Scripps Korea Antibody Institute

Chuncheon, South Korea

Scripps Korea Antibody Institute

Chuncheon, South Korea
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Xie J.,Scripps Research Institute | Zhang H.,Scripps Research Institute | Yea K.,Scripps Korea Antibody Institute | Lerner R.A.,Scripps Research Institute
Proceedings of the National Academy of Sciences of the United States of America | Year: 2013

We report here the generation of antibody agonists fromintracellular combinatorial libraries that transdifferentiate human stem cells. Antibodies that are agonists for the granulocyte colony stimulating factor receptor were selected from intracellular libraries on the basis of their ability to activate signaling pathways in reporter cells. We used a specialized 'near neighbor' approach in which the entire antibody library and its target receptor are cointegrated into the plasma membranes of a population of reporter cells. This format favors unusual interactions between receptors and their protein ligands and ensures that the antibody acts in an autocrine manner on the cells that produce it. Unlike the natural granulocyte-colony stimulating factor that activates cells to differentiate along a predetermined pathway, the isolated agonist antibodies transdifferentiated human myeloid lineage CD34+ bone marrow cells into neural progenitors. This transdifferentiation by agonist antibodies is different frommore commonly used methods because initiation is agenetic. Antibodies that act at the plasma membrane may have therapeutic potential as agents that transdifferentiate autologous cells.


Kim M.,Seoul National University | Yoon S.,Seoul National University | Lee S.,Scripps Korea Antibody Institute | Ha S.A.,Catholic University of Korea | And 3 more authors.
PLoS ONE | Year: 2012

Gremlin-1, a bone morphogenetic protein (BMP) antagonist, is overexpressed in various cancerous tissues but its role in carcinogenesis has not been established. Here, we report that gremlin-1 binds various cancer cell lines and this interaction is inhibited by our newly developed gremlin-1 antibody, GRE1. Gremlin-1 binding to cancer cells was unaffected by the presence of BMP-2, BMP-4, and BMP-7. In addition, the binding was independent of vascular endothelial growth factor receptor-2 (VEGFR2) expression on the cell surface. Addition of gremlin-1 to A549 cells induced a fibroblast-like morphology and decreased E-cadherin expression. In a scratch wound healing assay, A549 cells incubated with gremlin-1 or transfected with gremlin-1 showed increased migration, which was inhibited in the presence of the GRE1 antibody. Gremlin-1 transfected A549 cells also exhibited increased invasiveness as well as an increased growth rate. These effects were also inhibited by the addition of the GRE1 antibody. In conclusion, this study demonstrates that gremlin-1 directly interacts with cancer cells in a BMP- and VEGFR2-independent manner and can induce cell migration, invasion, and proliferation. © 2012 Kim et al.


Zhang H.,Scripps Research Institute | Yea K.,Scripps Korea Antibody Institute | Xie J.,Scripps Research Institute | Ruiz D.,Scripps Research Institute | And 2 more authors.
Chemistry and Biology | Year: 2013

We describe a system for direct selection of antibodies that are receptor agonists. Combinatorial antibody libraries in lentiviruses are used to infect eukaryotic cells that contain a fluorescent reporter system coupled to the receptor for which receptor agonist antibodies are sought. In this embodiment of the method, very large numbers of candidate antibodies expressing lentivirus and eukaryotic reporter cells are packaged together in a format where each is capable of replication, thereby forging a direct link between genotype and phenotype. Following infection, cells that fluoresce are sorted and the integrated genes encoding the agonist antibodies recovered. We validated the system by illustrating its ability to generate rapidly potent antibody agonists that are complete thrombopoietin phenocopies. The system should be generalizable to any pathway where its activation can be linked to production of a selectable phenotype. © 2013 Elsevier Ltd.


Peng Y.,Scripps Research Institute | Kim D.H.,Scripps Korea Antibody Institute | Jones T.M.,Scripps Research Institute | Ruiz D.I.,Scripps Research Institute | Lerner R.A.,Scripps Research Institute
Angewandte Chemie - International Edition | Year: 2013

Vectors have been constructed that express the chitin-binding domain (ChBD) on eukaryotic cell surfaces. The ChBD is linked to enhanced green fluorescent protein (EGFP) through a protein that spans the plasma membrane (see picture). This binding functionality does not have a counterpart in eukaryotes, thereby endowing the modified cell surface with a property that is orthogonal to animal cells. © 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.


Bowden T.A.,University of Oxford | Baruah K.,University of Oxford | Coles C.H.,University of Oxford | Harvey D.J.,University of Oxford | And 8 more authors.
Journal of the American Chemical Society | Year: 2012

Human IgG Fc glycosylation modulates immunological effector functions such as antibody-dependent cellular cytotoxicity and phagocytosis. Engineering of Fc glycans therefore enables fine-tuning of the therapeutic properties of monoclonal antibodies. The N-linked glycans of Fc are typically complex-type, forming a network of noncovalent interactions along the protein surface of the Cγ2 domain. Here, we manipulate the mammalian glycan-processing pathway to trap IgG1 Fc at sequential stages of maturation, from oligomannose- to hybrid- to complex-type glycans, and show that the Fc is structurally stabilized following the transition of glycans from their hybrid- to complex-type state. X-ray crystallographic analysis of this hybrid-type intermediate reveals that N-linked glycans undergo conformational changes upon maturation, including a flip within the trimannosyl core. Our crystal structure of this intermediate reveals a molecular basis for antibody biogenesis and provides a template for the structure-guided engineering of the protein-glycan interface of therapeutic antibodies. © 2012 American Chemical Society.


Yu X.,University of Oxford | Baruah K.,University of Oxford | Harvey D.J.,University of Oxford | Vasiljevic S.,University of Oxford | And 6 more authors.
Journal of the American Chemical Society | Year: 2013

Biologically active conformations of the IgG1 Fc homodimer are maintained by multiple hydrophobic interactions between the protein surface and the N-glycan. The Fc glycan modulates biological effector functions, including antibody-dependent cellular cytotoxicity (ADCC) which is mediated in part through the activatory Fc receptor, FcγRIIIA. Consistent with previous reports, we found that site-directed mutations disrupting the protein-carbohydrate interface (F241A, F243A, V262E, and V264E) increased galactosylation and sialylation of the Fc and, concomitantly, reduced the affinity for FcγRIIIA. We rationalized this effect by crystallographic analysis of the IgG1 Fc F241A mutant, determined here to a resolution of 1.9 Å, which revealed localized destabilization of this glycan-protein interface. Given that sialylation of Fc glycans decreases ADCC, one explanation for the effect of these mutants on FcγRIIIA binding is their increased sialylation. However, a glycan-engineered IgG1 with hypergalactosylated and hypersialylated glycans exhibited unchanged binding affinity to FcγRIIIA. Moreover, when we expressed these mutants as a chemically uniform (Man 5GlcNAc2) glycoform, the individual effect of each mutation on FcγRIIIA affinity was preserved. This effect was broadly recapitulated for other Fc receptors (FcγRI, FcγRIIA, FcγRIIB, and FcγRIIIB). These data indicate that destabilization of the glycan-protein interactions, rather than increased galactosylation and sialylation, modifies the Fc conformation(s) relevant for FcγR binding. Engineering of the protein-carbohydrate interface thus provides an independent parameter in the engineering of Fc effector functions and a route to the synthesis of new classes of Fc domain with novel combinations of affinities for activatory and inhibitory Fc receptors. © 2013 American Chemical Society.


Patent
Scripps Korea Antibody Institute | Date: 2014-01-24

The present invention relates to a method for constructing an Fv library based on a combination of proteins, a method of screening a desired antibody using the constructed Fv library, an Fv antibody screened by the screening method, and an Fv library constructed by the Fv library construction method. The Fv library of the present invention is based on a combination of proteins so that members thereof can be individually analyzed for their function. Moreover, the Fv library enables a desired Fv antibody to be screened without needing a target antigen preparation. In addition, the protein combination based Fv library makes it possible to significantly reduce the number of protein purification processes to thereby reduce costs and time, compared to conventional DNA-based libraries.


Patent
Scripps Korea Antibody Institute | Date: 2013-06-14

Provided is an antibody specifically binding to the CTLD (C-type lectin like domain) of clecl4a (C-type lectin domain family 14, member A), a method for preparing the antibody, a composition for suppressing angiogenesis comprising the antibody, a method for suppressing angiogenesis by administering the antibody or the composition, a composition for preventing or treating cancer comprising the antibody, a method for treating cancer by administering the antibody or the composition, a composition for diagnosing cancer comprising the antibody, a kit for diagnosing cancer comprising the composition, a method for diagnosing cancer using the composition, a composition for suppressing angiogenesis comprising a material for inhibiting expression of clecl4a, a kit for angiogenesis comprising the composition, a method for suppressing angiogenesis or treating cancer using the composition, and the use of the CTLD of clecl4a as an epitope for an antibody suppressive of angiogenesis.


Patent
Scripps Korea Antibody Institute | Date: 2016-03-02

The present invention relates to a self-cutting fusion protein including sequentially a target protein, a peptide consisting of an amino acid sequence of LPXTG, a sortase A cutting function domain, and a tag from the amino terminal; a coding nucleic acid thereof; an expression vector of the present invention including a nucleic acid; and a cell transformed by the expression vector of the present invention. The present invention also relates to a target protein refinement method including the steps of culturing, dissolving, and refining the transformed cell. The present invention relates to a self-cutting fusion protein including a self-cutting cassette which consists of a sortase A cutting function domain and a peptide consisting of an LPXTG amino acid sequence that is a recognition sequence thereof, and is very useful in that the steps of refining a target protein and eliminating the tag thereof can be accomplished by a single refinement step instead of separate steps. Particularly, the invention may be employed in various fields requiring a huge amount of a protein of a high purity in that a target protein existing in an amino terminal enhances the column union ability and the self-cutting ability of a fusion protein, a target protein rid of the tag is obtained with a high purity, a cutting-buffer makes the steps of refining and eliminating the tag be accomplished through a single step, significantly reducing the time and efforts needed for refining, and the accomplishment through a single step reduces the loss of the protein yield. Particularly, the invention is useful for manufacturing a therapeutic antibody-drug complex.


Patent
Scripps Korea Antibody Institute | Date: 2015-12-02

The present invention relates to a method for constructing an Fv library based on a combination of proteins, a method of screening a desired antibody using the constructed Fv library, an Fv antibody screened by the screening method, and an Fv library constructed by the Fv library construction method. The Fv library of the present invention is based on a combination of proteins so that members thereof can be individually analyzed for their function. Moreover, the Fv library enables a desired Fv antibody to be screened without needing a target antigen preparation. In addition, the protein combination based Fv library makes it possible to significantly reduce the number of protein purification processes to thereby reduce costs and time, compared to conventional DNA-based libraries.

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