Synaffix B.v.

Remon Van Geel

Floris Louis Van Delft

Maria Antonia Wijdeven

Sander Sebastiaan Van Berkel

Jorge Merijn Mathieu Verkade

Ryan Heesbeen

Inge Catharina Josephina Hurkmans

Petrus Josephus Jacobus Maria Van De Sande

Brian Maria Gerardus Janssen

Lianne Lelieveldt

The present invention concerns antibody-conjugates which are especially suitable for the targeting of PTK7-expressing cells, in particular tumour cells. The antibody-conjugates according to the invention have structure (1):-[()-{--]  1Herein, AB is an antibody capable of targeting PTK7-expressing tumours; L is a linker that links Z to D; Z is a connecting group; Lis -GlcNAc(Fuc)-(G)-S-(L)-, wherein G is a monosaccharide, j is an integer in the range of 0-10, S is a sugar or a sugar derivative, GlcNAc is N-acetylglucosamine and Fuc is fucose, w is 0 or 1, w' is 0, 1 or 2 and Lis —N(H)C(O)CH—, —N(H)C(O)CF— or —CH—; D is selected from the group consisting of anthracyclines, camptothecins, tubulysins, enediynes, amanitins, duocarmycins, maytansinoids, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof; b is 0 or 1; x is 1 or 2; and y is 1, 2, 3 or 4. The invention further concerns a method for preparing the antibody-conjugates of structure (1) and application of the antibody-conjugates of structure (1).

Antibody-conjugates for targeting of tumours expressing PTK7

The present invention relates to an antibody comprising a GlcNAc-S(A)substituent, wherein S(A)is a sugar derivative comprising x functional groups A wherein A is independently selected from the group consisting of an azido group, a keto group and an alkynyl group and x is 1, 2, 3 or 4, wherein said GlcNAc-S(A)substituent is bonded to the antibody via C1 of the N-acetylglucosamine of said GlcNAc-S(A)substituent, and wherein said N-acetylglucosamine is optionally fucosylated. The invention also relates to an antibody-conjugate, in particular to an antibody-conjugate according to the Formula (20) or (20), wherein AB is an antibody, S is a sugar or a sugar derivative, D is a molecule of interest, and wherein said N-acetylglucosamine is optionally fucosylated (b is 0 or 1). The invention further relates to a process for the preparation of a modified antibody, to a process for the preparation of an antibody-conjugate, and to said antibody-conjugate for use as a medicament. In addition, the invention relates to a kit of parts comprising an azide-modified antibody and a linker-conjugate, wherein said linker-conjugate comprises a (hetero)cycloalkynyl group and one or more molecules of interest.

Modified antibody, antibody-conjugate and process for the preparation thereof

The present invention relates to a process for the enzymatic modification of a glycoprotein. The process comprises the step of contacting a glycoprotein comprising a glycan comprising a terminal GlcNAc-moiety, in the presence of glycosyltransferase that is, or is derived from, a β-(1,4)-N-acetylgalactosaminyltransferase, with a non-natural sugar-derivative nucleotide. The non-natural sugar-derivative nucleotide is according to formula (3): wherein A is selected from the group consisting of —N, —C(O)R, —(CH)C≡C—R, —SH, —SC(O)R, —SC(O)OR, —SC(S)OR, —F, —Cl, —Br —I, —OS(O)R, terminal C-Calkenyl groups, C-Ccycloalkenyl groups, C-Calkadienyl groups, terminal C-Callenyl groups and amino groups. The invention further relates to a glycoprotein obtainable by the process according to the invention, to a bioconjugate that can be obtained by conjugating the glycoprotein with a linker-conjugate, and to β-(1,4)-N-acetylgalactosaminyltransferases that can be used in preparing the glycoprotein according to the invention.

Process for the modification of a glycoprotein using a glycosyltransferase that is or is derived from a β(1,4)-n-acetylgalactosaminyltransferase

The present invention is based on the surprising finding that the linker employed in a bioconjugate, such as an anti-body-drug conjugate, is not therapeutically inert but has an effect on the therapeutic index of the bioconjugate. The present invention thus concerns a method for increasing the therapeutic index of a bioconjugate and the bioconjugates for use in treatment, in particular cancer. The bioconjugates according to the invention have a sulfamide linker comprising a group according to formula (1).

Bioconjugates containing sulfamide linkers for use in treatment

The invention concerns fusion proteins, wherein two endoglycosidases are fused, possibly via a linker. The fusion enzymes according to the invention have structure (1): EndoX-(L)-EndoY (1), wherein EndoX is an endoglycosidase, EndoY is an endoglycosidase distinct from EndoX, L is a linker and p is 0 or 1. Such fusion enzymes capable of trimming glycoproteins comprising at least two distinct glycoforms in a single step. The invention further concerns the use of the fusion enzyme according to the invention for trimming glycoproteins. In another aspect, the invention relates to the process of production of the fusion enzyme. In a further aspect, the inventions concerns a process for trimming glycoproteins, comprising trimming the glycoprotein with a fusion enzyme according to the invention, to obtain a trimmed glycoprotein.

Enzymes for trimming of glycoproteins

The present invention concerns novel and improved antibody-conjugates for targeting HER2. The inventors found N that when antibody-conjugates were prepared using a specific mode of conjugation, they exhibit an improved therapeutic index. The mode of conjugation comprises a first step (i) of contacting a glycoprotein comprising 1-4 core N-acetylglucosamine moieties with a compound of the formula S(F)-P in the presence of a catalyst, wherein S(F)is a sugar derivative comprising x functional groups F1 capable of reacting with a functional group Q, x is 1 or 2 and P is a nucleoside mono- or diphosphate, and wherein the catalyst is capable of transferring the S(F)moiety to the core-GlcNAc moiety, to obtain a modified antibody; and a second step (ii) of reacting the modified antibody with a linker-conjugate comprising a functional group Qcapable of reacting with functional group Fand a target molecule D connected to Qvia a linker Lto obtain the antibody-conjugate wherein linker L comprises S&#x2014;Z-Land wherein Zis a connecting group resulting from the reaction between Qand F. The invention also relates to a use for improving the therapeutic index of an antibody-conjugate and to a method for targeting HER2-expressing cells.

Antibody-conjugates with improved therapeutic index for targeting HER2 tumours and method for improving therapeutic index of antibody-conjugates

The present invention relates to a process for the enzymatic modification of a glycoprotein. The process comprises the step of contacting a glycoprotein comprising a glycan comprising a terminal GlcNAc-moiety, in the presence of glycosyltransferase that is, or is derived from, a &#x3b2;-(1,4)-N-acetylgalactosaminyltransferase, with a non-natural sugar-derivative nucleotide. The non-natural sugar-derivative nucleotide is according to formula (3): wherein A is selected from the group consisting of &#x2014;N, &#x2014;C(O)R, &#x2014;(CH)C&#x2261;C&#x2014;R, &#x2014;SH, &#x2014;SC(O)R, &#x2014;SC(O)OR, &#x2014;SC(S)OR, &#x2014;F, &#x2014;Cl, &#x2014;Br &#x2014;I, &#x2014;OS(O)R, terminal C-Calkenyl groups, C-Ccycloalkenyl groups, C-Calkadienyl groups, terminal C-Callenyl groups and amino groups. The invention further relates to a glycoprotein obtainable by the process according to the invention, to a bioconjugate that can be obtained by conjugating the glycoprotein with a linker-conjugate, and to &#x3b2;-(1,4)-N-acetylgalactosaminyltransferases that can be used in preparing the glycoprotein according to the invention.

Process for the modification of a glycoprotein using a glycosyltransferase that is or is derived from a &#x3b2;(1,4)-n-acetylgalactosaminyltransferase

The present invention relates to an antibody comprising a GlcNAc-S(A)substituent, wherein S(A)is a sugar derivative comprising x functional groups A wherein A is independently selected from the group consisting of an azido group, a keto group and an alkynyl group and x is 1, 2, 3 or 4, wherein said GlcNAc-S(A)substituent is bonded to the antibody via CI of the N-acetylglucosamine of said GlcNAc-S(A)substituent, and wherein said N-acetylglucosamine is optionally fucosylated. The invention also relates to an antibody-conjugate, in particular to an antibody-conjugate according to the Formula (20) or (20b), wherein AB is an antibody, S is a sugar or a sugar derivative, D is a molecule of interest, and wherein said N-acetylglucosamine is optionally fucosylated (b is 0 or 1). The invention further relates to a process for the preparation of a modified antibody, to a process for the preparation of an antibody-conjugate, and to said antibody-conjugate for use as a medicament. In addition, the invention relates to a kit of parts comprising an azide-modified antibody and a linker-conjugate, wherein said linker-conjugate comprises a (hetero)cycloalkynyl group and one or more molecules of interest. 

The present invention relates to a cycloaddition process comprising the step of reacting a halogenated aliphatic 1,3-dipole compound with a (hetero)cycloalkyne according to Formula (1): Preferably, the (hetero)cycloalkyne according to Formula (1) is a (hetero)cyclooctyne. The invention also relates to the cycloaddition products obtainable by the process according to the invention. The invention further relates to halogenated aliphatic 1,3-dipole compounds, in particular to halogenated aliphatic 1,3-dipole compounds comprising N-acetylgalactosamine-UDP (GalNAc-UDP), and to halogenated 1,3-dipole compounds comprising (peracylated) N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), N-acetylmannosamine (ManNAc) and N-acetyl neuraminic acid (NeuNAc). 

Process for the cycloaddition of a halogenated 1,3-dipole compound with a (hetero)cycloalkyne

The invention relates to a glycoprotein comprising an optionally fucosylated glycan according to formula (105) or (106), wherein Su(A)x is a modified sugar moiety comprising one or more functional groups A. Functional group A is independently selected from the group consisting of a thiol group, a halogen, a sulfonyloxy group, a halogenated acetamido group, a mercaptoacetamido group and a sulfonated hydroxyacetamido group. The invention also relates to a glycoprotein-conjugate wherein a glycoprotein according to the invention is conjugated to a molecule of interest. Said molecule of interest may for example be an active substance. The invention further relates to a process for the preparation of a modified glycoprotein, and to a method for the preparation of a glycoprotein-conjugate. The invention particularly relates to modified antibodies, antibody-conjugates, antibody-drug conjugates and methods for the preparation thereof. 

Modified glycoprotein, protein-conjugate and process for the preparation thereof

Glycoproteins comprising a glycan of the formula (102) are disclosed; wherein b is 0 or 1; the GlcNAc residue optionally fucosylated; and Su(A)is a sugar derivative comprising x functional groups A, wherein x is 1, 2, 3 or 4 and A is independently selected from the group consisting of an azido group, a keto group, an alkynyl group, a thiol group, a halogen, a sulfonyloxy group, a halogenated acetamido group, a mercaptoacetamido group and a sulfonylated hydroxyacetamido group.  Protein-conjugates having glycoproteins according to the invention conjugated to a molecule of interest (e.g., an active substance) are also disclosed. Examples include modified antibodies, antibody-conjugates, and antibody-drug conjugates (ADCs). Processes for the preparation of the modified glycoproteins according to the invention and methods for the preparation of a protein-conjugate according to the invention are mentioned.

The present invention relates to a process for the enzymatic modification of a glycoprotein. The process comprises the step of contacting a glycoprotein comprising a glycan comprising a terminal GlcNAc-moiety, in the presence of glycosyltransferase that is, or is derived from, a &#x3b2;-(1,4)-N-acetylgalactosaminyltransferase, with a non-natural sugar-derivative nucleotide. The non-natural sugar-derivative nucleotide is according to formula (3): wherein A is selected from the group consisting of &#x2014;N, &#x2014;C(O)R, &#x2014;(CH)C&#x2261;C&#x2014;R, &#x2014;SH, &#x2014;SC(O)R, &#x2014;SC(O)OR8, &#x2014;SC(S)OR8, &#x2014;F, &#x2014;CI, &#x2014;Br &#x2014;I, &#x2014;OS(O)R, terminal C-Calkenyl groups, C-Ccycloalkenyl groups, C-Calkadienyl groups, terminal C-Callenyl groups and amino groups. The invention further relates to a glycoprotein obtainable by the process according to the invention, to a bioconjugate that can be obtained by conjugating the glycoprotein with a linker-conjugate, and to &#x3b2;-(1,4)-N-acetylgalactosaminyltransferases that can be used in preparing the the glycoprotein according to the invention. 

Process for the modification of a glycoprotein using a glycosyltransferase that is or is derived from A &#x3b2;(1,4)-N-acetylgalactosaminyltransferase

An antibody comprising a GlcNAc&#x2014;S(A)substituent is disclosed, wherein S(A)is a sugar derivative comprising x functional groups A wherein A is independently selected from the group consisting of an azido group, a keto group and an alkynyl group and x is 1, 2, 3 or 4, wherein said GlcNAc&#x2014;S(A)substituent is bonded to the antibody via C1 of the N-acetylglucosamine of said GlcNAc&#x2014;S(A)substituent, and wherein said N-acetylglucosamine is optionally fucosylated. Also disclosed is an antibody-conjugate, in particular to an antibody-conjugate according to the Formula (20) or (20b), wherein AB is an antibody, S is a sugar or a sugar derivative, D is a molecule of interest, and wherein said N-acetylglucosamine is optionally fucosylated (b is 0 or 1). Also disclosed is a process for the preparation of a modified antibody, to a process for the preparation of an antibody-conjugate, and to said antibody-conjugate for use as a medicament.

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Lith-Oijen, Netherlands

Remon Van Geel