raises lipid body status to critical regulators of different inflammatory diseases, key markers of leukocyte activation and attractive targets for novel anti-inflammatory therapies.
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lipopolysaccharide induced leukocyte lipid body formation in vivo innate immunity elicited intracellular loci involved in eicosanoid metabolism
Journal of Immunology, 2002Co-Authors: Patricia Pacheco, Peter F Weller, Fernando A Bozza, Rachel N Gomes, Marcelo T Bozza, Hugo C Castrofarianeto, Patricia T BozzaAbstract:Lipid bodies are rapidly inducible, specialized cytoplasmic domains for eicosanoid-forming Enzyme Localization, which we hypothesize to have specific roles in enhanced inflammatory mediator production during pathological conditions, including sepsis. However, little is known about the origins, composition, or functions of lipid bodies in vivo. We show that lipid body numbers were increased in leukocytes from septic patients in comparison with healthy subjects. Analogously, the intrathoracic administration of LPS into mice induced a dose- and time-dependent increase in lipid body numbers. Pretreatment with anti-CD14 or anti-CD11b/CD18 mAb drastically inhibited LPS-induced lipid body formation. Moreover, LPS failed to form lipid bodies in C3H/HeJ ( TLR4 mutated) mice, demonstrating a requisite role for LPS receptors in lipid body formation. LPS-induced lipid body formation was also inhibited by the platelet-activating factor-receptor antagonists, suggesting a role for endogenous platelet-activating factor. The eicosanoid-forming Enzymes, 5-lipoxygenase and cyclooxygenase-2, were immunolocalized within experimentally induced (LPS in mice) or naturally occurring (septic patients) lipid bodies. The proinflammatory cytokine involved in the pathogenesis of sepsis, TNF-α, was also shown to colocalize within lipid bodies. Prior stimulation of leukocytes to form lipid bodies enhanced the capacity of leukocytes to produce leukotriene B 4 and PGE 2 . In conclusion, our studies indicate that lipid bodies formed after LPS stimulation and sepsis are sites for eicosanoid-forming Enzymes and cytokine Localization and may develop and function as structurally distinct, intracellular sites for paracrine eicosanoid synthesis during inflammatory conditions.
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co compartmentalization of map kinases and cytosolic phospholipase a2 at cytoplasmic arachidonate rich lipid bodies
American Journal of Pathology, 1998Co-Authors: Wengui Yu, Patricia T Bozza, D M Tzizik, J P Gray, Jessica Cassara, Ann M Dvorak, Peter F WellerAbstract:Lipid bodies are inducible lipid domains abundantly present in leukocytes engaged in inflammation. They are rich in esterified arachidonate and are also potential sites for eicosanoid-forming Enzyme Localization. It is therefore of interest to know whether arachidonate-releasing cytosolic phospholipase A 2 (cPLA 2 ) localizes at lipid bodies. Here, we present evidence that cPLA 2 and its activating protein kinases, mitogen-activated protein (MAP) kinases, co-localize at lipid bodies. U937 cells express high levels of cPLA 2 and contain numerous cytoplasmic lipid bodies. Using double-labeling immunocytochemistry we demonstrated punctate cytoplasmic Localizations of both cPLA 2 and MAP kinases in U937 cells that were perfectly concordant with fluorescent fatty-acid-labeled lipid bodies. The co-Localization of cPLA 2 and MAP kinases at lipid bodies was confirmed by subcellular fractionation and immunoblot. Lipid body fractions free of cytosol and other organelles contained significant amounts of [ 14 C]arachidonate-labeled phosphatidylcholine and cPLA 2 enzymatic activities. Immunoblotting with specific antibodies identified cPLA 2 as well as MAP kinases, including ERK1, ERK2, p85, and p38, in lipid bodies. The co-compartmentalization within arachidonate-rich lipid bodies of cPLA 2 and its potentially activating protein kinases suggests that lipid bodies may be structurally distinct intracellular sites active in extracellular ligand-induced arachidonate release and eicosanoid formation.
Christianne Bandeiramelo - One of the best experts on this subject based on the ideXlab platform.
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leukocyte lipid bodies regulation and function contribution to allergy and host defense
Pharmacology & Therapeutics, 2007Co-Authors: Patricia T Bozza, Rossana C N Melo, Christianne BandeirameloAbstract:Lipid bodies are lipid-rich organelles found in the cytoplasm of a variety of cells, including leukocytes. Lipid body morphology, its ability to interact with other organelles and its functions are dictated by its lipid arrangement, as well as its protein composition. Both may vary according to the cell type and with the specific lipid body biogenic stimulatory pathways. Nascent lipid bodies, which are formed in vivo in the course of a variety of immunopathological conditions, are sites of Enzyme Localization, eicosanoid production, as well as, sites for cytokine storage in inflammatory leukocytes, suggesting that lipid bodies function as inducible intracellular platforms for spatial segregation and organization of signaling leading to inflammatory mediator secretion during inflammation. The emerging role of lipid bodies as inflammatory organelles raises lipid body status to critical regulators of different inflammatory diseases, key markers of leukocyte activation and attractive targets for novel anti-inflammatory therapies.
Ursula Puntener - One of the best experts on this subject based on the ideXlab platform.
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modulation of therapeutic antibody effector functions by glycosylation engineering influence of golgi Enzyme Localization domain and co expression of heterologous β1 4 n acetylglucosaminyltransferase iii and golgi α mannosidase ii
Biotechnology and Bioengineering, 2006Co-Authors: Claudia Ferrara, Peter Brunker, Tobias Suter, Samuel Moser, Ursula Puntener, Pablo UmanaAbstract:The effector functions elicited by IgG antibodies strongly depend on the carbohydrate moiety linked to the Fc region of the protein. Therefore several approaches have been developed to rationally manipulate these glycans and improve the biological functions of the antibody. Overexpression of recombinant beta1,4-N-acetylglucosaminyltransferase III (GnT-III) in production cell lines leads to antibodies enriched in bisected oligosaccharides. Moreover, GnT-III overexpression leads to increases in non-fucosylated and hybrid oligosaccharides. Such antibody glycovariants have increased antibody-dependent cellular cytotoxicity (ADCC). To explore a further variable besides overexpression of GnT-III, we exchanged the Localization domain of GnT-III with that of other Golgi-resident Enzymes. Our results indicate that chimeric GnT-III can compete even more efficiently against the endogenous core alpha1,6-fucosyltransferase (alpha1,6-FucT) and Golgi alpha-mannosidase II (ManII) leading to higher proportions of bisected non-fucosylated hybrid glycans ("Glyco-1" antibody). The co-expression of GnT-III and ManII led to a similar degree of non-fucosylation as that obtained for Glyco-1, but the majority of the oligosaccharides linked to this antibody ("Glyco-2") are of the complex type. These glycovariants feature strongly increased ADCC activity compared to the unmodified antibody, while Glyco-1 (hybrid-rich) features reduced complement-dependent cytotoxicity (CDC) compared to Glyco-2 or unmodified antibody. We show that apart from GnT-III overexpression, engineering of GnT-III Localization is a versatile tool to modulate the biological activities of antibodies relevant for their therapeutic application.
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modulation of therapeutic antibody effector functions by glycosylation engineering influence of golgi Enzyme Localization domain and co expression of heterologous β1 4 n acetylglucosaminyltransferase iii and golgi α mannosidase ii
Biotechnology and Bioengineering, 2006Co-Authors: Claudia Ferrara, Peter Brunker, Tobias Suter, Samuel Moser, Ursula Puntener, Pablo UmanaAbstract:The effector functions elicited by IgG antibodies strongly depend on the carbohydrate moiety linked to the Fc region of the protein. Therefore several approaches have been developed to rationally manipulate these glycans and improve the biological functions of the antibody. Overexpression of recombinant β1,4-N-acetylglucosaminyltransferase III (GnT-III) in production cell lines leads to antibodies enriched in bisected oligosaccharides. Moreover, GnT-III overexpression leads to increases in non-fucosylated and hybrid oligosaccharides. Such antibody glycovariants have increased antibody-dependent cellular cytotoxicity (ADCC). To explore a further variable besides overexpression of GnT-III, we exchanged the Localization domain of GnT-III with that of other Golgi-resident Enzymes. Our results indicate that chimeric GnT-III can compete even more efficiently against the endogenous core α1,6-fucosyltransferase (α1,6-FucT) and Golgi α-mannosidase II (ManII) leading to higher proportions of bisected non-fucosylated hybrid glycans (“Glyco-1” antibody). The co-expression of GnT-III and ManII led to a similar degree of non-fucosylation as that obtained for Glyco-1, but the majority of the oligosaccharides linked to this antibody (“Glyco-2”) are of the complex type. These glycovariants feature strongly increased ADCC activity compared to the unmodified antibody, while Glyco-1 (hybrid-rich) features reduced complement-dependent cytotoxicity (CDC) compared to Glyco-2 or unmodified antibody. We show that apart from GnT-III overexpression, engineering of GnT-III Localization is a versatile tool to modulate the biological activities of antibodies relevant for their therapeutic application. © 2006 Wiley Periodicals, Inc.