Processes for production of tumor infiltrating lymphocytes and uses of same in immunotherapy
The present invention provides improved and/or shortened methods for expanding TILs and producing therapeutic populations of TILs, including novel methods for expanding TIL populations in a closed system that lead to improved efficacy, improved phenotype, and increased metabolic health of the TILs in a shorter time period, while allowing for reduced microbial contamination as well as decreased costs. Such TILs find use in therapeutic treatment regimens.
1. A cryopreserved tumor infiltrating lymphocyte (TIL) composition comprising a therapeutic population of infiltrating lymphocytes (TILs), wherein the cryopreserved TIL composition is produced by a method comprising:
(a) obtaining a first population of TILs from a tumor resected from a subject by processing a tumor sample obtained from the subject into a tumor digest;
(b) adding the tumor digest into a closed system;
(c) performing a first expansion by culturing the first population of TILs in a cell culture medium comprising IL-2 to produce a second population of TILs, wherein the first expansion is performed in a closed container providing a first gas-permeable surface area, wherein the first expansion is performed for about 3-11 days to obtain the second population of TILs, and wherein the transition from step (b) to step (c) occurs without opening the system;
(d) performing a second expansion by supplementing the cell culture medium of the second population of TILs with additional IL-2, OKT-3, and antigen presenting cells (APCs), to produce a third population of TILs, wherein the second expansion is performed for about 7-11 days to obtain the third population of TILs, wherein the second expansion is performed in a closed container providing a second gas-permeable surface area, and wherein the transition from step (c) to step (d) occurs without opening the system;
(e) harvesting the therapeutic population of TILs obtained from step (d), wherein the transition from step (d) to step (e) occurs without opening the system;
(f) transferring the harvested TIL population from step (e) to an infusion bag, wherein the transfer from step (e) to (f) occurs without opening the system; and
(g) cryopreserving the infusion bag comprising the harvested TIL population from step (f) using a cryopreservation process to obtain the cryopreserved TIL composition.
2. The cryopreserved TIL composition according to claim 1 , wherein processing a tumor sample obtained from the subject into a tumor digest in step (a) comprises incubating the tumor sample in an enzymatic media.
3. The cryopreserved TIL composition according to claim 2 , wherein processing a tumor sample obtained from the subject into a tumor digest in step (a) further comprises disrupting the tumor sample mechanically so as to dissociate the tumor sample.
4. The cryopreserved TIL composition according to claim 3 , wherein processing a tumor sample obtained from the subject into a tumor digest in step (a) further comprises purifying the disassociated tumor sample using a density gradient separation.
5. The cryopreserved TIL composition according to claim 2 , wherein the enzymatic media comprises DNase.
6. The cryopreserved TIL composition according to claim 5 , wherein the enzymatic media comprises 30 units/mL of DNase.
7. The cryopreserved TIL composition according to claim 2 , wherein the enzymatic media comprises collagenase.
8. The cryopreserved TIL composition according to claim 7 , wherein the enzymatic media comprises 1.0 mg/mL of collagenase.
9. The cryopreserved TIL composition according to claim 1 , wherein the cell culture medium is CTS Optimizer.
10. The cryopreserved TIL composition according to claim 1 , wherein the medium in the first expansion and/or the second expansion is free of human serum.
11. The cryopreserved TIL composition according to claim 1 , wherein the therapeutic population of TILs harvested in step (e) comprises sufficient TILs for use in administering a therapeutically effective dosage to a subject.
12. The cryopreserved TIL composition according to claim 11 , wherein the therapeutically effective dosage comprises from about 1×10 9 to about 9×10 10 TILs.
13. The cryopreserved TIL composition according to claim 1 , wherein the APCs comprise peripheral blood mononuclear cells (PBMCs).
14. The cryopreserved TIL composition according to claim 1 , wherein the therapeutic population of TILs harvested in step (e) exhibits an increased subpopulation of CD8+ cells relative to the first and/or second population of TILs.
15. The cryopreserved TIL composition according to claim 1 , wherein the PBMCs are supplemented at a ratio of about 1:25 TIL:PBMCs.
16. The cryopreserved TIL composition according to claim 1 , wherein the first expansion in step (c) and the second expansion in step (d) are each individually performed within a period of 11 days.
17. The cryopreserved TIL composition according to claim 1 , wherein steps (a) through (f) are performed in about 10 days to about 22 days.
18. The cryopreserved TIL composition according to claim 1 , wherein steps (a) through (f) are performed in about 15 days to about 22 days.
19. The cryopreserved TIL composition according to claim 1 , wherein steps (a) through (f) are performed in about 20 days to about 22 days.
20. The cryopreserved TIL composition of claim 1 , wherein the composition comprises DMSO.
21. The cryopreserved TIL composition of claim 1 , wherein the composition comprises 7% to 10% DMSO.
22. The cryopreserved TIL composition of claim 1 , wherein the composition comprises cryopreservation medium.
23. The cryopreserved TIL composition of claim 22 , wherein the cryopreservation medium is CS10.
24. The cryopreserved TIL composition of claim 1 , wherein the second population of TILs is at least 50-fold greater in number than the first population of TILs.