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 method for treating a subject with melanoma, the method comprising administering expanded tumor infiltrating lymphocytes (TILs) comprising:
(a) adding a tumor digest or tumor fragments into a closed system, wherein the tumor digest or tumor fragments comprise a first population of TILs and are obtained from a tumor that was resected from the subject;
(b) 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 a first period of about 3 to 11 days to obtain the second population of TILs, and wherein the transition from step (a) to step (b) occurs without opening the system;
(c) performing a second expansion by supplementing the cell culture medium 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 a second period of about 7 to 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 (b) to step (c) occurs without opening the system;
(d) harvesting the third population of TILs obtained from step (c), wherein the transition from step (c) to step (d) occurs without opening the system;
(e) transferring the harvested third TIL population from step (d) to an infusion bag, wherein the transfer from step (d) to (e) occurs without opening the system;
(f) cryopreserving the infusion bag comprising the harvested TIL population from step (e) using a cryopreservation process, wherein no selection of the first population of TILs, second population of TILs, third population of TILs, harvested TIL population, and/or the therapeutic TIL population based on CD8 expression is performed during any of steps (a) to step (0, and
(g) administering a therapeutically effective dosage of the harvested TIL population from the infusion bag in step (f) to the subject.
2. The method according to claim 1 , wherein the tumor digest in step (a) was prepared by incubating a sample of the tumor that was resected from the subject in an enzymatic media.
3. The method according to claim 2 , wherein the preparation of the tumor digest comprises disrupting the tumor sample mechanically so as to dissociate the tumor sample, optionally further comprising purifying the disassociated tumor sample using a density gradient separation.
4. The method according to claim 2 , wherein the enzymatic media comprises DNase.
5. The method according to claim 4 , wherein the enzymatic media comprises 30 units/mL of DNase.
6. The method according to claim 2 , wherein the enzymatic media comprises collagenase.
7. The method according to claim 6 , wherein the enzymatic media comprises 1.0 mg/mL of collagenase.
8. The method according to claim 1 , wherein the third population of TILs harvested in step (d) comprises sufficient TILs for administering a therapeutically effective dosage of the TILs in step (g).
9. The method according to claim 8 , wherein the therapeutically effective dosage in step (g) comprises from about 1×10 9 to 5×10 10 TILs.
10. The method according to claim 8 , wherein the therapeutically effective dosage in step (g) comprises from about 1×10 9 to about 9×10 10 TILs.
11. The method according to claim 8 , wherein the therapeutically effective dosage in step (g) comprises from about 5×10 9 to 1×10 10 TILs.
12. The method according to claim 8 , wherein the therapeutically effective dosage in step (g) comprises from about 1×10 10 to 5×10 10 TILs.
13. The method according to claim 1 , wherein the APCs comprise peripheral blood mononuclear cells (PBMCs).
14. The method according to claim 13 , wherein the PBMCs are supplemented at a ratio of about 1:25 TIL:PBMCs.
15. The method according to claim 1 , wherein prior to administering a therapeutically effective dosage of TIL cells in step (g), a non-myeloablative lymphodepletion regimen has been administered to the subject.
16. The method according to claim 15 , where the non-myeloablative lymphodepletion regimen comprises the steps of administration of cyclophosphamide at a dose of 60 mg/m 2 /day for two days followed by administration of fludarabine at a dose of 25 mg/m 2 /day for five days.
17. The method according to claim 1 , further comprising the step of treating the subject with a high-dose IL-2 regimen starting on the day after administration of the TIL cells to the subject in step (g).
18. The method according to claim 17 , wherein the high-dose IL-2 regimen comprises 600,000 or 720,000 IU/kg administered as a 15-minute bolus intravenous infusion every eight hours until tolerance.
19. The method according to claim 1 , wherein the melanoma is metastatic melanoma.
20. The method according to claim 1 , wherein the first expansion in step (b) and the second expansion in step (c) are each individually performed within a period of 11 days.
21. The method according to claim 1 , wherein the first expansion is performed for about 10 days or for about 11 days.
22. The method according to claim 1 , wherein the second expansion is performed for about 10 days or for about 11 days.
23. The method according to claim 1 , wherein steps (a) through (e) are performed in about 10 days to about 22 days.
24. The method according to claim 1 , wherein steps (a) through (e) are performed in about 20 days or less.
25. The method according to claim 1 , wherein no selection of the first population of TILs, second population of TILs, third population of TILs, harvested TIL population, and/or the therapeutic TIL population based on CD4 and/or CD8 expression is performed during any of steps (a) to step (f).
26. The method according to claim 1 , wherein no selection of the first population of TILs, second population of TILs, third population of TILs, harvested TIL population, and/or the therapeutic TIL population is performed during any of steps (a) to step (f).
27. The method according to claim 1 , wherein the therapeutic population of TILs is capable of exhibiting IFN-gamma (IFN-γ) secretion greater than 1000 pg/ml.