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Bispecific asymmetric heterodimers comprising anti-cd3 constructs

A technology of heterodimers and constructs, applied in the direction of antibodies, antibody medical ingredients, medical preparations containing active ingredients, etc.

Inactive Publication Date: 2015-05-20
ZYMEWORKS BC INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The development of such multivalent and multispecific therapeutic proteins with favorable pharmacokinetics and functional activities has been a challenge

Method used

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  • Bispecific asymmetric heterodimers comprising anti-cd3 constructs
  • Bispecific asymmetric heterodimers comprising anti-cd3 constructs
  • Bispecific asymmetric heterodimers comprising anti-cd3 constructs

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0323] Example 1: Bispecific CD3-CD19 scFv fused to asymmetric IgGl Fc.

[0324] A bispecific CD3-CD19 scFv fused to an asymmetric IgG1 Fc heterodimer exhibiting stability comparable to native Fc homodimers is a novel composition identified as v873. v873 belongs to a novel family of CD3-based bispecific asymmetric IgGl antibodies that can be expressed and purified in mammalian CHO cells in significantly higher yields compared to the Amgen / Micromet bscCD19xCD3 BiTE bispecific. V873 exhibits unexpected effector cell: target cell binding, bridging, and target cell killing.

[0325] V873 and CD3-based bispecific asymmetric antibodies have utility in targeting T cell-mediated killing of diseased cells and thus may be applicable in the treatment of cancer as well as autoimmune and inflammatory diseases. v873 is a bispecific CD3-CD19 scFv fused to an asymmetric IgG1 Fc. v873 represents a novel class of bispecific asymmetric antibodies comprising an anti-CD3 warhead and a second war...

Embodiment 2

[0331] Example 2. Design, expression and purification of heteromultimeric constructs with heterodimeric Fc.

[0332] Exemplary bispecific anti-CD3 and anti-CD19 heterodimeric antibodies

[0333] An exemplary schematic representation of an anti-CD3 / anti-CD19 antibody is shown in Figure 1A.

[0334] v873, v874, v875 exemplify bispecific anti-CD3 / anti-CD19 heterodimeric Fc constructs and were prepared and tested as described below. When the description includes a reference to a BiTE, it means that the antibody construct has the same properties as the anti- The same amino acid sequence as the VH or VL of the CD3 anti-CD19 BiTE molecule.

[0335] v873 has an anti-CD19 BiTE(VL-VH) scFv on chain A of the heterodimeric Fc and a CD3 BiTE on chain B TM (VH-VL) scFv with mutation L351Y_F405A_Y407V on chain A and mutation T366L_K392M_T394W on chain B. [The polypeptide sequence corresponds to SEQ ID No: 26 and 28]

[0336] v874 has an anti-CD19 BiTE on chain A of the heterodimeric Fc...

Embodiment 3

[0407] Example 3: Heteromultimeric v873 can bridge Jurkat CD3 T cells and Raji CD19 B cells.

[0408] The ability of v873 to bridge T cells and B cells was tested by FACS analysis as follows.

[0409] Whole-cell bridging by FACS

[0410] 1x10 that will be suspended in RPMI 6 Cells / ml were labeled and mixed with 0.3 μM of the appropriate CellTrace marker and incubated for 25 min at 37 °C in a water bath.

[0411] The pellet was resuspended in 2ml of L10+GS1+NaN3 to a final concentration of 5x x106 cells / ml.

[0412] Cell suspensions were analyzed by flow cytometry (1 / 5 dilution) to verify proper cell labeling and laser settings. flow-check and flow-set fluorophores for verification of instrument standardization, optical alignment and fluidics

[0413] After validation by flow cytometry and prior to bridging, each cell line was split at the desired ratio at 1x106 cells / ml at a final concentration.

[0414] T:T bridging was scored with Jurkat-violet+Jurkat-far red, B:B bridg...

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Abstract

Disclosed herein are isolated multi-specific heteromultimer constructs that bind to CD3 expressed on T-cells and to an antigen expressed on B-cells. The multi-specific heteromultimer constructs are capable of bridging T- and B-cells and mediating killing of B-cells. The multi-specific heteromultimer constructs are based on a heterodimeric Fc scaffold or on a segmented albumin scaffold. Also disclosed herein are multi-specific heteromultimer constructs that bind to HER2 and HER3.

Description

[0001] Cross References to Related Applications [0002] This application claims the benefit of US Application Serial No. 61 / 671,640, filed July 13, 2012, and US Application Serial No. 61 / 845,948, filed July 13, 2013, which are hereby incorporated by reference in their entireties. technical field [0003] The field of the invention is the rational design of multispecific scaffolds comprising CD3 binding domains for custom development of biotherapeutics. Background technique [0004] In the field of therapeutic proteins, antibodies with their multivalent target binding characteristics are excellent scaffolds for designing drug candidates. Further advancing these features, designed bispecific antibodies and other fusion multispecific therapeutics exhibit dual or multiple target specificities and the opportunity to generate drugs with novel modes of action. The development of such multivalent and multispecific therapeutic proteins with favorable pharmacokinetics and functional...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/00A61K39/395C12P21/08
CPCC07K16/2803C07K16/2887C07K16/32C07K2317/31C07K2317/35C07K2317/526C07K2317/528C07K2317/622C07K2317/64C07K2317/71C07K2317/72C07K2317/73C07K2317/732C07K2317/74C07K2317/92C07K2317/94C07K2319/31C07K2317/52C07K2317/524C07K16/2809A61P29/00A61P31/00A61P31/12A61P33/14A61P35/00A61P35/02A61P37/06A61P37/08C07K2317/60C07K2317/50
Inventor S·B·迪克西特T·斯普雷特冯克罗登斯泰恩G·Y·K·吴N·E·韦塞
Owner ZYMEWORKS BC INC
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