PHARMACEUTICAL COMPOSITIONS COMPRISING BISPECIFIC ANTIBODIES BINDING TO B7H4 AND CD3

1 . A pharmaceutical composition comprising:

a) an antibody comprising an antigen-binding region capable of binding to human B7H4 and an antigen-binding region capable of binding to human CD3,

b) a buffering agent,

wherein the pH of the composition is from 4.0 to 8.0.

2 . A pharmaceutical composition according to claim 1 , wherein said antigen-binding regions comprise heavy and light chain variable regions, wherein said heavy and light chain variable regions are humanized and/or human.

3 . (canceled)

4 . A pharmaceutical composition according to claim 1 , wherein the buffering agent is selected from the group consisting of histidine, glutamate, and mixtures thereof.

5 . A pharmaceutical composition according to claim 1 , wherein the pharmaceutical composition further comprises c) a non-ionic excipient, such as a sugar or a sugar alcohol, such as sorbitol, sucrose or mixtures thereof.

6 . (canceled)

7 . A pharmaceutical composition according to claim 5 , wherein the non-ionic excipient is present at a concentration of 100 to 300 mM, such as 125-250 mM preferably 250 mM.

8 . A pharmaceutical composition according to claim 1 , wherein the pharmaceutical composition further comprises d) a surfactant.

9 . A pharmaceutical composition according to claim 8 , wherein the surfactant is selected from the group consisting of glycerol monooleate, benzethonium chloride, sodium docusate, phospholipids, polyethylene alkyl ethers, sodium lauryl sulfate and tricaprylin, benzalkonium chloride, citrimide, cetylpyridinium chloride and phospholipids, alpha tocopherol, glycerol monooleate, myristyl alcohol, phospholipids, poloxamers, polyoxyethylene alkyl ethers, polyoxyethylene castor oil derivatives, polyoxyethylene sorbintan fatty acid esters, polyoxyethylene sterarates, polyoxyl hydroxystearate, polyoxylglycerides, polysorbates, propylene glycol dilaurate, propylene glycol monolaurate, sorbitan esters sucrose palmitate, sucrose stearate, tricaprylin and TPGS, and mixtures thereof.

10 . (canceled)

11 . A pharmaceutical composition according to claim 8 , wherein the surfactant is present at a concentration from about 0.005% to 0.4% w/v, such as from about 0.01 to 0.1% w/v, such as from about 0.01 to 0.09% w/v such as from about 0.01 to 0.06% w/v such as from about 0.01 to 0.05% w/v such as 0.02% w/v or 0.03% w/v or 0.04% w/v or 0.05% w/v, preferably 0.02% w/v.

12 . A pharmaceutical composition according to claim 1 , wherein the concentration of the antibody is 0.5 to 100 mg/ml, such as 1.0 to 50 mg/ml, or such as 5 to 30 mg/ml, such as 5 mg/ml, or 6 mg/ml, or 7 mg/ml, or 8 mg/ml, or 9 mg/ml, or 10 mg/ml, or 11 mg/ml, or 12 mg/ml, or 13 mg/ml, or 14 mg/ml, or 15 mg/ml, or 16 mg/ml, or 17 mg/ml, or 18 mg/ml, or 19 mg/ml, or 20 mg/ml, or 21 mg/ml, or 22 mg/ml, or 23 mg/ml, or 24 mg/ml, or 25 mg/ml, or 26 mg/ml, or 27 mg/ml, or 28 mg/ml, or 29 mg/ml, 30 mg/ml, 31 mg/ml, 32 mg/ml, 33 mg/ml, 34 mg/ml, 35 mg/ml, 36 mg/ml, 37 mg/ml, 38 mg/ml, 39 mg/ml, 40 mg/ml, 41 mg/ml, 42 mg/ml, 43 mg/ml, 44 mg/ml, 45 mg/ml, 46 mg/ml, 47 mg/ml, 48 mg/ml, 49 mg/ml, 50 mg/ml, 51 mg/ml, 52 mg/ml, 53 mg/ml, 54 mg/ml, 55 mg/ml, 56 mg/ml, 57 mg/ml, 58 mg/ml, 59 mg/ml, or such as 60 mg/ml.

13 . A pharmaceutical composition according to claim 1 , wherein the buffering agent is present at a concentration of 5 to 40 mM, such as 10-30 mM, preferably 20 mM.

14 . A pharmaceutical composition according to claim 1 , wherein the pharmaceutical composition is an aqueous composition.

15 . A pharmaceutical composition according to claim 1 , comprising:

a) 60_mg/ml of the antibody,

b) 10 to 20 mM glutamate or histidine,

c) 150 to 350 mM sorbitol or sucrose,

d) a polysorbate,

wherein the pH of the composition is from 5.0 to 6.0.

16 . A pharmaceutical composition according to claim 1 , selected from the group consisting of

(i) a pharmaceutical composition comprising a) 20-60 mg/ml of the antibody, b) 20 mM glutamate, c) 250 mM sorbitol, d) 0.02% w/v polysorbate 80, wherein the pH of the composition is from 5.1-5.3, and

(ii) a pharmaceutical composition comprising a) 20-60 mg/ml of the antibody, b) 20 mM histidine, c) 250 mM sucrose, d) 0.02% w/v polysorbate 80, wherein the pH of the composition is from 5.4-5.6.

17 - 21 . (canceled)

22 . A pharmaceutical composition according to claim 1 , wherein the antigen-binding region that binds to CD3 comprises:

a heavy chain variable region (VH) comprising the CDR1, CDR2, and CDR3 regions of SEQ ID NO. 16 or 17,

and,

a light chain variable region (VL) comprising the CDR1, CDR2, and CDR3 regions of SEQ ID NO: 22.

23 . A pharmaceutical composition according to claim 1 , wherein the antigen-binding region that binds to CD3 comprises:

a heavy chain variable region (VH) comprising the CDR1, CDR2, and CDR3 sequences of SEQ ID NOs.: 18, 19 and 21 or

a heavy chain variable region (VH) comprising the CDR1, CDR2, and CDR3 sequences of SEQ ID NOs.: 18, 19 and 20,

and,

a light chain variable region (VL) comprising the CDR1, CDR2, and CDR3 sequences of SEQ ID NO: 23, GTN and 24, respectively.

24 . A pharmaceutical composition according to claim 1 , wherein the antigen-binding region that binds to CD3 comprises:

a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 16 or 17, or a sequence having at least 90%, at least 95%, at least 97%, or at least 99% amino acid sequence identity to the sequence of SEQ ID NO: 16 or 17;

and,

a light chain variable region (VL) comprising the sequence of SEQ ID NO: 22 or a sequence having at least 90%, at least 95%, at least 97%, or at least 99% amino acid sequence identity to the sequence of SEQ ID NO: 22.

25 . A pharmaceutical composition according to claim 1 , wherein the antigen-binding region capable of binding to human B7H4 comprises:

a) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions of SEQ ID NO. 25 and a variable light chain region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NO. 33;

b) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions of SEQ ID NO. 29 and a variable light chain region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NO. 33;

c) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions of SEQ ID NO. 36 and a variable light chain region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NO. 40;

d) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions of SEQ ID NO. 43 and a variable light chain region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NO. 47;

e) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions of SEQ ID NO. 50 and a variable light chain region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NO. 54;

f) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions of SEQ ID NO. 31 and a variable light chain region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NO. 33; or

g) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions of SEQ ID NO. 65 and a variable light chain region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NO. 69.

26 . (canceled)

27 . A pharmaceutical composition according to claim 1 , wherein the antigen-binding region capable of binding to human B7H4 comprises:

a) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NOs.: 26, 27 and 28, and a variable light chain region comprising the CDR1, CDR2 and CDR3 respectively of SEQ ID NO. 34, GAS and SEQ ID NO. 35;

b) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NOs.: 26, 30 and 28, and a variable light chain region comprising the CDR1, CDR2 and CDR3 respectively of SEQ ID NO. 34, GAS and SEQ ID NO. 35;

c) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NOs.: 37, 38 and 39, and a variable light chain region comprising the CDR1, CDR2 and CDR3 respectively of SEQ ID NO. 41, DTS and SEQ ID NO. 42;

d) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NOs.: 44, 45 and 46, and a variable light chain region comprising the CDR1, CDR2 and CDR3 respectively of SEQ ID NO. 48, YTS and SEQ ID NO. 49;

e) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NOs.: 51, 52 and 53, and a variable light chain region comprising the CDR1, CDR2 and CDR3 respectively of SEQ ID NO. 55, GAS and SEQ ID NO. 56;

f) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NOs.: 26, 32 and 28, and a variable light chain region comprising the CDR1, CDR2 and CDR3 respectively of SEQ ID NO. 34, GAS and SEQ ID NO. 35; or

g) a variable heavy chain (VH) region comprising the CDR1, CDR2 and CDR3 regions respectively of SEQ ID NOs.: 66, 67 and 68, and a variable light chain region comprising the CDR1, CDR2 and CDR3 respectively of SEQ ID NO. 70, GAS and SEQ ID NO. 71.

28 . (canceled)

29 . A pharmaceutical composition according to claim 1 , wherein the antigen-binding region capable of binding to human B7H4 comprises:

a) a variable heavy chain (VH) region of SEQ ID NO. 25 and a variable light chain region of SEQ ID NO. 33;

b) a variable heavy chain (VH) region of SEQ ID NO. 29 and a variable light chain region of SEQ ID NO. 33;

c) a variable heavy chain (VH) region of SEQ ID NO. 36—and a variable light chain region of SEQ ID NO. 40;

d) a variable heavy chain (VH) region of SEQ ID NO. 43 and a variable light chain region of SEQ ID NO. 47;

e) a variable heavy chain (VH) region of SEQ ID NO. 50 and a variable light chain region of SEQ ID NO. 54;

f) a variable heavy chain (VH) region of SEQ ID NO. 29 and a variable light chain region of SEQ ID NO. 33; or

g) a variable heavy chain (VH) region of SEQ ID NO. 65 and a variable light chain region of SEQ ID NO. 69.

30 . (canceled)

31 . A pharmaceutical composition according to claim 1 , wherein the antibody comprises an antigen-binding region capable of binding to human B7H4 and an antigen-binding region capable of binding to human CD3, wherein the antigen-binding region that binds to CD3 comprises

a heavy chain variable region (VH), wherein CDR1 comprises the amino acid sequence of SEQ ID NO. 18, wherein CDR2 comprises the amino acid sequence of SEQ ID NO. 19, and wherein CDR3 comprises the amino acid sequence of SEQ ID NO. 20 or 21, and a light chain variable region (VL), wherein CDR1 comprises the amino acid sequence of SEQ ID NO. 23, wherein CDR2 comprises the amino acid sequence of GTN, and wherein CDR3 comprises the amino acid sequence of SEQ ID NO: 24, and wherein the antigen-binding region capable of binding to B7H4 comprises:

a variable heavy chain (VH) region, wherein comprises the amino acid sequence of SEQ ID NO. 26, wherein CDR2 comprises the amino acid sequence of SEQ ID NO. 30, and wherein CDR3 comprises the amino acid sequence of SEQ ID NO. 28, and

a light chain variable region (VL), wherein CDR1 comprises the amino acid sequence of SEQ ID NO. 34, wherein CDR2 comprises the amino acid sequence of GAS, and wherein CDR3 comprises the amino acid sequence of SEQ ID NO: 35.

32 . (canceled)

33 . A pharmaceutical composition according to claim 1 , wherein the antibody comprises an antigen-binding region capable of binding to human B7H4 and an antigen-binding region capable of binding to human CD3, wherein the antigen-binding region that binds to CD3 comprises a variable heavy chain (VH) region of SEQ ID NO: 16 or 17 and a variable light chain region of SEQ ID NO: 22 and wherein the antigen-binding region capable of binding to human B7H4 comprises a variable heavy chain (VH) region of SEQ ID NO: 29: and a variable light chain region of SEQ ID NO: 33.

34 . (canceled)

35 . A pharmaceutical composition according to claim 1 , wherein the antibody comprises two heavy chain constant regions (CH), and two light chain constant regions (CL), preferably wherein the two heavy chain constant domains, and two light chain constant regions are derived from human.

36 - 37 . (canceled)

38 . A pharmaceutical composition according to claim 1 , wherein the antibody comprises a first and a second heavy chain, each of said first and second heavy chain comprises at least a hinge region, a CH2 and CH3 region, wherein in said first heavy chain at least one of the amino acids in the positions corresponding to positions selected from the group consisting of T366, L368, K370, D399, F405, Y407 and K409 in a human IgG1 heavy chain has been substituted, and in said second heavy chain at least one of the amino acids in the positions corresponding to a position selected from the group consisting of T366, L368, K370, D399, F405, Y407, and K409 in a human IgG1 heavy chain has been substituted, wherein said substitutions of said first and said second heavy chains are not in the same positions, and wherein the amino acid positions are numbered according to Eu numbering.

39 . A pharmaceutical composition according to claim 38 , wherein the amino acid in the position corresponding to K409 in a human IgG1 heavy chain is R in said first heavy chain, and the amino acid in the position corresponding to F405 in a human IgG1 heavy chain is L in said second heavy chain, or wherein the amino acid in the position corresponding to F405 in a human IgG1 heavy chain is L in said first heavy chain, and the amino acid in the position corresponding to K409 in a human IgG1 heavy chain is R in said second heavy chain.

40 . (canceled)

41 . A pharmaceutical composition according to claim 1 , wherein the antibody comprises a first and a second heavy chain, and wherein in both the first and the second heavy chain,

(i) the amino acid residues at the positions corresponding to positions L234 and L235 in a human IgG1 heavy chain according to Eu numbering are F and E, respectively,

(ii) the amino acid residue at the position corresponding to position D265 in a human IgG1 heavy chain according to Eu numbering is A, or

(iii) the amino acid residues at the positions corresponding to positions L234, L235, and D265 in a human IgG1 heavy chain according to Eu numbering are F, E, and A, respectively.

42 . (canceled)

43 . A pharmaceutical composition according to claim 1 , wherein the antibody comprises a lambda (k) light chain and a kappa (x) light chain; e.g. an antibody with a heavy chain and a lambda light chain which comprise the binding region capable of binding to CD3, and a heavy chain and a kappa light chain which comprise the binding region capable of binding to B7H4.

44 . A pharmaceutical composition according to claim 1 , wherein the antigen binding region capable of binding to human B7H4 is comprised in an heavy chain and a light chain, said heavy chain comprising said VH region and an IgG1 heavy chain constant region and said light chain comprising said VL region and a kappa light chain constant region; and wherein said antigen binding region capable of binding to human CD3 is comprised in a heavy chain and a light chain, said heavy chain comprising said VH region and an IgG1 heavy chain constant region and said light chain comprising said VL region and a lambda light chain constant region.

45 . A pharmaceutical composition according to claim 44 , wherein one IgG1 heavy chain constant region is as defined in SEQ ID NO. 60 and the other is as defined in SEQ ID NO. 61, and wherein said kappa light chain constant region is as defined in SEQ ID NO. 63 and said lambda light chain constant region is as defined in SEQ ID NO. 64.

46 . A pharmaceutical composition according to claim 45 , wherein said IgG1 heavy chain constant regions as defined in SEQ ID NO. 60 and 61 have their terminal lysines deleted.

47 - 48 . (canceled)

49 . A method for treating a disease comprising administering to a subject in need thereof an effective amount of the pharmaceutical composition according to claim 1 .

50 . The method according to claim 49 , wherein the disease is cancer, optionally wherein the cancer is characterized by expression of B7H4 in cancer cells, optionally wherein the cancer is a solid tumor.

51 . The method according to claim 50 , wherein the cancer is selected from the group consisting of lung cancer, NSCLC (ADC or SQCC), stomach cancer, pancreas cancer, cholangiocarcinoma, bladder cancer, cervical cancer, head and neck cancer, breast cancer, ovarian cancer and uterine cancer.

52 - 54 . (canceled)

55 . A unit dosage form, comprising:

a) an antibody comprising an antigen-binding region capable of binding to human B7H4 and an antigen-binding region capable of binding to human CD3, wherein said antigen-binding regions comprise heavy and light chain variable regions, wherein said heavy and light chain variable regions are humanized and/or human, in an amount of from 5 pg to 1200 mg, and

b) a buffering agent, preferably selected from the group consisting of histidine, glutamate, and mixtures thereof,

wherein the pH of the unit dosage form is from 4.0 to 8.0, preferably 4.5 to 6.5, more preferably 5.0 to 6.0.

56 - 57 . (canceled)

58 . A kit-of-parts comprising:

a. the pharmaceutical composition of claim 1 ,

b. a receptacle for the pharmaceutical composition or for the unit dosage form

c. directions for dilution and/or for use.

59 . A method of preparing a pharmaceutical composition according to claim 1 , comprising the steps of mixing in water for injection:

a. 0.5 to 120 mg/ml of the antibody, and

b. a buffering agent

and adjusting the pH to 4.0-8.0, preferably to 5.0-6.0.