Expansion of renewable stem cell populations using modulators of PI 3-kinase
First Claim
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1. A method of ex vivo expanding and inhibiting differentiation of a population of stem cells, the method comprising:
- (a) providing the cells ex vivo with conditions for cell proliferation;
(b) ex vivo providing the cells with an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase;
thereby ex vivo expanding and inhibiting differentiation of the population of stem cells.
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Abstract
Provided are ex vivo and in vivo methods of expanding renewable stem cells using modulators of PI 3-kinase activity, expanded populations of renewable stem cells, and uses thereof.
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Citations
147 Claims
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1. A method of ex vivo expanding and inhibiting differentiation of a population of stem cells, the method comprising:
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(a) providing the cells ex vivo with conditions for cell proliferation;
(b) ex vivo providing the cells with an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase;
thereby ex vivo expanding and inhibiting differentiation of the population of stem cells. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 46, 47, 48, 49, 50)
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23. A method of transducing expanded, undifferentiated stem cells with an exogene, the method comprising:
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(a) obtaining a population of stem cells;
(b) expanding and inhibiting differentiation of said stem cells by;
(i) providing said stem cells with conditions for cell proliferation;
(ii) providing said stem cells with an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase;
wherein steps (i) and (ii) are effected in vitro or ex vivo, thereby expanding and inhibiting differentiation of said stem cells; and
(c) transducing said expanded, undifferentiated stem cells with the exogene. - View Dependent Claims (24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45)
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51. A method of hematopoietic stem cells transplantation into a recipient, the method comprising:
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(a) obtaining a population of hematopoietic stem cells;
(b) ex vivo expanding and inhibiting differentiation of said hematopoietic stem cells by;
(i) ex vivo providing said hematopoietic stem cells with conditions for cell proliferation;
(ii) providing said hematopoietic stem cells ex vivo with an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase;
thereby expanding and inhibiting differentiation of said stem hematopoietic cells; and
(c) transplanting said hematopoietic stem cells into the recipient. - View Dependent Claims (52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72)
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73. A method of adoptive immunotherapy comprising:
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(a) obtaining progenitor hematopoietic cells from a patient;
(b) ex vivo expanding and inhibiting differentiation of said hematopoietic cells by;
(i) providing said progenitor hematopoietic cells ex vivo with conditions for cell proliferation;
(ii) providing said progenitor hematopoietic cells with an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase;
thereby expanding and inhibiting differentiation of said progenitor hematopoietic cells; and
(c) transplanting said progenitor hematopoietic cells into a recipient. - View Dependent Claims (74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94)
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95. A method of mobilization of bone marrow stem cells into the peripheral blood of a donor for harvesting the cells comprising:
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(a) administering to the donor an effective concentration of a a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase, thereby expanding and inhibiting differentiation of a population of bone marrow stem cells; and
(b) harvesting the cells by leukopheresis.
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96. A method of inhibiting maturation/differentiation of erythroid precursor cells for treatment of a β
- -hemoglobinopathic patient comprising administering to said patient an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase, thereby expanding and inhibiting differentiation of a population of stem cells of said patient such that upon removal of said modulator of PI 3-kinse from said patient, said stem cells undergo accelerated maturation resulting in elevated fetal hemoglobin production,
thereby ameliorating symptoms of β
-hemoglobinopathy in said patient. - View Dependent Claims (98, 99, 100, 101, 102)
- -hemoglobinopathic patient comprising administering to said patient an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase, thereby expanding and inhibiting differentiation of a population of stem cells of said patient such that upon removal of said modulator of PI 3-kinse from said patient, said stem cells undergo accelerated maturation resulting in elevated fetal hemoglobin production,
- 97. The method of 96, further comprising the step of administering to said patient a cytokine.
- 108. A method of preservation of undifferentiated stem cells comprising providing the undifferentiated stem cells with an effective concentration of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase of said undifferentiated stem cells, wherein said providing is performed in at least one of the steps of harvesting, isolating and storage of the undifferentiated hematopoietic cells.
- 129. Stem cell collection bags, stem cell separation and stem cell washing buffers supplemented with an amount of a modulator of PI 3-kinase activity, said modulator selected capable of downregulating a PI 3-kinase activity or an expression of a gene encoding a PI 3-kinase, said amount sufficient to inhibit differentiation of a population of undifferentiated hematopoietic cells.
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139. An assay for determining whether a modulator of PI 3-kinase activity is capable of inhibiting differentiation of cells, the assay comprising:
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(a) culturing a population of cells capable of differentiating, in the presence or absence of said modulator of PI 3-kinase activity; and
(b) assessing changes in differentiation of said cells, wherein an increase in differentiation as compared to untreated cells indicates a modulator of PI 3-kinase activity incapable of inhibiting differentiation, and whereas a lack of or decrease in differentiation as compared to untreated cells, indicates a modulator of PI 3-kinase activity capable of inhibiting differentiation. - View Dependent Claims (140, 141, 142, 143, 144, 145, 146)
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147. A method of ex vivo expanding and inhibiting differentiation of a population of stem cells, the method comprising:
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(a) providing the cells ex vivo with conditions for cell proliferation;
(b) ex vivo reducing a capacity of said stem cells in responding to signaling pathways involving a PI 3-kinase activity;
thereby ex vivo expanding and inhibiting differentiation of the population of stem cells.
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Specification