Processes for generating til products using pd-1/tigit talen double knockdown
AssigneesIovance Biotherapeutics, Inc., Cellectis
InventorsRafael CUBAS, Frederick G. VOGT, Hequn YIN, Marcus MACHIN, Anand VEERAPATHRAN, Brittany BUNCH, Viktoria GONTCHAROVA, Rongsu QI, Alex BOYNE, Alexandre JUILLERAT, Laurent POIROT
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Abstract
The present invention provides methods for preparing expanded tumor infiltrating lymphocytes (TILs) having reduced expression of PD-1 and TIGIT using sequential electroporation of two TALEN systems targeting PD-1 and TIGIT. Such TILs find use in therapeutic treatment regimens for cancer patients.
Claims27
A method for preparing expanded tumor infiltrating lymphocytes (TILs) obtained from a tumor resected from a patient having reduced expression of PD-1 and TIGIT, comprising:
- (a) culturing a first population of TILs from the tumor from the patient in a first cell culture medium comprising IL-2 for about 5-7 days to produce a second population of TILs;
- (b) activating the second population of TILs for about 2-4 days, to produce a third population of TILs;
- (c) introducing a first TALE nuclease (TALEN) system targeting a first gene selected from the group consisting of a gene encoding PD-1 and a gene encoding TIGIT into at least a portion of the third population of TILs, to produce a fourth population of TILs;
- (d) resting the fourth population of TILs in the first cell culture medium comprising IL-2 for about 2 to 3 days;
- (e) introducing a second TALEN system targeting a second gene selected from the group consisting of the gene encoding PD-1 and the gene encoding TIGIT into at least a portion of the fourth population of TILs to produce a fifth population of TILs, wherein the first gene and the second gene are different;
- and
- (f) culturing the fifth population of TILs in a second cell culture medium comprising antigen presenting cells (APCs), OKT-3, and IL-2 for about 7-11 days, to produce sixth population of TILs having reduced expression of the first gene and the second gene.
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(canceled)
-25. (canceled)
The method of claim 24 or 251, wherein the target sequence of the PD-1 targeting TALEN system comprise the nucleic acid sequence of SEQ ID NO. 18 and the target sequence of the TIGIT targeting TALEN system comprise the nucleic acid sequence of SEQ ID NO. 23 or 28.
33-81. (canceled)
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(canceled)
A method for treating a cancer patient, comprising:
- (a) obtaining a first population of TILs from a tumor sample resected from the cancer patient by processing a tumor sample obtained from the cancer patient into multiple tumor fragments, or from a tumor sample obtained from the cancer patient by surgical resection, needle biopsy, core biopsy, small biopsy, or other means;
- (b) culturing the first population of TILs in a first cell culture medium comprising IL-2 for about 5-7 days to produce a second population of TILs;
- (c) activating the second population of TILs for about 2-4 days, to produce a third population of TILs;
- (d) introducing a first TALE nuclease (TALEN) system targeting a first gene selected from the group consisting of a gene encoding PD-1 and a gene encoding TIGIT into at least a portion of the third population of TILs, to produce a fourth population of TILs;
- (e) resting the fourth population of TILs in the first cell culture medium comprising IL-2 for about 2 to 3 days;
- (f) introducing a second TALEN system targeting a second gene selected from the group consisting of the gene encoding PD-1 and the gene encoding TIGIT into at least a portion of the fourth population of TILs to produce a fifth population of TILs, wherein the first gene and the second gene are different;
- (g) culturing the fifth population of TILs in a second cell culture medium comprising antigen presenting cells (APCs), OKT-3, and IL-2 for about 7-11 days, to produce sixth population of TILs having reduced expression of the first gene and the second gene;
- and
- (h) administering a therapeutically effective dosage of the sixth population of TILs to the cancer patient.
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Dependencies are inferred from the claim text.