Recombinant production of mixtures of antibodies
Provided is methods for producing mixtures of antibodies from a single host cell clone, wherein, a nucleic acid sequence encoding a light chain and nucleic acid sequences encoding different heavy chains are expressed in a recombinant host cell. The recombinantly produced antibodies in the mixtures according to the invention suitably comprise identical light chains paired to different heavy chains capable of pairing to the light chain, thereby forming functional antigen-binding domains. Mixtures of the recombinantly produced antibodies are also provided by the invention. Such mixtures can be used in a variety of fields.
1. A method of producing a mixture of two or three non-identical antibodies in a recombinant host cell clone, the method comprising:
expressing in the recombinant host cell clone a polynucleotide or polynucleotides encoding at least one light chain and two different heavy chains that are able to pair with the at least one light chain, wherein the at least one light chain and two different heavy chains are preselected from a display library comprising said one light chain, wherein the two or three non-identical antibodies specifically bind EP-CAM and/or CD46 and the at least one light chain comprises a sequence selected from SEQ ID NO: 2 and SEQ ID NO: 4, and the two different heavy chains comprise a sequence presented as SEQ ID NO: 6 and 8;
wherein the two or three non-identical antibodies specifically bind EP-CAM and/or CD46 and the at least one light chain comprises a sequence selected from SEQ ID NO: 2 and SEQ ID NO: 4, and the two different heavy chains comprise the sequence of SEQ ID NO: 6 and 10;
wherein the two or three non-identical antibodies specifically bind CD46 and the at least one light chain comprises a sequence selected from SEQ ID NO: 2 and SEQ ID NO: 4, and the two different heavy chains comprise the sequence of SEQ ID NO: 8 and 10;
wherein the two or three non-identical antibodies specifically bind CD22, and/or CD72 and the at least one light chain comprises a sequence presented as SEQ ID NO: 18, and the two different heavy chains comprise the sequence of SEQ ID NO: 12 and 14;
wherein the two or three non-identical antibodies specifically bind CD22, and/or HLA-DR and the at least one light chain comprises a sequence presented as SEQ ID NO: 18, and the two different heavy chains comprise the sequence of SEQ ID NO: 12 and 16; or
wherein the two or three non-identical antibodies specifically bind CD72, and/or HLA-DR and the at least one light chain comprises a sequence presented as SEQ ID NO: 18, and the two different heavy chains comprise the sequence of SEQ ID NO: 14 and 16.
2. The method according to claim 1 , wherein the recombinant host cell clone comprises a polynucleotide encoding a common light chain that is able to pair with the two different heavy chains, such that the produced antibodies comprise common light chains.
3. The method according to claim 1 , wherein the mixture of two or three non-identical antibodies comprises a bispecific antibody and at least one monospecific antibody.
4. The method according to claim 1 , wherein the mixture comprises a bispecific antibody and two different monospecific antibodies.
5. The method according to claim 1 , further comprising: recovering antibodies separately or as a mixture from the host cell clone or a culture comprising the host cell clone.
6. The method according to claim 5 , wherein the antibodies are recovered separately from the host cell clone or a culture comprising the host cell clone.
7. The method according to claim 5 , wherein the antibodies are recovered as a mixture from the host cell clone or a culture comprising the host cell clone.
8. The method according to claim 1 , wherein at least two antibodies comprising a heavy light chain dimer in the mixture of antibodies have different specificities and affinities for their respective antigens.
9. The method according to claim 1 , wherein the method does not require amplification of said polynucleotide or polynucleotides encoding at least one light chain and two different heavy chains in the cell.
10. The method according to claim 1 , wherein the mixture comprises three non-identical antibodies.
11. The method according to claim 2 , wherein the common light chain is identical in each light chain/heavy chain pair of the two or three non-identical antibodies.
12. The method according to claim 2 , wherein the common light chain is the only light chain present in the mixture.
13. The method according to claim 1 , wherein the two different heavy chains differ in their variable region.
14. The method according to claim 1 , wherein the two different heavy chains differ in both the variable region and constant region.
15. The method according to claim 2 , wherein:
the mixture comprises three non-identical antibodies;
the common light chain is identical in each light chain/heavy chain pair of the three non-identical antibodies;
the common light chain is the only light chain present in the mixture; and
the two different heavy chains differ in their variable region.
16. The method of claim 1 , wherein said two different heavy chains differ in their constant regions sufficiently so that the amount of bispecific antibodies is decreased as compared to the theoretical percentage of bispecific antibodies.
17. The method of claim 8 , wherein the different specificities are directed to different epitopes on the same antigen.
18. The method of claim 8 , wherein the different specificities are directed to different antigens present in one antigen comprising mixture.
19. The method of claim 8 , wherein at least two of the antibodies have different affinities for the same epitope.
20. The method of claim 1 , wherein the two different heavy chains are of IgG isotype.