Methods of producing T memory stem cell populations
Inventors
Gattinoni, Luca • Lugli, Enrico • Roederer, Mario • Restifo, Nicholas P.
Assignees
US Department of Health and Human Services
Publication Number
US-11111478-B2
Publication Date
2021-09-07
Expiration Date
2032-09-06
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Abstract
Provided are methods of producing an isolated T memory stem cell population, the method comprising a) isolating naïve T cells from a mammal, wherein the mammal is not a mouse; b) activating the naïve T cells and expanding the numbers of naïve T cells in the presence of one or more non-specific T cell stimuli, one or more cytokines, and a GSK-3beta inhibitor. Also provided are methods of producing an isolated T memory stem cell population, the method comprising a) isolating lymphocytes from a mammal; b) sorting the lymphocytes using flow cytometry into a population comprising a phenotype comprising i) CD95+, CD45RO−, and CCR7+; and ii) CD62L+ or one or more of CD27+, CD28+, CD45RA+, and CD127+ to produce an isolated T memory stem cell population. Further embodiments of the invention provide related cells, populations of cells, pharmaceutical compositions, and methods of treating or preventing cancer.
Core Innovation
The invention provides methods and compositions for producing isolated T memory stem cell (TSCM) populations from mammals, excluding mice. One method involves isolating naïve T cells from the mammal, then activating and expanding these cells using one or more non-specific T cell stimuli, cytokines, and a glycogen synthase kinase (GSK)-3beta inhibitor. Another method involves isolating lymphocytes from a mammal and sorting them by flow cytometry to obtain a population characterized by a specific phenotype, including markers such as CD95+, CD45RO−, CCR7+, and CD62L+ or other combinations of markers like CD27+, CD28+, CD45RA+, and CD127+.
The problem addressed is that adoptive cell therapy (ACT) using tumor-reactive T cells faces challenges because T cells isolated from peripheral blood may lack sufficient tumor-specific reactivity or persistence after reinfusion. Improved methods are needed to obtain antigen-specific T cells from peripheral blood that exhibit enhanced tumor-specific activity and persistence in patients.
The invention further characterizes TSCM cells as a distinct memory T cell population with enhanced stem cell-like qualities, including increased self-renewal, multipotency, and capacity to repopulate differentiated effector lymphocytes in response to antigenic stimuli. The use of GSK-3beta inhibitors, believed to activate Wnt signaling, allows effective in vitro generation of TSCM cells by delaying or preventing T cell differentiation. The invention also describes methods for expanding these cells and their use in pharmaceutical compositions and cancer treatment.
Claims Coverage
The patent discloses three main inventive features focused on methods for producing isolated T memory stem cell populations, each involving particular sorting phenotypes and optional expansion or genetic modification steps.
Method of producing isolated T memory stem cells by flow cytometry sorting in non-human primates
A method comprising isolating lymphocytes from a mammal and sorting them using flow cytometry into a population characterized by the phenotype: (i) CD95+ and/or CXCR3+; and (ii) CD45RA+, CCR7+, and CD28+, thereby producing an isolated T memory stem cell population.
In vitro expansion of isolated T memory stem cells
The method further includes expanding the isolated T memory stem cells in vitro to increase their numbers.
Genetic modification of isolated T memory stem cells with antigen-specific receptors
The method further comprises transducing the isolated T memory stem cells with a nucleotide sequence encoding a chimeric antigen receptor (CAR) or a T cell receptor (TCR), where the CAR or TCR has antigenic specificity for a cancer antigen or a viral antigen.
Collectively, these inventive features describe methods to isolate, expand, and genetically modify T memory stem cells with defined surface phenotypes for therapeutic use, particularly in non-mouse mammals such as humans and non-human primates.
Stated Advantages
TSCM cells possess enhanced capacity for self-renewal and multipotency compared with central memory T cells.
TSCM cells exhibit improved persistence and survival upon adoptive transfer compared with other T cell subsets.
TSCM cells have superior proliferative ability and rapid effector function acquisition after stimulation.
Methods allow for expansion and generation of TSCM cells in vitro using GSK-3beta inhibitors, enabling production of large numbers of these cells.
Adoptive transfer of TSCM cells leads to superior antitumor activity and treatment efficacy compared to other memory or effector T cells.
TSCM cells can be genetically modified with antigen-specific receptors, improving specificity and therapeutic utility.
Documented Applications
Use of TSCM cells and methods for producing them in adoptive cell therapy for treating or preventing various cancers, including melanoma, mesothelioma, leukemia, lymphomas, and solid tumors.
Production of pharmaceutical compositions containing isolated TSCM cells for administration to mammals.
Genetic modification of TSCM cells with chimeric antigen receptors or T cell receptors specific for cancer or viral antigens, for therapeutic immunotherapy.
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