Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 11 de 11
Filter
Add more filters










Publication year range
1.
Front Pharmacol ; 13: 867457, 2022.
Article in English | MEDLINE | ID: mdl-36120380

ABSTRACT

Disrupted tau proteostasis and transneuronal spread is a pathological hallmark of Alzheimer's disease. Neurodegenerative diseases remain an unmet medical need and novel disease modifying therapeutics are paramount. Our objective was to develop a mechanistic mathematical model to enhance our understanding of tau antibody pharmacokinetics and pharmacodynamics in animals and humans. A physiologically-based pharmacokinetic-pharmacodynamic (PBPK-PD) modeling approach was employed to support the preclinical development and clinical translation of therapeutic antibodies targeting tau for the treatment of Alzheimer's disease. The pharmacokinetics of a tau antibody was evaluated in rat and non-human primate microdialysis studies. Model validation for humans was performed using publicly available clinical data for gosuranemab. In-silico analyses were performed to predict tau engagement in human brain for a range of tau antibody affinities and various dosing regimens. PBPK-PD modeling enabled a quantitative understanding for the relationship between dose, affinity, and target engagement, which supported lead candidate optimization and predictions of clinically efficacious dosing regimens.

2.
Bioconjug Chem ; 29(7): 2357-2369, 2018 07 18.
Article in English | MEDLINE | ID: mdl-29923706

ABSTRACT

Glucocorticoids (GCs) are excellent anti-inflammatory drugs but are dose-limited by on-target toxicity. We sought to solve this problem by delivering GCs to immune cells with antibody-drug conjugates (ADCs) using antibodies containing site-specific incorporation of a non-natural amino acid, novel linker chemistry for in vitro and in vivo stability, and existing and novel glucocorticoid receptor (GR) agonists as payloads. We directed fluticasone propionate to human antigen-presenting immune cells to afford GR activation that was dependent on the targeted antigen. However, mechanism of action studies pointed to accumulation of free payload in the tissue culture supernatant as the dominant driver of activity and indeed administration of the ADC to human CD74 transgenic mice failed to activate GR target genes in splenic B cells. Suspecting dissipation of released payload, we designed an ADC bearing a novel GR agonist payload with reduced permeability which afforded cell-intrinsic activity in human B cells. Our work shows that antibody-targeting offers significant potential for rescuing existing and new dose-limited drugs outside the field of oncology.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Antigens, Differentiation, B-Lymphocyte/immunology , B-Lymphocytes/metabolism , Drug Delivery Systems/methods , Glucocorticoids/administration & dosage , Histocompatibility Antigens Class II/immunology , Immunoconjugates/therapeutic use , Animals , Anti-Inflammatory Agents/therapeutic use , B-Lymphocytes/drug effects , Drug Development , Drug Stability , Fluticasone/administration & dosage , Humans , Mice , Mice, Transgenic , Receptors, Glucocorticoid/agonists
3.
J Pharm Sci ; 104(12): 4002-4014, 2015 Dec.
Article in English | MEDLINE | ID: mdl-26308749

ABSTRACT

IgG4 antibodies are evolving as an important class of cancer immunotherapies. However, human IgG4 can undergo Fab arm (half molecule) exchange with other IgG4 molecules in vivo. The hinge modification by a point mutation (S228P) prevents half molecule exchange of IgG4. However, the experimental confirmation is still expected by regulatory agencies. Here, we report for the first time the extensive analysis of half molecule exchange for a hinge-modified therapeutic IgG4 molecule, pembrolizumab (Keytruda) targeting programmed death 1 (PD1) receptor that was approved for advanced melanoma. Studies were performed in buffer or human serum using multiple exchange partners including natalizumab (Tysabri) and human IgG4 pool. Formation of bispecific antibodies was monitored by fluorescence resonance energy transfer, exchange with Fc fragments, mixed mode chromatography, immunoassays, and liquid chromatography-mass spectrometry. The half molecule exchange was also examined in vivo in SCID (severe combined immunodeficiency) mice. Both in vitro and in vivo results indicate that the hinge modification in pembrolizumab prevented half molecule exchange, whereas the unmodified counterpart anti-PD1 wt showed active exchange activity with other IgG4 antibodies or self-exchange activity with its own molecules. Our work, as an example expected for meeting regulatory requirements, contributes to establish without ambiguity that hinge-modified IgG4 antibodies are suitable for biotherapeutic applications.


Subject(s)
Antibodies, Monoclonal, Humanized/immunology , Immunoglobulin G/immunology , Animals , Chromatography/methods , Female , Humans , Immunoassay/methods , Immunoglobulin Fab Fragments/immunology , Immunoglobulin Fc Fragments/immunology , Mice , Mice, SCID , Programmed Cell Death 1 Receptor/immunology
4.
J Exp Med ; 207(3): 579-89, 2010 Mar 15.
Article in English | MEDLINE | ID: mdl-20212065

ABSTRACT

DNAX adaptor protein 12 (DAP12) is a trans-membrane adaptor molecule that transduces activating signals in NK and myeloid cells. Absence of functional Dap12 results in osteoclast defects and bone abnormalities. Because DAP12 has no extracelluar binding domains, it must pair with cell surface receptors for signal transduction. There are at least 15 known DAP12-associating cell surface receptors with distinct temporal and cell type-specific expression patterns. Our aim was to determine which receptors may be important in DAP12-associated bone pathologies. Here, we identify myeloid DAP12-associating lectin (MDL)-1 receptor (also known as CLEC5A) as a key regulator of synovial injury and bone erosion during autoimmune joint inflammation. Activation of MDL-1 leads to enhanced recruitment of inflammatory macrophages and neutrophils to the joint and promotes bone erosion. Functional blockade of MDL-1 receptor via Mdl1 deletion or treatment with MDL-1-Ig fusion protein reduces the clinical signs of autoimmune joint inflammation. These findings suggest that MDL-1 receptor may be a therapeutic target for treatment of immune-mediated skeletal disorders.


Subject(s)
Adaptor Proteins, Signal Transducing/physiology , Inflammation/immunology , Lectins, C-Type/physiology , Membrane Proteins/physiology , Receptors, Cell Surface/physiology , Adaptor Proteins, Signal Transducing/immunology , Animals , Antibodies, Monoclonal/immunology , Arthritis, Rheumatoid/immunology , Arthritis, Rheumatoid/physiopathology , Autoimmune Diseases/immunology , Autoimmune Diseases/therapy , Bone Marrow Cells/immunology , Bone Marrow Cells/physiology , Gene Expression Regulation , Humans , Inflammation/physiopathology , Inflammation/therapy , Joint Diseases/immunology , Killer Cells, Natural/immunology , Lectins, C-Type/genetics , Lectins, C-Type/immunology , Membrane Proteins/immunology , Mice , Mice, Inbred C57BL , Mice, Inbred Strains , Osteoblasts/physiology , Receptors, Cell Surface/genetics , Receptors, Cell Surface/immunology
5.
Infect Immun ; 78(3): 1353-63, 2010 Mar.
Article in English | MEDLINE | ID: mdl-20065029

ABSTRACT

Paired immunoglobulin-like type 2 receptors (PILRs) inhibitory PILRalpha and activating PILRbeta are predominantly expressed on myeloid cells. Their functions in host defense and inflammation are largely unknown, and in this study, we evaluated their roles in an acute Staphylococcus aureus pneumonia model. Compared to their respective controls, Pilrb(-/-) mice or mice in which PILRalpha was activated with an agonistic antibody showed improved clearance of pulmonary staphylococci and improved survival. These mice had reduced serum or bronchoalveolar lavage fluid levels of interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNF-alpha), and IL-6 and elevated levels of gamma interferon (IFN-gamma), IL-12, and IL-10. In contrast, mice in which PILRbeta was activated had increased lung bacterial burdens and higher mortality coupled with an intense proinflammatory response with highly elevated levels of IL-1beta, TNF-alpha, and IL-6. Treatment groups with reduced bacterial burdens had higher levels of Keratinocyte-derived chemokine (KC), macrophage inflammatory protein 2 (MIP-2), and MIP-1alpha in bronchoalveolar lavage fluid and an increased influx of neutrophils and macrophages to the lungs. Consistent with our in vivo findings, bone marrow-derived macrophages from Pilrb(-/-) mice released significantly less IL-1beta and TNF-alpha and more IFN-gamma and IL-12 than did the wild-type macrophages when directly stimulated with heat-killed S. aureus. To our knowledge, this is the first evidence that S. aureus directly interacts with PILRbeta. It provides a mechanism by which manipulating the balance in favor of an inhibitory PILR signal, by activation of PILRalpha or deletion of PILRbeta, helps to control acute S. aureus-mediated pneumonia and attenuate the inflammatory response. These results highlight the importance of PILRs in innate immunity and the control of inflammation.


Subject(s)
Pneumonia, Staphylococcal/immunology , Pneumonia, Staphylococcal/pathology , Receptors, Immunologic/metabolism , Signal Transduction , Staphylococcus aureus/immunology , Animals , Blood Chemical Analysis , Bronchoalveolar Lavage Fluid/chemistry , Colony Count, Microbial , Cytokines/analysis , Cytokines/blood , Female , Lung/microbiology , Macrophages/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Receptors, Immunologic/deficiency , Receptors, Immunologic/immunology , Survival Analysis
6.
J Exp Med ; 204(6): 1463-74, 2007 Jun 11.
Article in English | MEDLINE | ID: mdl-17548519

ABSTRACT

Polymicrobial sepsis alters the adaptive immune response and induces T cell suppression and Th2 immune polarization. We identify a GR-1(+)CD11b(+) population whose numbers dramatically increase and remain elevated in the spleen, lymph nodes, and bone marrow during polymicrobial sepsis. Phenotypically, these cells are heterogeneous, immature, predominantly myeloid progenitors that express interleukin 10 and several other cytokines and chemokines. Splenic GR-1(+) cells effectively suppress antigen-specific CD8(+) T cell interferon (IFN) gamma production but only modestly suppress antigen-specific and nonspecific CD4(+) T cell proliferation. GR-1(+) cell depletion in vivo prevents both the sepsis-induced augmentation of Th2 cell-dependent and depression of Th1 cell-dependent antibody production. Signaling through MyD88, but not Toll-like receptor 4, TIR domain-containing adaptor-inducing IFN-beta, or the IFN-alpha/beta receptor, is required for complete GR-1(+)CD11b(+) expansion. GR-1(+)CD11b(+) cells contribute to sepsis-induced T cell suppression and preferential Th2 polarization.


Subject(s)
CD11b Antigen/metabolism , Immune Tolerance/immunology , Myeloid Differentiation Factor 88/metabolism , Myeloid Progenitor Cells/immunology , Receptors, Chemokine/metabolism , Sepsis/immunology , Th2 Cells/immunology , Animals , Cell Proliferation , Colony-Forming Units Assay , Cytokines/metabolism , Flow Cytometry , Lymphoid Tissue/cytology , Lymphoid Tissue/immunology , Mice , Mice, Inbred C57BL , Myeloid Progenitor Cells/metabolism , Signal Transduction/immunology
7.
Blood ; 105(12): 4749-51, 2005 Jun 15.
Article in English | MEDLINE | ID: mdl-15741223

ABSTRACT

Interleukin-4 (IL-4), a major T-helper type 2 (Th2) cytokine, primes dendritic cells (DCs) for IL-12 production, suggesting a negative feedback loop to prevent dysregulated Th2 inflammation, such as allergy. We previously showed that human thymic stromal lymphopoietin (TSLP), highly expressed by keratinocytes of atopic dermatitis, activates CD11c(+) DCs to induce the differentiation of naive CD4(+) and CD8(+) T cells into proallergic effectors. Here we show that TSLP primes DCs to produce large amounts of IL-12 after CD40 ligand stimulation, similar to IL-4 priming of DCs. In contrast to IL-4 priming, DCs activated with TSLP and CD40 ligand induce the differentiation of naive CD4(+) T cells into effectors producing both Th1 and Th2 cytokines, a unique profile that is reminiscent of the late phase of allergy. Thus, TSLP is a major regulatory cytokine for IL-12 production by DCs, and TSLP-activated DCs could promote the persistence of Th2 inflammation even in the presence of IL-12-inducing signals.


Subject(s)
CD40 Ligand/metabolism , Cytokines/physiology , Dendritic Cells/cytology , Interleukin-12/metabolism , Myeloid Cells/metabolism , Th2 Cells/cytology , CD11c Antigen/biosynthesis , CD4 Antigens/biosynthesis , CD40 Antigens/biosynthesis , CD40 Ligand/biosynthesis , CD8 Antigens/biosynthesis , Cell Differentiation , Cells, Cultured , Coculture Techniques , Cytokines/metabolism , Dendritic Cells/metabolism , Dermatitis, Atopic/metabolism , Humans , Inflammation , Interleukin-4/metabolism , Keratinocytes/cytology , Signal Transduction , Time Factors , Thymic Stromal Lymphopoietin
8.
Blood ; 103(7): 2547-53, 2004 Apr 01.
Article in English | MEDLINE | ID: mdl-14670916

ABSTRACT

Type 1 interferon-producing cells (IPCs), also known as plasmacytoid dendritic cell (DC) precursors, represent the key effectors in antiviral innate immunity and triggers for adaptive immune responses. IPCs play important roles in the pathogenesis of systemic lupus erythematosus (SLE) and in modulating immune responses after hematopoietic stem cell transplantation. Understanding IPC development from hematopoietic progenitor cells (HPCs) may provide critical information in controlling viral infection, autoimmune SLE, and graft-versus-host disease. FLT3-ligand (FLT3-L) represents a key IPC differentiation factor from HPCs. Although hematopoietic cytokines such as interleukin-3 (IL-3), IL-7, stem cell factor (SCF), macrophage-colony-stimulating factor (M-CSF), and granulocyte M-CSF (GM-CSF) promote the expansion of CD34+ HPCs in FLT3-L culture, they strongly inhibit HPC differentiation into IPCs. Here we show that thrombopoietin (TPO) cooperates with FLT3-L, inducing CD34+ HPCs to undergo a 400-fold expansion in cell numbers and to generate more than 6 x 10(6) IPCs per 10(6) CD34+ HPCs within 30 days in culture. IPCs derived from HPCs in FLT3-L/TPO cultures display blood IPC phenotype and have the capacity to produce large amounts of interferon-alpha (IFN-alpha) and to differentiate into mature DCs. This culture system, combined with the use of adult peripheral blood CD34+ HPCs purified from G-CSF-mobilized donors, permits the generation of more than 10(9) IPCs from a single blood donor.


Subject(s)
Dendritic Cells/cytology , Hematopoiesis/physiology , Hematopoietic Stem Cells/cytology , Membrane Proteins/physiology , Thrombopoietin/physiology , Antigens, CD/analysis , Antigens, CD34/analysis , Cell Culture Techniques/methods , Cell Differentiation/immunology , Cell Division/physiology , Cells, Cultured , Dendritic Cells/immunology , Fetus , Gestational Age , Granulocyte Colony-Stimulating Factor/pharmacology , Hematopoietic Stem Cell Mobilization/methods , Humans , Lymphocyte Activation , Plasma Cells/cytology , Plasma Cells/immunology , T-Lymphocytes/immunology
9.
J Exp Med ; 197(8): 1059-63, 2003 Apr 21.
Article in English | MEDLINE | ID: mdl-12707303

ABSTRACT

Human thymic stromal lymphopoietin (TSLP) is a novel epithelial cell-derived cytokine, which induces dendritic cell (DC)-mediated CD4+ T cell responses with a proallergic phenotype. Although the participation of CD8+ T cells in allergic inflammation is well documented, their functional properties as well as the pathways leading to their generation remain poorly understood. Here, we show that TSLP-activated CD11c+ DCs potently activate and expand naive CD8+ T cells, and induce their differentiation into interleukin (IL)-5 and IL-13-producing effectors exhibiting poor cytolytic activity. Additional CD40L triggering of TSLP-activated DCs induced CD8+ T cells with potent cytolytic activity, producing large amounts of interferon (IFN)-gamma, while retaining their capacity to produce IL-5 and IL-13. These data further support the role of TSLP as initial trigger of allergic T cell responses and suggest that CD40L-expressing cells may act in combination with TSLP to amplify and sustain pro-allergic responses and cause tissue damage by promoting the generation of IFN-gamma-producing cytotoxic effectors.


Subject(s)
CD40 Ligand/metabolism , CD8-Positive T-Lymphocytes/immunology , Cytokines/metabolism , Dendritic Cells/immunology , Hypersensitivity/immunology , Lymphocyte Activation , CD11c Antigen/immunology , CD11c Antigen/metabolism , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/metabolism , Cell Differentiation/physiology , Cells, Cultured , Dendritic Cells/cytology , Dendritic Cells/metabolism , Humans , Inflammation/immunology , Interferon-gamma/metabolism , Interleukin-13/metabolism , Interleukin-5/metabolism , Thymic Stromal Lymphopoietin
10.
Nat Immunol ; 3(7): 673-80, 2002 Jul.
Article in English | MEDLINE | ID: mdl-12055625

ABSTRACT

Whether epithelial cells play a role in triggering the immune cascade leading to T helper 2 (T(H)2)-type allergic inflammation is not known. We show here that human thymic stromal lymphopoietin (TSLP) potently activated CD11c(+) dendritic cells (DCs) and induced production of the T(H)2-attracting chemokines TARC (thymus and activation-regulated chemokine; also known as CCL17) and MDC (macrophage-derived chemokine; CCL22). TSLP-activated DCs primed naïve T(H) cells to produce the proallergic cytokines interleukin 4 (IL-4), IL-5, IL-13 and tumor necrosis factor-alpha, while down-regulating IL-10 and interferon-gamma. TSLP was highly expressed by epithelial cells, especially keratinocytes from patients with atopic dermatitis. TSLP expression was associated with Langerhans cell migration and activation in situ. These findings shed new light on the function of human TSLP and the role played by epithelial cells and DCs in initiating allergic inflammation.


Subject(s)
CD11 Antigens , Cytokines/immunology , Dendritic Cells/immunology , Dermatitis, Atopic/immunology , Interleukin-7/immunology , Th2 Cells/immunology , Thymus Gland/immunology , Adult , Biomarkers , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/immunology , Cell Division , Cells, Cultured , Coculture Techniques , Cytokines/genetics , Cytokines/pharmacology , Dendritic Cells/cytology , Dendritic Cells/drug effects , Dermatitis, Atopic/pathology , Epithelial Cells/cytology , Epithelial Cells/immunology , Gene Expression , Humans , Interleukin-7/pharmacology , Langerhans Cells/cytology , Langerhans Cells/immunology , Palatine Tonsil/immunology , RNA, Messenger , Skin/immunology , Skin/pathology , Th2 Cells/cytology , Thymic Stromal Lymphopoietin
11.
J Exp Med ; 195(7): 953-8, 2002 Apr 01.
Article in English | MEDLINE | ID: mdl-11927638

ABSTRACT

Plasmacytoid predendritic cells or type 1 interferon (IFN)-producing cells (IPCs) have recently been identified in mice. Although culture systems giving rise to different murine dendritic cell subsets have been established, the developmental regulation of murine plasmacytoid IPCs and the culture conditions leading to their generation remain unknown. Here we show that large numbers of over 40% pure CD11c(+)CD11b(-)B220(+)Gr-1(+) IPCs can be generated from mouse bone marrow cultures with FLT3-ligand. By contrast GM-CSF or TNF-alpha, which promote the generation of CD11c(+)CD11b(+)B220(-) myeloid DCs, block completely the development of IPCs. IPCs generated display similar features to human IPCs, such as the plasmacytoid morphology, the ability to produce large amounts of IFN-alpha in responses to herpes simplex virus, and the capacity to respond to ligands for Toll-like receptor 9 (TLR-9; CpG ODN 1668), but not to ligands for TLR-4 (lipopolysaccharide [LPS]). Unlike human IPCs which produce little IL-12p70, mouse IPCs produce IL-12p70 in response to CpG ODN 1668 and herpes simplex virus. This study demonstrates that the development of murine CD11c(+)CD11b(-)B220(+)Gr-1(+) IPCs and CD11c(+)CD11b(+)B220(-) myeloid DCs is differentially regulated by FLT3-ligand and granulocyte/macrophage colony-stimulating factor. Human IPCs and mouse IPCs display different ability to produce IL-12p70. Large numbers of mouse IPCs can now be obtained from total bone marrow culture.


Subject(s)
Dendritic Cells/physiology , Granulocyte-Macrophage Colony-Stimulating Factor/pharmacology , Animals , Antigens, Differentiation/analysis , Bone Marrow Cells/cytology , Bone Marrow Cells/drug effects , Bone Marrow Cells/immunology , Cell Differentiation , Cells, Cultured , Dendritic Cells/drug effects , Female , Gene Expression Regulation , Hematopoiesis , Humans , Kinetics , Macrophage-1 Antigen/analysis , Membrane Proteins , Mice , Mice, Inbred BALB C , Species Specificity , Time Factors , Tumor Necrosis Factor-alpha/pharmacology
SELECTION OF CITATIONS
SEARCH DETAIL
...