Enzymatic conjugation of polypeptides
Inventors
Bregeon, Delphine • Dennler, Patrick • Belmant, Christian • Fischer, Eliane • Gauthier, Laurent • ROMAGNÉ, FRANCOIS • Schibli, Roger
Assignees
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Publication Number
US-9717803-B2
Publication Date
2017-08-01
Expiration Date
Abstract
The present application relates to methods for the functionalization of immunoglobulins, in particular with drugs. Also disclosed herein are linking reagents, functionalized antibodies, pharmaceutical compositions, and method of treating disease and/or conditions.
Core Innovation
The invention relates to compositions comprising a population of antibodies or antibody fragments having the same primary amino acid sequence, wherein at least 90% of the population has (m) functionalized acceptor glutamine residues (Q) per antibody or fragment, with m selected from 2 or 4. Each functionalized acceptor glutamine residue is present within or substituted into the primary amino acid sequence of a constant region and has the structure of Formula IVb or a related lysine-based linker form.
The functionalized acceptor glutamine residues include defined linker and attachment architecture with linker and spacer system elements, including (C)n, X, L, V, Y, and M components, and RR2 as an addition product between a reactive moiety R and a complementary reactive moiety R2. The structures further allow non-cleavable or conditionally-cleavable moieties, and embodiments include heavy-chain-specific and sequence-based positioning constraints, including EU residue numbering and N297X or EU position 295 variants.
The compositions are tumor-binding, where the antibodies or antibody fragments specifically bind to a tumor antigen, and Z is selected from taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, nitrogen mustards, maytansinoids, calicheamycins, duocarmycins, tubulysines, amatoxins, dolastatins, auristatins, enediynes, pyrrolobenzodiazepines, and ethylenimines. The disclosure also relates to TGase-mediated conjugation concepts, antibody-drug conjugates, and the engineering of acceptor glutamine reactivity in immunoglobulins.
Claims Coverage
The consolidated claim coverage centers on antibody or antibody fragment compositions with the same primary amino acid sequence and tumor antigen binding, where at least 90% of the population carries functionalized acceptor glutamine residues in the constant region. The independent claim features converge on defined Formula IVb or lysine-based linker structures, the RR2 addition-product linkage concept, m selected from 2 or 4, and payload selection through Z.
Population with same primary amino acid sequence and high functionalized acceptor glutamine occupancy
A composition comprising a population of antibodies or antibody fragments having the same primary amino acid sequence, where at least 90% of the antibodies or antibody fragments have (m) functionalized acceptor glutamine residues (Q) per antibody or fragment, with m selected from 2 or 4.
Functionalized acceptor glutamine residues in the constant region
Each functionalized acceptor glutamine residue has the structure of Formula IVb or a related lysine-based linker form and is a glutamine residue present within or substituted into the primary amino acid sequence of a constant region of the antibodies or antibody fragments.
Defined linker architecture with reactive moiety addition product
The functionalized acceptor glutamine residues include structured components with linker and spacer system elements, where RR2 is an addition product between a reactive moiety R and a complementary reactive moiety R2, and where V and Y may be absent, non-cleavable, or conditionally-cleavable moieties or spacer system components.
Tumor antigen binding and Z payload classes
The antibodies or antibody fragments specifically bind to a tumor antigen, and Z is selected from taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, nitrogen mustards, maytansinoids, calicheamycins, duocarmycins, tubulysines, amatoxins, dolastatins, auristatins, enediynes, pyrrolobenzodiazepines, and ethylenimines.
Heavy-chain localized functionalized glutamine and sequence constraints
At least 90% of the antibodies or antibody fragments comprise on each heavy chain at least one functionalized acceptor glutamine residue having the structure of Formula IVb, with narrower embodiments including EU residue numbering, N297X substitution constraints, and EU position 295 variants.
The independent claims cover same-primary-sequence antibody or antibody-fragment populations that bind to a tumor antigen and are functionalized at constant-region acceptor glutamine residues using defined Formula IVb or lysine-based linker structures. The claims further specify at least 90% occupancy, m selected as 2 or 4, heavy-chain-focused embodiments, and payload selection via Z from a defined group.
Stated Advantages
Enables near-complete loading with narrow drug-antibody ratio (DAR) distributions.
Addresses partial and heterogeneous conjugation with large, charged, or hydrophobic payloads.
Provides fully and stoichiometrically conjugated acceptor glutamines after removing N297-linked glycosylation.
Documented Applications
Antibody-drug conjugates (ADCs) formed by transglutaminase (TGase/BTG)-mediated, site-specific functionalization of immunoglobulins for tumor antigen binding.
Pharmaceutical composition and method of treating disease.
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