# US 11,998,568 B2 · Processes for production of tumor infiltrating lymphocytes and uses of same in immunotherapy

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- Number: US 11,998,568 B2
- Document ID: US11998568B2
- Status: Granted
- Kind: B2
- Application number: 17/817,232
- Filed: 2022-08-03
- Published: 2022-12-08
- Granted: 2024-06-04
- Earliest claimed: 2017-03-29
- Assignees: Iovance Biotherapeutics, Inc.
- Applicants: Iovance Biotherapeutics, Inc.
- Inventors: Seth Wardell; James Bender; Michael T. Lotze
- Web page: https://patentgrep.com/patent/US11998568B2

## CPC

- 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)
- A01N 1/162: Temperature processes, e.g. following predefined temperature changes over time
- A61K 9/0019: {Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner (non-active ingredients are additionally classified in A61K47/00)}
- A61K 31/675: having nitrogen as a ring hetero atom, e.g. pyridoxal phosphate
- A61K 31/7076: containing purines, e.g. adenosine, adenylic acid
- A61K 38/2013: {IL-2}
- A61K 40/11: T-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer [LAK] cells
- A61K 40/428: {Undefined tumor antigens, e.g. tumor lysate or antigens targeted by cells isolated from tumor}
- A61P 35/00: Antineoplastic agents
- C12N 5/0634: {Cells from the blood or the immune system}
- C12N 5/0636: {T lymphocytes}
- C12N 5/0638: {Cytotoxic T lymphocytes [CTL] or lymphokine activated killer cells [LAK]}
- A61K 38/217: {IFN-gamma}
- A61K 39/0011: {Cancer antigens}
- A61K 40/50: characterised by the use of allogeneic cells
- A61K 2039/5154: {Antigen presenting cells [APCs], e.g. dendritic cells or macrophages}
- A61K 2039/5156: {expressing foreign proteins}
- A61K 2039/5158: {Antigen-pulsed cells, e.g. T-cells}
- A61K 2039/55533: {IL-2}
- C12N 2500/90: Serum-free medium, which may still contain naturally-sourced components
- C12N 2501/04: Immunosuppressors, e.g. cyclosporin, tacrolimus
- C12N 2501/2302: Interleukin-2 (IL-2)
- C12N 2501/2315: Interleukin-15 (IL-15)
- C12N 2501/2321: Interleukin-21 (IL-21)
- C12N 2501/24: Interferons [IFN]
- C12N 2501/603: Oct-3/4
- C12N 2502/11: blood or immune system cells
- C12N 2506/30: from cancer cells, e.g. reversion of tumour cells
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A01N: PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF (preservation of food or foodstuff A23); BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES (preparations for medical, dental or toiletry purposes which kill or prevent the growth or proliferation of unwanted organisms A61K); PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

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.

## Claims

### Claim 1

A method for treating a subject with a cancer, the method comprising administering expanded tumor infiltrating lymphocytes (TILs) comprising:
(a) performing a first expansion by (i) thawing cryopreserved dissociated tumor materials comprising a first population of TILs from a tumor that was resected from the subject, dissociated after the resection, and cryopreserved after the dissociation, and (ii) 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;
(b) 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 third population of TILs is a therapeutic 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 (a) to step (b) occurs without opening the system;
(c) harvesting the third population of TILs obtained from step (b), wherein the transition from step (b) to step (c) occurs without opening the system;
(d) transferring the harvested third TIL population from step (c) to an infusion bag, wherein the transfer from step (c) to (d) occurs without opening the system;
(e) cryopreserving the infusion bag comprising the harvested TIL population from step (d) using a cryopreservation process; and
(f) administering a therapeutically effective dosage of the harvested TIL population from the infusion bag in step (e) to the subject.

### Claim 2 (depends on 1)

The method according to claim 1, wherein the dissociated tumor materials comprise a tumor digest.

### Claim 3 (depends on 1)

The method according to claim 1, wherein the dissociated tumor materials comprise one or more tumor fragments.

### Claim 4 (depends on 1)

The method according to claim 1, wherein obtaining the dissociated tumor materials comprises mechanically disrupting the tumor resected from the subject.

### Claim 5 (depends on 1)

The method according to claim 1, wherein obtaining the dissociated tumor materials comprises enzymatically disrupting the tumor resected from the subject.

### Claim 6 (depends on 1)

The method according to claim 1, wherein the third population of TILs harvested in step (c) comprises sufficient TILs for administering a therapeutically effective dosage of the TILs in step (f).

### Claim 7 (depends on 6)

The method according to claim 6, wherein the therapeutically effective dosage in step (f) comprises from about 1×109 to about 9×1010 TILs.

### Claim 8 (depends on 6)

The method according to claim 6, wherein the therapeutically effective dosage in step (f) comprises from about 1×109 to about 5×109 TILs.

### Claim 9 (depends on 6)

The method according to claim 6, wherein the therapeutically effective dosage in step (f) comprises from about 5×109 to about 1×1010 TILs.

### Claim 10 (depends on 6)

The method according to claim 6, wherein the therapeutically effective dosage in step (f) comprises from about 1×1010 to about 5×1010 TILs.

### Claim 11 (depends on 1)

The method according to claim 1, wherein the APCs comprise peripheral blood mononuclear cells (PBMCs).

### Claim 12 (depends on 11)

The method according to claim 11, wherein the PBMCs are supplemented at a ratio of about 1:25 TIL:PBMCs.

### Claim 13 (depends on 1)

The method according to claim 1, wherein prior to administering a therapeutically effective dosage of TIL cells in step (f), a non-myeloablative lymphodepletion regimen has been administered to the subject.

### Claim 14 (depends on 13)

The method according to claim 13, where the non-myeloablative lymphodepletion regimen comprises the steps of administration of cyclophosphamide at a dose of 60 mg/m2/day for two days followed by administration of fludarabine at a dose of 25 mg/m2/day for five days.

### Claim 15 (depends on 1)

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 (f).

### Claim 16 (depends on 15)

The method according to claim 15, 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.

### Claim 17 (depends on 1)

The method according to claim 1, wherein the cancer is selected from the group consisting of melanoma (including metastatic melanoma), ovarian cancer, cervical cancer, non-small-cell lung cancer (NSCLC), lung cancer, bladder cancer, breast cancer, cancer caused by human papilloma virus, head and neck cancer (including head and neck squamous cell carcinoma (HNSCC)), renal cancer, and renal cell carcinoma.

### Claim 18 (depends on 1)

The method according to claim 1, wherein the cancer is melanoma (including metastatic melanoma).

### Claim 19 (depends on 1)

The method according to claim 1, wherein the first expansion in step (a) and the second expansion in step (b) are each individually performed within a period of 11 days.

### Claim 20 (depends on 1)

The method according to claim 1, wherein steps (a) through (d) are performed in about 10 days to about 22 days.

### Claim 21 (depends on 1)

The method according to claim 1, wherein steps (a) through (d) are performed in about 15 days to about 22 days.

### Claim 22 (depends on 1)

The method according to claim 1, wherein steps (a) through (d) are performed in about 20 days to about 22 days.

### Claim 23 (depends on 1)

The method according to claim 1, wherein the closed container in step (a) and/or step (b) is a gas-permeable bag.