Engineered artificial antigen presenting cells for tumor infiltrating lymphocyte expansion
In some embodiments, compositions and methods re¬lating to isolated artificial antigen presenting cells (aAPCs) are dis¬closed, 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-A/B/C, ICOS-L, and CD58, and wherein the one or more viral vectors com¬prise 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.
1 . A method of expanding a population of lymphocytes, the method comprising:
(i) transducing a population of myeloid cells with one or more viral vectors comprising (a) a nucleic acid encoding CD86, (b) one or more nucleic acids encoding one or more costimulatory molecules, and (c) a nucleic acid encoding an anti-OKT-3 antibody scFv binding domain, to obtain a population of artificial antigen presenting cells (aAPCs) expressing a protein encoded by each of the nucleic acids of (a), (b), and (c); and
(ii) contacting the population of lymphocytes with the population of aAPCs in a cell culture medium for a period of time to obtain an expanded population of lymphocytes.
2 . The method of claim 1 , wherein the population of lymphocytes comprises tumor-infiltrating lymphocytes (TILs).
3 . The method of claim 1 , wherein the cell culture medium comprises IL-2.
4 . The method of claim 3 , wherein the IL-2 is at an initial concentration of about 3000 IU/mL.
5 . The method of claim 1 , wherein the cell culture medium comprises OKT-3 antibody.
6 . The method of claim 5 , wherein the OKT-3 antibody is at an initial concentration of about 30 ng/mL.
7 . The method of claim 1 , wherein the population of lymphocytes is expanded by at least 50-fold over a period of about 7 days.
8 . The method of claim 1 , wherein the population of aAPCs endogenously expresses HLA-A/B/C, ICOS-L, and CD58.
9 . The method of claim 1 , wherein the anti-OKT-3 antibody scFv binding domain comprises the peptide sequence of SEQ ID NO:27 or SEQ ID NO:28.
10 . The method of claim 1 , wherein the one or more costimulatory molecules are independently selected from the group consisting of 4-1BB (CD137), OX40 (CD134), CD1a, CD1b, CD1c, CD1d, CD2, CD3γ, CD3δ, CD3ε, CD4, CD5, CD6, CD7, CD8α, CD8β, CD9, CD10, CD11a, CD11b, CD11c, CDw12, CD13, CD14, CD15, CD15s, CD16a, CD16b, CD18, CD19, CD20, CD21, CD22, CD23, CD24, CD25, CD26, CD27, CD28, CD29, CD30, CD31, CD32, CD33, CD34, CD35, CD36, CD37, CD38, CD39, CD40, CD41, CD42a, CD42b, CD42c, CD42d, CD43, CD44, CD45, CD45R, CD46, CD47, CD48, CD49a, CD49b, CD49c, CD49d, CD49e, CD49f, CD50, CD51, CD52, CD53, CD54, CD55, CD56, CD57, CD58, CD59, CDw60, CD61, CD62E, CD62L, CD62P, CD63, CD64, CD65, CD66a, CD66b, CD66c, CD66d, CD66e, CD66f, CD67, CD68, CD69, CDw70, CD71, CD72, CD73, CD74, CDw75, CDw76, CD77, CD79a, CD79B, CD80, CD81, CD82, CD83, CD84, CD85, CD86, CD87, CD88, CD89, CD90, CD91, CDw92, CD93, CD94, CD95, CD96, CD97, CD98, CD99, CD100, CD101, CD102, CD103, CD104, CD105, CD106, CD107a, CD107b, CDw108, CDw109, CD114, CD115, CD116, CD117, CD118, CD119, CD120a, CD120b, CD121a, CD121b, CD122, CD123, CDw124, CD125, CD126, CDw127, CDw128a, CDw128b, CDw130, CDw131, CD132, CD133, CD135, CD136, CD138, CD139, CD140a, CD140b, CD141, CD142, CD143, CD144, CDw145, CD146, CD147, CD148, CDw149, CD150, CD151, CD152, CD153, CD154, CD155, CD156, CD157, CD158a, CD158b, CD161, CD162, CD163, CD164, CD165, CD166, and TCRζ.
11 . The method of claim 10 , wherein the one or more costimulatory molecules are independently selected from the group consisting of CD28, 4-1BB (CD137), and OX40 (CD134).
12 . The method of claim 1 , wherein the expansion is performed using a gas permeable container.
13 . The method of claim 1 , wherein the ratio of the population of lymphocytes to the population of APCs is between about 1 to 200 and about 1 to 400.
14 . The method of claim 13 , wherein the ratio of the population of lymphocytes to the population of APCs is about 1 to 300.
15 . The method of claim 1 , wherein the one or more viral vectors comprise a lentiviral vector.
16 . The method of claim 1 , wherein the expanded population of lymphocytes is cryopreserved.
17 . The method of claim 2 , wherein the expanded population of TILs is cryopreserved.
18 . The method of claim 1 , wherein the population of myeloid cells comprises a MOLM-14 cell, an EM-3 cell, a K562 cell, a MOLM-13 cell, a KG1-246 cell, aKG1-8031 cell, or an EM-2 cell.
19 . The method of claim 1 , wherein the CD86 protein comprises a sequence selected from the group consisting of SEQ ID NO:8 and SEQ ID NO:19, or a sequence comprising one or more conservative amino acid substitutions thereof.
20 . The method of claim 1 , wherein the one or more costimulatory molecules comprises 4-1BBL protein.
21 . The method of claim 20 , wherein the 4-1BBL protein comprises a sequence as set forth in SEQ ID NO:9 or SEQ ID NO:16, or a sequence comprising one or more conservative amino acid substitutions thereof.
22 . The method of claim 1 , wherein the one or more costimulatory molecules comprises OX40L protein.
23 . The method of claim 21 , 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.