Compositions and methods for in vitro activation and expansion of serial killer T cell populations and passive immunization of a cancer patient with tumor cell killing cells
AssigneeAlloplex Biotherapeutics, Inc.
InventorFrank Borriello
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Abstract
The described invention provides a method for in vitro immunoactivation of mononuclear cells by contact with one or more populations of engineered leukocyte stimulator cells genetically engineered to express a core of 3 essential immunomodulator peptides, and optionally additional R immunomodulator peptides, and use of a cell product comprising the expanded and activated mononuclear cell population comprising one or more subpopulations of cytotoxic serial killer cells for passive immunization of a cancer patient not currently under the influence of an immunosuppressive regimen.
Claims18
A method of treating a patient with melanoma, wherein the patient is not currently under the influence of an immunosuppressive regimen, the method comprising:
- preparing a composition comprising a cell product comprising an activated and expanded population of mononuclear cells (MNCs), the MNCs comprising activated and expanded subpopulations of cytotoxic serial killer cells by:
- (a) inducing an immune response in vitro by:
- (1) isolating a population of MNCs derived from peripheral blood or cord blood;
- (2) preparing a population of engineered leukocyte stimulator (ENLST™) cells comprising a population of tumor cells derived from a primary SKMEL-2 melanoma tumor cell line that is genetically engineered by transfecting or transducing recombinant DNA sequences coding for at least three core immunomodulator peptides comprising OX40 Ligand (OX40L), CD27 Ligand (CD70) and CD28 Ligand (CD28L) and selecting for ENLST™ cell clones that stably express the three core immunomodulator peptides, wherein the amino acid sequence of a wild type OX40 Ligand comprises SEQ ID NO:
- 108, the amino acid sequence of a wild type CD27 ligand comprises SEQ ID NO:
- 109, and the amino acid sequence of a wild type CD28 ligand comprises SEQ ID NO:
- 110, SEQ ID NO:
- 111, or both;
- (3) contacting the population of MNCs of step (a)(1) with the ENLST™ cells of step (a)(2) in vitro in a mixed lymphocyte tumor cell reaction (MLTR) assay;
- wherein the contacting is effective to stimulate synergistic expansion of cytotoxic serial killer cells, to form an activated population of MNCs comprising activated subpopulations of cytotoxic serial killer cells;
- (b) culturing the activated MNCs to form a cell product containing an activated and expanded population of MNCs including expanded and activated subpopulations of cytotoxic serial killer cells;
- (c) preparing a unit dose package comprising an individual dose of the activated and expanded cell product, freezing the unit dose packages, and storing the frozen unit dose packages in cryostorage;
- (d) thawing a therapeutic amount of the frozen unit dose packages comprising the cell product under controlled conditions;
- (e) optionally combining the frozen and thawed cell product with a pharmaceutically acceptable carrier to form a pharmaceutical composition;
- and
- (f) administering the therapeutic amount of the cell product of (d) or the pharmaceutical composition of (e) comprising the activated and expanded cell product to the patient with melanoma,
- wherein the therapeutic amount is effective to reduce melanoma burden.
A cell product comprising a population of expanded and activated mononuclear cells comprising activated subpopulations of cytotoxic serial killer cells prepared by a process comprising:
- (a) isolating a population of MNCs derived from peripheral blood or cord blood;
- (b) preparing a population of engineered leukocyte stimulator (ENLST™) cells comprising a population of tumor cells derived from a primary SKMEL-2 melanoma tumor cell line that is genetically engineered by transfecting or transducing recombinant DNA sequences coding for at least three core immunomodulator peptides comprising OX40 Ligand (OX40L), CD27 Ligand (CD70) and CD28 Ligand (CD28L) and selecting for ENLST™ cell clones that stably express the three core immunomodulator peptides, wherein the amino acid sequence of a wild type OX40 Ligand comprises SEQ ID NO:
- 108, the amino acid sequence of a wild type CD27 ligand comprises SEQ ID NO:
- 109, and the amino acid sequence of a wild type CD28 ligand comprises SEQ ID NO:
- 110, SEQ ID NO:
- 111, or both;
- (c) contacting the population of MNCs of step (a) with the ENLST™ cells of step cells of step (b) in vitro in a mixed lymphocyte tumor cell reaction (MLTR) assay to form an activated population of MNCs comprising activated subpopulations of cytotoxic serial killer cells;
- (d) expanding the activated population of MNCs comprising activated subpopulations of serial killer cells in vitro by culturing the activated MNCs to form the cell product comprising an activated and expanded population of MNCs comprising expanded and activated subpopulations of cytotoxic serial killer cells.
Dependencies are inferred from the claim text.