Designed ankyrin repeat domains with binding specificity for serum albumin

A technology of repeating domain and binding specificity, applied in the field of designed ankyrin repeating domain with binding specificity to serum albumin, which can solve the problems of cost and the like

Pending Publication Date: 2017-12-08
MOLECULAR PARTNERS AG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This is technically difficult to achieve with antibody drugs, which further require cost-intensive production in mammalian cells

Method used

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  • Designed ankyrin repeat domains with binding specificity for serum albumin
  • Designed ankyrin repeat domains with binding specificity for serum albumin
  • Designed ankyrin repeat domains with binding specificity for serum albumin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0140] Example 1: Selection, Expression, Purification and Analysis of Designer Ankyrin Repeat Domains with Binding Specificity for VEGF-A, HGF, or Serum Albumin

[0141] Using ribosome display (Binz et al., 2004 (cit. above)), by the method described in WO2010 / 060748 for generating a designer ankyrin repeat domain with binding specificity for VEGF-A, and WO2010 / 060748 The method described in 2014 / 191574 for generating a designer ankyrin repeat domain with binding specificity for HGF, and the method for generating an ankyrin repeat domain with binding specificity for serum albumin described in WO 2012 / 069654 Ankyrin repeat domain approach, selecting from combinatorial libraries designer ankyrin repeat domains with binding specificity for VEGF-A, HGF or serum albumin. Binding of selected clones to specific (VEGF-A, HGF, or serum albumin, respectively) and nonspecific (e.g., MBP, E. Hundreds of designer ankyrin repeat domains with binding specificity for VEGF-A, HGF, or serum al...

Embodiment 2

[0143] Example 2: Characterization of Designer Ankyrin Repeat Domains Using Surface Plasmon Resonance

[0144] SPR was measured using a ProteOn instrument (BioRad) and the measurements were performed according to standard protocols known to those skilled in the art. The Kd values ​​measured for selected proteins are listed in Tables 1-3.

[0145] Table 1: Examples of dissociation constants for designer ankyrin repeat domains that bind to human VEGF-A

[0146]

[0147] * Proteins #12, #13 and #16 in this table represent designer ankyrins consisting of the corresponding amino acid sequences of SEQ ID NOs: 12, 13 and 16 additionally comprising an N-terminal His-tag (SEQ ID NO: 1) repeat domain.

[0148] Similar VEGF-A dissociation constant values ​​were obtained for proteins #14, #15 and #17 to #22.

[0149] Table 2: Examples of dissociation constants for designer ankyrin repeat domains binding to human HGF

[0150]

[0151] * Proteins #23 to #29 in this table represe...

Embodiment 3

[0157] Example 3: Competition Binding Assay and Receptor Competition Binding Assay

[0158]Designer ankyrin repeat domains with binding specificity for VEGF-A, HGF, or serum albumin, respectively, were characterized by competition assays. These assays are well known to those skilled in the art. For designer ankyrin repeat domains with binding specificity for VEGF-A, quantitative sandwich enzyme immunoassay technique (VEGF-A Quantikine kit DVE00, R&D Systems, Inc.) was used according to the manufacturer's instructions. A monoclonal antibody specific for VEGF-A was pre-coated onto a microwell plate. VEGF-A standards and mixtures of various concentrations of VEGF-A (20 pM) and proteins #18, #19, or #20 were applied to the wells, and any free VEGF-A present (i.e., not bound to the engineered The ankyrin repeat domain) is bound by the immobilized antibody. After washing away any unbound material, an enzyme-linked polyclonal antibody specific for VEGF-A is added to the wells. Af...

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PUM

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Abstract

New designed ankyrin repeat domains with binding specificity for serum albumin, recombinant binding proteins comprising at least two designed ankyrin repeat domains with binding specificity for serum albumin, as well as recombinant binding proteins comprising at least one designed ankyrin repeat domain with binding specificity for hepatocyte growth factor (HGF), at least one designed ankyrin repeat domain with binding specificity for vascular endothelial growth factor (VEGF-A), and at least two designed ankyrin repeat domain with binding specificity for serum albumin are described, as well as nucleic acids encoding such designed ankyrin repeat domains and recombinant binding proteins, pharmaceutical compositions comprising such designed ankyrin repeat domains, recombinant binding proteins or nucleic acids and the use of such designed ankyrin repeat domains, recombinant binding proteins, nucleic acids or pharmaceutical compositions in the treatment of diseases.

Description

[0001] Cross references to related patent applications [0002] This application claims the benefit of and priority to European Patent Application EP15162502 filed with the European Patent Office on April 2, 2015. This application also claims the benefit and priority of European Patent Application EP15162511 filed with the European Patent Office on April 2, 2015. The contents of European Patent Applications EP15162502 and EP 15162511 are hereby incorporated by reference in their entirety for all purposes, including all tables, drawings and claims, pursuant to PCT Article 4.18, also including any part of the description, claims or drawings not contained in Incorporation of any element or part referred to herein but in PCT Article 20.5(a). technical field [0003] A newly designed ankyrin repeat domain with binding specificity for serum albumin that exhibits improved storage stability is provided. Also provided are recombinant binding proteins comprising at least two designer ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/00C07K19/00A61K38/16A61P35/00
CPCC07K14/001A61K38/00C07K2319/31C07K2319/70C07K2319/00C07K14/47C07K2318/20A61P25/28A61P27/02A61P29/00A61P31/00A61P35/00A61P37/06A61P43/00A61P9/10C07K14/4703C12N15/62C07K14/765
Inventor T·巴克M·T·斯通普H·K·宾茨D·菲利普斯I·多拉多P·福雷尔F·W·默茨I·松德雷格D·斯泰纳M·古罗迪-格奥尔吉耶娃J·阿布拉姆萨力巴
Owner MOLECULAR PARTNERS AG
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