Method of producing lymphocytes

• US 8,765,469 B2
• Filed: 08/10/2006
• Issued: 07/01/2014
• Est. Priority Date: 08/17/2005
• Status: Expired due to Fees

• Current Assignee: Takara Bio Incorporated (Ningbo Junsheng Investment Group Co., Ltd.)
• Original Assignee: Takara Bio Incorporated (Ningbo Junsheng Investment Group Co., Ltd.)
• Inventors: Marui, Takahiro, Nagamine, Kinuko, Muraki, Nobuko, Kato, Akiko, Enoki, Tatsuji, Sagawa, Hiroaki, Kato, Ikunoshin
• Primary Examiner(s): LI, QIAN JANICE
• Application Number:
  US11/990,443
• Publication Number:
  US 20100150886A1
• Time in Patent Office: 2,882 Days
• Field of Search: 435/325, 435/375
• US Class Current: 435/375
• CPC Class Codes: A61K 2039/5154 Antigen presenting cells [APCs], e.g. dendritic cells or macrophages
• CPC Class Codes: A61K 35/17 Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes when activated by a specific antigen A61K39/00
• CPC Class Codes: A61K 35/26 Lymph; Lymph nodes; Thymus; Spleen; Splenocytes; Thymocytes
• CPC Class Codes: A61K 39/4611 T-cells, e.g. tumor infiltrating lymphocytes [TIL], lymphokine-activated killer cells [LAK] or regulatory T cells [Treg]
• CPC Class Codes: A61K 39/464411 Immunoglobulin superfamily
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: C07K 14/78 Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
• CPC Class Codes: C12N 2501/51 B7 molecules, e.g. CD80, CD86, CD28 (ligand), CD152 (ligand)
• CPC Class Codes: C12N 2501/515 CD3, T-cell receptor complex
• CPC Class Codes: C12N 2501/58 Adhesion molecules, e.g. ICAM, VCAM, CD18 (ligand), CD11 (ligand), CD49 (ligand)
• CPC Class Codes: C12N 2531/00 Microcarriers
• CPC Class Codes: C12N 2533/52 Fibronectin; Laminin
• CPC Class Codes: C12N 5/0636 T lymphocytes
• CPC Class Codes: G01N 33/547 with antigen or antibody attached to the carrier via a bridging agent

Abstract

A method for preparing lymphocytes characterized in that the method comprises the step of carrying out expansion in the presence of (a) fibronectin, a fragment thereof or a mixture thereof, (b) a CD3 ligand, and (c) a CD28 ligand.

Claims

1. A method for preparing lymphocytes suitable for adoptive immunotherapy comprising the step of carrying out expansion in the simultaneous presence of (a) fibronectin, a fragment thereof or a mixture thereof, (b) an immobilized CD3 ligand, and (c) an immobilized CD28 ligand.

2. The method according to claim 1, wherein the CD3 ligand is an anti-CD3 antibody.

3. The method according to claim 1, wherein the CD28 ligand is an anti-CD28 antibody.

4. The method according to claim 1, wherein the fibronectin fragment is a polypeptide (m) comprising at least any one of the amino acid sequences shown in SEQ ID NOs:

1 to 8 of Sequence Listing, or a polypeptide (n) comprising at least one amino acid sequence having substitution, deletion, insertion or addition of one or plural number of amino acids in any one of said amino acid sequences, wherein the polypeptide (n) has a function equivalent to said polypeptide (m).

5. The method according to claim 4, wherein the fibronectin fragment has a cell adhesion activity and/or a heparin binding activity.

6. The method according to claim 5, wherein the fibronectin fragment is at least one polypeptide selected from the group consisting of polypeptides having any one of the amino acid sequences shown in SEQ ID NOs:

9 to 21 of Sequence Listing.

7. The method according to claim 1, wherein the lymphocytes are lymphokine-activated cells.

8. The method according to claim 1, further comprising the step of transducing a foreign gene into lymphocytes.

9. The method according to claim 8, wherein the foreign gene is transduced using retrovirus, adenovirus, adeno-associated virus, lentivirus or simian virus.

10. A method for preparing a solid phase comprising:

contacting the solid phase with (a) fibronectin, a fragment thereof or a mixture thereof, (b) a CD3 ligand and (c) a CD28 ligand under an acidic condition.

11. The method according to claim 10, wherein the solid phase is a cell culture carrier.

12. A solid phase to which (a) fibronectin, a fragment thereof or a mixture thereof, (b) a CD3 ligand, and (c) a CD28 ligand are immobilized.

13. The solid phase according to claim 12, wherein the solid phase is a cell culture plate, a petri dish, a flask, a bag, beads, a membrane or a slide glass.