Engineered Artificial Antigen Presenting Cells for Tumor Infiltrating Lymphocyte Expansion

• US 20180127715A1
• Filed: 11/01/2017
• Published: 05/10/2018
• Est. Priority Date: 10/31/2016
• Status: Active Grant

• Current Assignee: Iovance Biotherapeutics, Inc.
• Original Assignee: Iovance Biotherapeutics, Inc.
• Inventors: Veerapathran, Anand, Gokuldass, Aishwarya, Rabinovich, Brian, Lotze, Michael T.
• Granted Patent: US 10,415,015 B2
• Time in Patent Office: Days
• CPC Class Codes: A61K 2039/515 Animal cells
• CPC Class Codes: A61K 2039/5158 Antigen-pulsed cells, e.g. T-cells
• CPC Class Codes: A61K 39/0011 Cancer antigens
• 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 39/4644 Cancer antigens
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 14/70532 B7 molecules, e.g. CD80, CD86
• CPC Class Codes: C07K 14/70575 NGF/TNF-superfamily, e.g. CD70, CD95L, CD153, CD154 NGF C07K14/48, TNF C07K14/525
• CPC Class Codes: C07K 16/4283 against an allotypic or isotypic determinant on Ig
• CPC Class Codes: C07K 2317/622 Single chain antibody (scFv)
• CPC Class Codes: C12N 2501/06 Anti-neoplasic drugs, anti-retroviral drugs, e.g. azacytidine, cyclophosphamide
• CPC Class Codes: C12N 2501/2302 Interleukin-2 (IL-2)
• CPC Class Codes: C12N 2501/25 Tumour necrosing factors [TNF]
• 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 2501/998 Proteins not provided for elsewhere
• CPC Class Codes: C12N 2502/11 blood or immune system cells
• CPC Class Codes: C12N 2502/99 genetically modified cells
• CPC Class Codes: C12N 5/0635 B lymphocytes
• CPC Class Codes: C12N 5/0639 Dendritic cells, e.g. Langherhans cells in the epidermis
• CPC Class Codes: C12N 5/065 : Thymocytes

Abstract

In some embodiments, compositions and methods relating to isolated artificial antigen presenting cells (aAPCs) are disclosed, including aAPCs comprising a myeloid cell transduced with one or more viral vectors, such as a MOLM-14 or a EM-3 myeloid cell, wherein the myeloid cell endogenously expresses HLA-AB/C, ICOS-L, and CD58, and wherein the one or more viral vectors comprise a nucleic acid encoding CD86 and a nucleic acid encoding 4-1BBL and/or OX40L and transduce the myeloid cell to express CD86 and 4-1BBL and/or OX40L proteins. In some embodiments, methods of expanding tumor infiltrating lymphocytes (TILs) with aAPCs and methods of treating cancers using TILs after expansion with aAPCs are also disclosed.

Claims

1. An artificial antigen presenting cell (aAPC) comprising a myeloid cell transduced with one or more viral vectors, wherein the one or more viral vectors comprise a nucleic acid encoding CD86 and one or more nucleic acids encoding a costimulatory molecule, and wherein the myeloid cell expresses a CD86 protein and one or more costimulatory molecules.

2. The aAPC of claim 1, wherein the aAPC can stimulate and expand tumor infiltrating lymphocytes (TILs) contacted with the aAPC.

3. The aAPC of claim 1, wherein the aAPC expands a population of TILs by at least 50-fold over a period of 7 days in a cell culture medium comprising IL-2 at a concentration of about 3000 IU/mL and OKT-3 antibody at a concentration of about 30 ng/mL.

4. The aAPC of claim 1, wherein the aAPC can stimulate and expand a T cell contacted with the aAPC.

5. The aAPC of claim 1, wherein the myeloid cell endogenously expresses HLA-A/B/C, ICOS-L, and CD58.

6. The aAPC of claim 1, wherein the myeloid cell is a MOLM-14 cell.

7. The aAPC of claim 1, wherein the myeloid cell is a EM-3 cell.

8. The aAPC of claim 7, wherein the EM-3 cell is further transduced to express a single chain fragment variable (scFv) binding domain capable of binding the Fc domain of OKT-3 antibody.

9. The aAPC of claim 8, wherein the scFv binding domain comprises clone 7C12 (SEQ ID NO:

27) or clone 8B3 (SEQ ID NO;

28), or a sequence comprising one or more conservative amino acid substitutions thereof.

10. The aAPC of claim 1, wherein the CD86 protein comprises a sequence as set forth in SEQ ID NO:

8, or a sequence comprising one or more conservative amino acid substitutions thereof.

11. The aAPC of claim 1, wherein the nucleic acid encoding CD86 comprises SEQ ID NO:

19.

12. The aAPC of claim 1, wherein the one or more costimulatory molecules comprises a 4-1BBL protein.

13. The aAPC of claim 12, wherein the 4-1BBL protein comprises a sequence as set forth in SEQ ID NO:

9, or a sequence comprising one or more conservative amino acid substitutions thereof.

14. The aAPC of claim 12, wherein the one or more nucleic acids encoding the 4-1BBL protein comprises SEQ ID NO:

16.

15. The aAPC of claim 1, wherein the one or more costimulatory molecules comprises an OX40L protein.

16. The aAPC of claim 15, wherein the OX40L protein comprises a sequence as set forth in SEQ ID NO:

10, or a sequence comprising one or more conservative amino acid substitutions thereof.

17. A method of expanding a population of tumor infiltrating lymphocytes (TILs), the method comprising the step of contacting the population of TILs with a population of aAPCs according to claim 1 in a cell culture medium.

18. The method of claim 17, wherein the cell culture medium further comprises IL-2 at an initial concentration of about 3000 IU/mL and OKT-3 antibody at an initial concentration of about 30 ng/mL.

19. The method of claim 17, wherein the population of APCs expands the population of TILs by at least 50-fold over a period of 7 days in a cell culture medium.

20. The method of claim 17, wherein the myeloid cell endogenously expresses HLA-A/B/C, ICOS-L, and CD58.

21. The method of claim 17, wherein the myeloid cell is a MOLM-14 cell.

22. The method of claim 17, wherein the myeloid cell is a EM-3 cell.

23. The method of claim 22, wherein the EM-3 cell is further transduced to express a single chain fragment variable (scFv) binding domain capable of binding the Fc domain of OKT-3 antibody.

24. The method of claim 23, wherein the scFv binding domain comprises clone 7C12 (SEQ ID NO:

27) or clone 8B3 (SEQ ID NO;

28), or a sequence comprising one or more conservative amino acid substitutions thereof.

25. The method of claim 17, wherein the CD86 protein comprises SEQ ID NO:

8, or a sequence comprising one or more conservative amino acid substitutions thereof.

26. The method of claim 17, wherein the nucleic acid encoding CD86 comprises SEQ ID NO:

19.

27. The method of claim 17, wherein the one or more costimulatory molecules comprises a 4-1BBL protein.

28. The method of claim 27, wherein the 4-1BBL protein comprises a sequence as set forth in SEQ ID NO:

9, or a sequence comprising one or more conservative amino acid substitutions thereof.

29. A method of treating a cancer with a population of tumor infiltrating lymphocytes (TILs) comprising the steps of:

(a) obtaining a first population of TILs from a tumor resected from a patient;

(b) performing an initial expansion of the first population of TILs in a first cell culture medium to obtain a second population of TILs, wherein the second population of TILs is at least 5-fold greater in number than the first population of TILs, and wherein the first cell culture medium comprises IL-2;

(c) performing a rapid expansion of the second population of TILs using a population of myeloid artificial antigen presenting cells (myeloid aAPCs) in a second cell culture medium to obtain a third population of TILs, wherein the third population of TILs is at least 50-fold greater in number than the second population of TILs after 7 days from the start of the rapid expansion; and

wherein the second cell culture medium comprises IL-2 and OKT-3;

(d) administering a therapeutically effective portion of the third population of TILs to a patient with the cancer.

30. An artificial antigen presenting cell (aAPC) comprising a myeloid cell genetically engineered to express CD86 and one or more costimulatory molecules selected from the group consisting of 4-1BBL, OX40L, and a combination thereof.