Bispecific molecules recognizing lymphocyte antigen CD2 and tumor antigens

• US 5,798,229 A
• Filed: 08/02/1994
• Issued: 08/25/1998
• Est. Priority Date: 08/02/1993
• Status: Expired due to Fees

• Current Assignee: Merck Patent GmbH (Merck & Co., Inc.)
• Original Assignee: Merck Patent GmbH (Merck & Co., Inc.)
• Inventors: Wild, Martin, Shraven, Burkhart, Strittmatter, Wolfgang, Meuer, Stefan, Jaggle, Carlota-Silvia
• Primary Examiner(s): LORING, SUSAN
• Application Number:
  US08/284,947
• Time in Patent Office: 1,484 Days
• Field of Search: 530/387.3, 530/388.2, 530/388.22, 530/388.75, 530/388.8, 530/388.85, 424/136.1, 435/70.21
• US Class Current: 435/70.21
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 16/2806 against CD2
• CPC Class Codes: C07K 16/30 from tumour cells
• CPC Class Codes: C07K 2317/74 Inducing cell proliferation

Abstract

The present invention relates to novel and highly effective bispecific antibody fragments which recognize lymphocyte CD2 antigen and any variable tumor antigen. Moreover, the invention relates to two new monoclonal antibodies terms AICD2.M1 and AICD2.M2. A combination of these antibodies, whereby at least one of them is a bispecific antibody fragment, can be used successfully in tumor therapy and diagnostics.

Claims

1. A molecule which is a bispecific antibody fragment, comprising

(a) a first binding site specific for an epitope on a tumor cell antigen and

(b) a second binding site specific for an epitope of antigen CD2, selected from the group consisting of AIDC2.M1 and AICD2.M2,

of the formulae BAb<

X,AICD2.M1>

Y or BAb<

X,AICD2.M2>

Y, wherein

BAb≦

indicates the molecule is a bispecific antibody,

X is a first antibody determinant specific for an epitope on a tumor cell antigen,

AICD2.M1 and AICD2.M2 are antibody determinants of antibodies AICD2.M1 and AICD2.M2, respectively, which are specific for antigen CD2, and

≧

Y indicates the molecule is an antibody fragment.

2. A bispecific antibody fragment of claim 1, wherein X is an antibody determinant derived from monoclonal antibodies MAb 425 (ATCC HB 9629) or MAb 361 (ATCC HB 9325).

3. A bispecific antibody fragment of claim 1, wherein Y indicates the molecule is an F(ab'"'"')₂ molecule.

4. A bispecific antibody fragment of claim 3, selected from

BAb<

425,AICD2.M1>

F(ab'"'"')₂,

BAb<

425,AICD2.M2>

F(ab'"'"')₂,

Bab<

361,AICD2.M1>

F(ab'"'"')₂, or

BAb<

361 ,AICD2.M2>

F(ab'"'"')₂.

5. A bispecific antibody fragment of claim 1, wherein Y indicates the molecule is an Fv molecule.

6. A bispecific antibody fragment of claim 1, wherein said bispecific antibody fragment is cytotoxic for a tumor cell carrying said tumor cell epitope.

7. A bispecific antibody fragment of claim 1, wherein said bispecific antibody fragment is capable of triggering T-cell or NK-cell mediated cell lysis of a tumor cell carrying said tumor cell epitope.

8. A bispecific antibody fragment of claim 1, wherein said bispecific antibody fragment does not compete with LFA3 (CD58) for binding to CD2.

9. A bispecific antibody fragment of claim 1, wherein said bispecific antibody fragment demonstrates strong avidity for said tumor cell, strong avidity for T-cells and NK-cells, no receptor modulation, and/or high NK- and T-cell specificity, whereby said bispecific antibody fragment alone is ineffective or marginally effective to activate T-cell or NK-cell mediated cell lysis of a tumor cell carrying said tumor cell epitope, but is therapeutically effective to activate T-cell or NK-cell mediated cell lysis of a tumor cell carrying said tumor cell epitope only via a two-step activation with a second, different antibody, antibody fragment, bispecific antibody or bispecific antibody fragment having a binding site specific for an epitope of antigen CD2.

10. A method for preparing a bispecific antibody fragment of claim 1, comprising enzymatically converting each of two different monoclonal antibodies (a'"'"') and (b'"'"'),

wherein

(a'"'"') has a first binding site specific for an epitope on a tumor cell antigen, and

(b'"'"') has a second binding site specific for an epitope of leukocyte antigen CD2, and

wherein each antibody comprises two identical light chain and heavy chain half molecules linked by one or more disulfide bonds, into two F(ab'"'"')₂ molecules,

splitting each F(ab'"'"')₂ molecule under reducing conditions into Fab'"'"' thiols,

derivatizing the Fab'"'"' molecules derived from either antibody (a'"'"') or (b'"'"') with a thiol activating agent, and

combining the thus-derivatized Fab'"'"' molecule with the corresponding other Fab'"'"' molecule derived from antibody (b'"'"') or (a'"'"') to produce a bispecific antibody F(ab'"'"')₂ fragment bearing both tumor specificity and leukocyte specificity,

wherein said monoclonal antibody (b'"'"') specific for an epitope of leukocyte antigen CD2 is selected from the group consisting of AICD2.M1 and AICD2.M2.

11. A method of claim 10, wherein the monoclonal antibody (a'"'"') is MAb 425 (ATCC HB 9629), MAb 361 (ATCC HB 9325).

14. A pharmaceutical formulation comprising a bispecific antibody fragment of claim 1, and a pharmaceutically acceptable excipient.

15. A kit comprising

a first pharmaceutical formulation (I) of claim 14, wherein the bispecific antibody fragment is BAb<

X,AICD2.M2>

Y and

a second pharmaceutical formulation (II) comprising

MAb,AICD2.M1, or MAb<

AICD2.M1>

Y, or BAb<

X,AICD2.M1>

Y,

wherein X and Y have the indicated meanings,

together with a pharmaceutically acceptable excipient.

16. A kit comprising

a first pharmaceutical formulation (I) of claim 14, wherein the bispecific antibody fragment is BAb<

X,AICD2.M1>

Y and

a second pharmaceutical formulation (II) comprising

MAb,AICD2.M2, or MAb<

AICD2.M2>

Y, or BAb<

X.AICD2.M2>

Y,

wherein X and Y have the indicated meanings,

together with a pharmaceutically acceptable excipient.

17. A kit of claim 16, wherein

formulation (1) comprises a bispecific antibody fragment selected from

Bab<

425,AICD2.M1>

F(ab'"'"')₂ or

BAb<

361,AICD2.M1>

F(ab'"'"')₂,

and

formulation (II) comprises MAb<

AICD2.M2>

F(ab'"'"')₂.

12. Monoclonal antibody AICD2.M1, which is specific for an epitope of antigen CD2, and is obtainable by isolation from hybridoma cell line 1 H 10 (DSM ACC2118).

13. Monoclonal antibody AICD2.M2, which is specific for an epitope of antigen CD2, and is obtainable by isolation from hybridoma cell line 7 D 3 (DSM ACC2119).