Cysteine peptide-enabled antibodies

A technology of amino acid and phenylalanine, applied in the direction of peptide, specific peptide, anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, etc.

Pending Publication Date: 2019-09-17
CITY OF HOPE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These approaches are limited by the available chemistry of the parental mAb and / or require extensive protein engineering

Method used

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  • Cysteine peptide-enabled antibodies
  • Cysteine peptide-enabled antibodies
  • Cysteine peptide-enabled antibodies

Examples

Experimental program
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preparation example Construction

[0179] For the preparation of suitable antibodies of the invention and uses according to the invention, e.g. recombinant, monoclonal or polyclonal antibodies, a number of techniques known in the art can be used (see, e.g., Kohler & Milstein, Nature 256:495-497 (1975); People such as Kozbor, Immunology Today 4:72 (1983); People such as Cole, pages 77-96 in Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc. (1985); Coligan, Current Protocols in Immunology (1991); Harlow & Lane, Antibodies, A Laboratory Manual (1988); and Goding, Monoclonal Antibodies: Principles and Practice (2nd Edition, 1986)). Genes encoding the heavy and light chains of an antibody of interest can be cloned from cells, for example, genes encoding monoclonal antibodies can be cloned from hybridomas and used to produce recombinant monoclonal antibodies. Gene libraries encoding the heavy and light chains of monoclonal antibodies can also be prepared from hybridomas or plasma cells. Random combination...

Embodiment 1

[0378] Example 1: Cys-Meditope

[0379]In some cases, it is desirable to add functionality to monoclonal antibodies through covalent linkages. To achieve this, Applicants used a meditope / meside-enabled Fab (meFab) interaction to generate a disulfide bond ( figure 1 with Figure 2A ).

[0380] Based on careful examination of the meditope-meFab interaction and the previous observation that the cysteine ​​(Cys) introduced at K208 by site-directed mutagenesis was not oxidized, Applicants' meditope-enabled monoclonal antibody to trastuzumab Cys was introduced at position 175 of alanine (Ala) in (memAb)V2(I83E). In addition, Applicants synthesized a linear meditope (SEQ. ID NO: 4) in which X is diphenylalanine and the serine (Ser) at position 6 is replaced by Cys. Applicants hypothesize that the Ala175Cys mutation in the memAb heavy chain and the Ser6Cys mutation in the meditope bring the sulfur groups into close proximity, promoting disulfide bond formation ( Figure 2A ).

...

Embodiment 2

[0395] Example 2: Stable site-specific modification of disulfide-conjugated monoclonal antibodies using meditope peptide-assisted

[0396] The high specificity and favorable pharmacological properties of monoclonal antibodies (mAbs) have generated significant interest in reengineering such molecules to enhance their therapeutic and diagnostic potential. Herein, applicants use high-affinity interactions between meditope peptides and meditope-enabled mAbs (memAbs) to drive rapid, efficient and robust site-specific formation of disulfide bonds. Applicants use this meditope, peptide-assisted conjugation technology (mPACT) platform to attach fluorescent dyes, cytotoxins, or "click" chemistry to memAbs and meFabs. More importantly, Applicants developed genetically encoded meditope-tagged biologics to generate stable bifunctional Fabs and mAbs. This includes conjugation of bacterially expressed fluorescent proteins, Nanobodies and Affibodies to memAbs and meFabs containing N-, C- or...

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Abstract

Provided herein are functionalized monoclonal antibodies (mAbs) including antibody fragments covalently linked to a peptide compound through a disulfide linkage. The disulfide linkage is between a cysteine in the Fab region of the antibody or fragment thereof and a thiol moiety of a side chain amino acid of the peptide compound. The covalently formed complexes including provided herein form highly stable and versatile drug delivery and diagnostic compositions.

Description

[0001] Cross References to Related Applications [0002] This application claims the benefit of U.S. Provisional Application No. 62 / 430,848, filed December 6, 2016, and U.S. Provisional Application No. 62 / 531,825, filed July 12, 2017, both in their entirety and for all purposes This article is incorporated by reference. [0003] References to "sequence listings," table or computer program listing appendices filed as ASCII files [0004] Created on December 5, 2017, 41,300 bytes, machine format IBM-PC, the sequence listing written in the file 48440-627001WO Sequence Listing_ST25.TXT of the MS-Windows operating system, hereby incorporated as a reference. Background technique [0005] Because of their specificity and favorable pharmacokinetics and pharmacodynamics, considerable efforts have been made to equip monoclonal antibodies (mAbs) with potent cytotoxins or biologics to enhance their therapeutic efficacy, or with radionuclides to equip monoclonal antibodies to image di...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28C07K7/08C07K7/06
CPCC07K7/06C07K7/08C07K16/32C07K2317/522C07K2317/55C07K2317/624A61K47/6889A61K47/6803A61K47/6851C07K16/283
Inventor J·C·威廉斯K·布兹伊梅克Y·马D·霍内J·金
Owner CITY OF HOPE
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