Methods of growing tumor infiltrating lymphocytes in gas-permeable containers
Inventors
Rosenberg, Steven A. • Dudley, Mark E. • Stroncek, David • Sabatino, Marianna • Jin, Jianjian • Somerville, Robert • Wilson, John R.
Assignees
Wilson Wolf Manufacturing LLC • US Department of Health and Human Services
Publication Number
US-11401503-B2
Publication Date
2022-08-02
Expiration Date
2032-03-20
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Abstract
An embodiment of the invention provides a method of promoting regression of cancer in a mammal comprising obtaining a tumor tissue sample from the mammal; culturing the tumor tissue sample in a first gas permeable container containing cell medium therein; obtaining tumor infiltrating lymphocytes (TIL) from the tumor tissue sample; expanding the number of TIL in a second gas permeable container containing cell medium therein using irradiated allogeneic feeder cells and/or irradiated autologous feeder cells; and administering the expanded number of TIL to the mammal. Methods of obtaining an expanded number of TIL from a mammal for adoptive cell immunotherapy are also provided.
Core Innovation
The invention provides a method of promoting regression of cancer in a mammal by obtaining a tumor tissue sample; culturing this sample in a first gas permeable container with cell medium; isolating tumor infiltrating lymphocytes (TIL) from the tumor tissue; expanding the number of TIL in a second gas permeable container using irradiated allogeneic and/or autologous feeder cells; and administering the expanded TIL to the mammal. This method involves culturing tumor fragments submerged under a suitable volume of medium on gas permeable material that facilitates gas exchange, growth, and viability of TIL.
The problem addressed is the complexity and technical challenges in growing TIL for adoptive cell therapy using existing methods involving non-gas permeable containers. Traditional procedures are sophisticated, labor-intensive, use more reagents, involve frequent feeding, and require specialized equipment, all of which introduce technical, regulatory, and logistic challenges limiting the successful use of TIL therapy.
This invention solves these problems by using gas permeable containers which simplify procedures, reduce labor and reagent use, protect cells better from microbial contamination, and may reduce the number of containers needed. This approach decreases final culture volume, incubator capacity, and frequency of feeding (every three to four days), and is potentially automatable. The method also allows for expansion of TIL expressing specific antigenicities, including modifications with T-cell growth factors or receptors, providing broader applicability in treating various cancers.
Claims Coverage
The patent includes two independent claims describing methods of obtaining and expanding tumor infiltrating lymphocytes (TIL) using gas permeable containers, selecting TIL based on IFN-γ release, and subsequent expansion with feeder cells and specific culture mediums for adoptive immunotherapy.
Method of obtaining selected TIL capable of lysing cancer cells using gas permeable containers
The method comprises obtaining a tumor tissue sample; culturing the sample for 14 to 15 days in a first gas permeable container with cell medium comprising IL-2; co-culturing TIL with autologous tumor cells; selecting TIL based on IFN-γ release; expanding the selected TIL in a second gas permeable container with IL-2 and OKT3 using irradiated allogeneic and/or autologous feeder cells for about 14 days; and wherein at least a portion of the expanded TIL express CD3 and CD8.
Features enhancing TIL expansion
Expanding TIL by at least about 1000-fold; using about 1×10^9 to 4×10^9 irradiated feeder cells at ratios of about 1 TIL to 20-100 feeder cells; culturing with irradiated peripheral blood mononuclear cells; feeding cells every third or fourth day; and seeding containers with about 1×10^6 TIL.
Method of obtaining expanded TIL from a human including fragmentation and expansion
Obtaining a tumor tissue sample from a human and fragmenting it, culturing fragments for 14 to 15 days in a first gas permeable container with IL-2 to obtain TIL; co-culturing with autologous tumor cells; selecting TIL by IFN-γ release; seeding second gas permeable container with about 1×10^6 selected TIL; and culturing with IL-2, OKT3, and irradiated allogeneic feeder cells at about 1:50 ratio for 14 days to obtain expanded TIL expressing CD3 and CD8.
Scope of applicable tumors and mammals
The tumor tissue sample can be derived from a broad list of cancers including melanoma and others recited explicitly. The mammal can be a human.
Duration of the method
The method duration is about 42 days.
The claims cover methods of culturing, selecting, and expanding TIL in gas permeable containers using irradiated feeder cells and specific culture conditions including IL-2 and OKT3, with features enhancing yield, viability, and specificity of TIL for adoptive immunotherapy, particularly in human cancers including melanoma.
Stated Advantages
Simpler and less labor-intensive methods of growing TIL compared to non-gas permeable containers.
Reduced use of reagents and simplified equipment requirements.
Enhanced protection against microbial contamination due to closed gas permeable systems.
Reduced number of containers used, lowering labor and contamination risk.
Lower final culture volume reduces incubator space and media needed, simplifying cell processing.
Reduced feeding frequency (every 3-4 days) maintains cell viability and improves culture reproducibility.
Potential for automation of culturing and expansion processes.
Facilitates faster and more efficient adoptive cell therapy allowing treatment of more patients earlier in disease.
Enables treating patients previously not treatable due to insufficient TIL numbers.
Methods are more compliant with cGMP, aiding regulatory approval for clinical use.
Documented Applications
Treatment and prevention of various cancers including melanoma, metastatic melanoma, and a wide range of human cancers such as acute lymphocytic leukemia, breast cancer, colon cancer, lung cancer, ovarian cancer, prostate cancer, and others.
Use in adoptive cell immunotherapy to generate and administer expanded TIL to cancer patients.
Adoptive cell therapy involving administration of TIL expanded using the inventive methods with potential co-administration of T-cell growth factors like IL-2.
Modification of TIL for enhanced expression of T-cell growth factors or T-cell receptors for targeted immunotherapy.
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