Enzymatic conjugation of antibodies

Patent metadata

Field	Value
Publication number	US-10675359-B2
Application number	US-201715702586-A
Country	US
Kind code	B2
Filing date	Sep 12, 2017
Priority date	Dec 23, 2011
Publication date	Jun 9, 2020
Grant date	Jun 9, 2020

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.

First claim

The invention claimed is:

1. A human or humanized antibody comprising one or more solvent-exposed glutamine residues in a variable region and at least one acceptor glutamine residue in its constant region or in a sequence fused to a variable or constant region, wherein said one or more solvent-exposed glutamine residues are present in a light chain variable domain (VL) CDR at a Kabat position selected from the group consisting of 27, 55, and a combination thereof,
wherein said antibody is conjugated via said acceptor glutamine residue to one or more moieties-of-interest (Z) through a linker that comprises a NH—(C) n — moiety,
wherein
(C) n is a substituted or unsubstituted alkyl or heteroalkyl chain, optionally wherein any carbon of the chain is substituted with an alkoxy, hydroxyl, alkylcarbonyloxy, alkyl-S—, thiol, alkyl-C(O)S—, amine, alkylamine, amide, or alkylamide, and n is an integer selected from among the range of 2 to 20;
Z is a reactive moiety or a moiety that improves the pharmacokinetic properties, a therapeutic moiety or a diagnostic moiety, and
wherein the antibody comprises heavy chain variable region sequence selected from the group consisting of SEQ ID NOs: 3-239 and light chain variable region sequence selected from group consisting of SEQ ID NO: 240-299.

2. The antibody of claim 1 , wherein the antibody is a tetrameric antibody and said constant region is a heavy chain constant region.

3. The antibody of claim 1 , wherein the antibody comprises:
a heavy chain framework region 1 (FR-H1) comprising a glutamine residue at Kabat position 1, 3, 5, 6, 10, 11, 12, 13 and/or 16;
a heavy chain framework region 2 (FR-H2) comprising a glutamine residue at Kabat position 38, 39, 43 and/or 45;
a heavy chain framework region 3 (FR-H3) comprising a glutamine residue at Kabat position 66, 75, 77, 81 and/or 85;
a heavy chain framework region 1 (CDR-H1) comprising a glutamine residue at Kabat position 26 and/or 35; and/or
a heavy chain framework region 2 (CDR-H2) comprising a glutamine residue at Kabat position 50, 52, 56, 61 and/or 64, wherein said glutamine residue is not an acceptor glutamine.

4. A composition comprising a plurality of human or humanized antibodies of claim 1 , wherein each of said human or humanized antibodies comprises one acceptor glutamine on each heavy chain, wherein at least 80% of the antibodies in the composition comprise on each heavy chain one functionalized acceptor glutamine residue (Q) having Formula IVa,
(Q)-NH—(C) n —X-L- (V—(Y—(Z) z ) q ) r   Formula IVa,

or a pharmaceutically acceptable salt or solvate thereof,
wherein:
Q is a glutamine residue present in the human or humanized antibody;
(C) n is a substituted or unsubstituted alkyl or heteroalkyl chain, optionally wherein any carbon of the chain is substituted with an alkoxy, hydroxyl, alkylcarbonyloxy, alkyl-S—, thiol, alkyl-C(O)S—, amine, alkylamine, amide, or alkylamide;
n is an integer selected from among the range of 2 to 20;
X is NH, O, S, absent, or a bond;
L is independently absent, a bond or a continuation of a bond, or a carbon comprising framework of 5 to 200 atoms substituted at one or more atoms;
r is an integer selected from among 1, 2, 3 or 4;
q is an integer selected from among 1 , 2, 3 or 4;
z is an integer selected from among 1, 2, 3 or 4; and
V is independently absent, a bond or a continuation of a bond, a non-cleavable moiety or a conditionally-cleavable moiety;
Y is independently absent, a bond or a continuation of a bond, or a spacer system which is comprised of 1 or more spacers; and
Z is a moiety that improves pharmacokinetic properties, a therapeutic moiety or a diagnostic moiety.

5. A composition comprising a plurality of human or humanized antibodies of claim 1 , wherein each of said human or humanized antibodies comprises one acceptor glutamine on each heavy chain, wherein at least 80% of the antibodies in the composition comprise on each heavy chain two functionalized acceptor glutamine residues (Q) having Formula IVb,
(Q)-NH—(C) n —X-L-(V—(Y-(M) z ) q ) r   Formula IVb

or a pharmaceutically acceptable salt or solvate thereof,
wherein:
Q is a glutamine residue present in the human or humanized antibody;
(C) n is a substituted or unsubstituted alkyl or heteroalkyl chain, optionally wherein any carbon of the chain is substituted with an alkoxy, hydroxyl, alkylcarbonyloxy, alkyl-S—, thiol, alkyl-C(O)S—, amine, alkylamine, amide, or alkylamide;
n is an integer selected from among the range of 2 to 20;
X is NH, O, S, absent, or a bond;
L is independently absent, a bond or a continuation of a bond, or a carbon comprising framework of 5 to 200 atoms substituted at one or more atoms;
r is an integer selected from among 1, 2, 3 or 4;
q is an integer selected from among 1 , 2, 3 or 4;
z is an integer selected from among 1, 2, 3 or 4; and
V is independently absent, a bond or a continuation of a bond, a non-cleavable moiety or a conditionally-cleavable moiety;
Y is independently absent, a bond or a continuation of a bond, or a spacer system which is comprised of 1 or more spacers; and
M is independently: R or (RR′) -L′-(V′—(Y′—(Z)z′)q′)r′), wherein R is a reactive moiety, wherein each of L′, V′, Y′, z′, q′, and r′ are as defined for L, V, Y, z, q, and r, wherein RR′ is an addition product between a reactive moiety R and a complementary reactive moiety R′, and wherein Z is a moiety that improves pharmacokinetic properties, a therapeutic moiety or a diagnostic moiety.

6. The composition of claim 5 , wherein the composition has a mean Z:antibody ratio of at least 1.5, wherein less than 10% of the antibodies comprise more than two moieties of interest (Z) per antibody and less than 25% comprise less than two moieties of interest (Z) per antibody.

7. A composition comprising a plurality of human or humanized antibodies of claim 1 , wherein each of said human or humanized antibodies is linked to a moiety of interest (Z), wherein the composition has a mean Z: antibody ratio of at least 1.5 wherein less than 10% of the antibodies comprise more than two moieties of interest (Z) per antibody.

8. The composition of claim 7 , wherein less than 25% comprise less than two moieties of interest (Z) per antibody.

9. The composition of claim 7 , wherein the antibodies are linked to said moiety of interest (Z) via one functionalized acceptor glutamine on each heavy chain of the antibody.

10. A composition comprising a plurality of human or humanized antibodies of claim 1 , wherein at least 80% of the antibodies in the composition comprise on each heavy chain one functionalized acceptor glutamine residue (Q) having Formula IVa or Formula IVb,
(Q)-NH—(C) n —X-L-(V—(Y—(Z) z ) q ) r   Formula IVa;

(Q)-NH—(C) n —X-L-(V—(Y-(M) z ) q ) r   Formula IVb,

or a pharmaceutically acceptable salt or solvate thereof,
wherein:
Q is a glutamine residue present in the human or humanized antibody;
(C) n is a substituted or unsubstituted alkyl or heteroalkyl chain, optionally wherein any carbon of the chain is substituted with an alkoxy, hydroxyl, alkylcarbonyloxy, alkyl-S—, thiol, alkyl-C(O)S—, amine, alkylamine, amide, or alkylamide;
n is an integer selected from among the range of 2 to 20;
X is NH, O, S, absent, or a bond;
L is independently absent, a bond or a continuation of a bond, or a carbon comprising framework of 5 to 200 atoms substituted at one or more atoms;
r is an integer selected from among 1, 2, 3 or 4;
q is an integer selected from among 1 , 2, 3 or 4;
z is an integer selected from among 1, 2, 3 or 4; and
V is independently absent, a bond or a continuation of a bond, a non-cleavable moiety or a conditionally-cleavable moiety;
Y is independentl

Assignees

• Innate Pharma
• Scherrer Inst Paul

Inventors

• Dennler Patrick
• Bregeon Delphine
• Gauthier Laurent
• Romagné François
• Belmant Christian
• Fischer Eliane
• Schibli Roger

Classifications

• C07F9/5022

  Aromatic phosphines (P-C aromatic linkage) · CPC title

• A61K47/6817

  Toxins · CPC title

• C07D257/08

  Six-membered rings · CPC title

• C12P21/005

  Glycopeptides, glycoproteins · CPC title

• C07C323/60

  with the carbon atom of at least one of the carboxyl groups bound to nitrogen atoms · CPC title