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1.
J Immunol ; 202(8): 2307-2319, 2019 04 15.
Article in English | MEDLINE | ID: mdl-30796180

ABSTRACT

Suboptimal immune responses to pathogens contribute to chronic infections. One way to improve immune responses is to boost Ag presentation. In this study, we investigate the potential of the tripartite motif-containing 21 (TRIM21) pathway. TRIM21 is a ubiquitously expressed cytosolic protein that recognizes the Fc region of Abs. When Abs that are bound to pathogens enter the cell as immune complexes, binding of TRIM21 to Fc initiates downstream inflammatory signaling and targets the immune complexes for proteasomal degradation. In APCs, peptides generated by proteasomes are loaded onto MHC class I molecules to stimulate CD8 T cell responses, which are crucial for effective immunity to pathogens. We hypothesized that increasing the affinity between immune complexes and TRIM21 might markedly improve CD8 T cell responses to Ags processed by the TRIM21 pathway. Using phage display technology, we engineered the human IgG1 Fc to increase its affinity for TRIM21 by 100-fold. Adenovirus immune complexes with the engineered Fc induced greater maturation of human dendritic cells (DC) than immune complexes with unmodified Fc and stimulated increased Ag-specific CD8 T cell proliferation and IFN-γ release in cocultures of DC-PBMC. Thus, by increasing the affinity between Fc and TRIM21, Ags from immune complexes undergo enhanced cross-presentation on DC, leading to greater CD8 T cell responses. Our study reveals an approach that could potentially be used in vaccines to increase cytotoxic T cell responses against Ags that are targeted or delivered by Fc-modified Abs.


Subject(s)
Antigen Presentation , Dendritic Cells/immunology , Monocytes/immunology , Ribonucleoproteins/immunology , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/immunology , Dendritic Cells/cytology , Humans , Immunoglobulin Fc Fragments/immunology , Immunoglobulin G/immunology , Monocytes/cytology , Ribonucleoproteins/genetics
2.
MAbs ; 6(3): 765-73, 2014.
Article in English | MEDLINE | ID: mdl-24671001

ABSTRACT

The pro-inflammatory cytokine interleukin (IL)-1ß is a clinical target in many conditions involving dysregulation of the immune system; therapeutics that block IL-1ß have been approved to treat diseases such as rheumatoid arthritis (RA), neonatal onset multisystem inflammatory diseases, cryopyrin-associated periodic syndromes, active systemic juvenile idiopathic arthritis. Here, we report the generation and engineering of a new fully human antibody that binds tightly to IL-1ß with a neutralization potency more than 10 times higher than that of the marketed antibody canakinumab. After affinity maturation, the derived antibody shows a>30-fold increased affinity to human IL-1ß compared with its parent antibody. This anti-human IL-1ß IgG also cross-reacts with mouse and monkey IL-1ß, hence facilitating preclinical development. In a number of mouse models, this antibody efficiently reduced or abolished signs of disease associated with IL-1ß pathology. Due to its high affinity for the cytokine and its potency both in vitro and in vivo, we propose that this novel fully human anti-IL-1ß monoclonal antibody is a promising therapeutic candidate and a potential alternative to the current therapeutic arsenal.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Interleukin-1beta/antagonists & inhibitors , Interleukin-1beta/immunology , Animals , Antibodies, Monoclonal/genetics , Antibodies, Monoclonal/therapeutic use , Antibodies, Monoclonal, Humanized , Antibodies, Neutralizing/genetics , Antibodies, Neutralizing/therapeutic use , Antibody Affinity , Arthritis, Experimental/immunology , Arthritis, Experimental/therapy , Cross Reactions , Disease Models, Animal , Epitopes/immunology , Female , Humans , Immunoglobulin G/genetics , Immunoglobulin G/immunology , Immunoglobulin G/therapeutic use , Inflammation/immunology , Inflammation/therapy , Macaca mulatta , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Inbred NOD , Mice, SCID , Multiple Myeloma/immunology , Multiple Myeloma/therapy , Peptide Library , Peritonitis/immunology , Peritonitis/therapy , Protein Engineering
3.
J Neurosci ; 30(25): 8457-67, 2010 Jun 23.
Article in English | MEDLINE | ID: mdl-20573893

ABSTRACT

Cyclin-dependent kinase 5 (Cdk5)-mediated phosphorylation plays an important role in proper synaptic function and transmission. Loss of Cdk5 activity results in abnormal development of the nervous system accompanied by massive disruptions in cortical migration and lamination, therefore impacting synaptic activity. The Cdk5 activator p35 associates with delta-catenin, the synaptic adherens junction protein that serves as part of the anchorage complex of AMPA receptor at the postsynaptic membrane. However, the implications of Cdk5-mediated phosphorylation of delta-catenin have not been fully elucidated. Here we show that Cdk5-mediated phosphorylation of delta-catenin regulates its subcellular localization accompanied by changes in dendritic morphogenesis and synaptic activity. We identified two Cdk5 phosphorylation sites in mouse delta-catenin, serines 300 and 357, and report that loss of Cdk5 phosphorylation of delta-catenin increased its localization to the membrane. Furthermore, mutations of the serines 300 and 357 to alanines to mimic nonphosphorylated delta-catenin resulted in increased dendritic protrusions accompanied by increased AMPA receptor subunit GluR2 localization at the membrane. Consistent with these observations, loss of Cdk5 phosphorylation of delta-catenin increased the AMPA/NMDA ratio. This study reveals how Cdk5 phosphorylation of the synaptic mediator protein delta-catenin can alter its localization at the synapse to impact neuronal synaptic activity.


Subject(s)
Catenins/metabolism , Cyclin-Dependent Kinase 5/metabolism , Neurons/metabolism , Receptors, AMPA/metabolism , Synapses/metabolism , Synaptic Transmission/physiology , Animals , Cells, Cultured , Cerebral Cortex/metabolism , Electrophysiology , Immunohistochemistry , Mice , Mutation , Nerve Tissue Proteins/metabolism , Neurons/cytology , Phosphorylation , Radioligand Assay , Delta Catenin
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