Anti-rabbit CD19 antibodies and methods of use

Patent metadata

Field	Value
Publication number	US-12378315-B2
Application number	US-202117362126-A
Country	US
Kind code	B2
Filing date	Jun 29, 2021
Priority date	Dec 30, 2018
Publication date	Aug 5, 2025
Grant date	Aug 5, 2025

Abstract

Herein is reported an antibody binding to rabbit CD19 comprising (a) a HVR-H1 comprising the amino acid sequence of SEQ ID NO: 32 or 33 or 34, (b) a HVR-H2 comprising the amino acid sequence of SEQ ID NO: 35 or 36, (c) a HVR-H3 comprising the amino acid sequence of SEQ ID NO: 37, (d) a HVR-L1 comprising the amino acid sequence of SEQ ID NO: 38, (e) a HVR-L2 comprising the amino acid sequence of SEQ ID NO: 39, and (f) a HVR-L3 comprising the amino acid sequence of SEQ ID NO: 40, as well as methods of using the same, especially in the identification and selection of antibody producing rabbit B-cells.

First claim

The invention claimed is:

1. An antibody binding to rabbit CD19 comprising
(a) a HVR-H1 comprising the amino acid sequence of SEQ ID NO: 32 or 33 or 34,
(b) a HVR-H2 comprising the amino acid sequence of SEQ ID NO: 35 or 36,
(c) a HVR-H3 comprising the amino acid sequence of SEQ ID NO: 37,
(d) a HVR-L1 comprising the amino acid sequence of SEQ ID NO: 38,
(e) a HVR-L2 comprising the amino acid sequence of SEQ ID NO: 39, and
(f) a HVR-L3 comprising the amino acid sequence of SEQ ID NO: 40.

2. The antibody according to claim 1 , wherein the antibody is a monoclonal antibody.

3. The antibody according to claim 1 , wherein the antibody is a chimeric or humanized antibody.

4. The antibody according to claim 1 , wherein the antibody comprises a variable heavy chain domain of SEQ ID NO: 30 and a variable light chain domain of SEQ ID NO: 26.

5. The antibody according to claim 1 , wherein the antibody is a full length antibody or an antibody fragment.

6. The antibody according to claim 1 , wherein the antibody is conjugated to a detectable label.

7. The antibody according to claim 6 , wherein the detectable label is a fluorescent dye.

8. A method for selecting a rabbit B-cell comprising the following steps:
a) incubating a multitude of rabbit B-cells with an antibody according to claim 1
b) selecting one or more B-cells to which the antibody according to claim 1 is bound and thereby selecting a rabbit B-cell.

9. The method according to claim 8 further comprising one or more of the following steps:
after step b) and prior to step c): incubating the rabbit B-cells at 37° C. for one hour in co-cultivation medium, and/or
c) single depositing one or more rabbit B-cells to which the antibody according to claim 1 is bound, and/or
d) co-cultivating the single deposited rabbit B-cells with feeder cells in a co-cultivation medium, and/or
e) selecting a rabbit B-cell proliferating in step d) and thereby selecting a rabbit B-cell.

10. A method for removing non B-cells for a cultivation comprising the following steps:
a) co-cultivating rabbit B-cells, which have been deposited either as single cells or as pool of cells, with feeder cells,
b) incubating the cells from the co-cultivation obtained in step a) with the antibody according to claim 1 , and
c) selecting one or more rabbit B-cells to which the antibody according to claim 1 is bound and thereby removing non-B-cells.

11. A method for determining the number B-cells in a co-cultivation of a single deposited B-cell with feeder cells comprising the following steps:
a) co-cultivating a single deposited rabbit B-cell with feeder cells,
b) incubating the cells from the co-cultivation obtained in step a) with the antibody according to claim 1 , and
c) determining the number of B-cells in the cultivation by counting the number of cells to which the antibody according to claim 1 is bound.

12. A method for co-cultivating one or more rabbit B-cells comprising the steps of
incubating a multitude of rabbit B-cells/labelling individual B-cells of a multitude of rabbit B-cells with an antibody according to claim 1 ,
selecting/depositing one or more rabbit B-cells that have the antibody according to any one of claims 1 to 7 bound to their surface/that have been labelled either as individual B-cells (single deposited B-cell) or as a pool of B-cells, and
co-cultivating the single deposited rabbit B-cells or the pool of rabbit B-cells with feeder cells.

13. The method according to claim 9 , wherein the co-cultivating is in the presence of a synthetic feeder mix that comprises IL-1β, TNFα, IL-10, and one or more selected from IL-21, SAC, BAFF, IL-2, IL-4, and IL-6.

14. A method for selecting B-cells comprising the following steps:
a) obtaining B-cells from the blood of a rabbit,
b) incubating the B-cells with an antibody according to claim 1 , which is bound to a bead,
c) removing non-bound B-cells,
d) recovering the bound B-cells from the beads and thereby selecting one or more B-cells.

15. The method according to claim 14 further comprising the following steps
e) optionally incubating the recovered B-cells at 37° C. for one hour in co-cultivation medium,
f) depositing one or more recovered B-cells in an individual container,
g) co-cultivating the deposited cells with a feeder cell in a co-cultivation medium,
h) selecting a B-cell proliferating in step g) and thereby selecting a B-cell.

Assignees

• Hoffmann La Roche

Inventors

• Haas Alexander
• Jung Friederike
• Klostermann Stefan
• Offner Sonja

Classifications

• C12N2502/30

  tumour cells · CPC title

• C12N2500/80

  from animals · CPC title

• C12N5/0635

  B lymphocytes · CPC title

• C07K2317/24

  containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered · CPC title

• C07K2317/565

  Complementarity determining region [CDR] · CPC title