GENERATION OF T-CELLS BY DIRECT REPROGRAMMING FROM FIBROBLASTS AND MSC
The invention relates to a method comprising (a) providing a vertebrate somatic cell, (b) inducing expression in said cell of at least three transcription factors, wherein said transcription factors are: i. Tbet (Tbx21), ii. Tcf7, and iii. Ets1; (c) incubating said cell to allow transdifferentiation. Suitably said cell is transdifferentiated to an immune effector cell. The invention also relates to cells, nucleic acids, pharmaceutical compositions, and methods of treatment.
1 . A method comprising
(a) providing a vertebrate somatic cell,
(b) inducing expression in said cell of at least three transcription factors, wherein said transcription factors are:
Tbet (Tbx21),
Tcf7, and
Ets1;
(c) incubating said cell to allow transdifferentiation.
2 - 12 . (canceled)
13 . A method according to claim 1 wherein said cell is transdifferentiated to an immune effector cell.
14 . A method according to claim 1 wherein said cell is transdifferentiated to an alpha/beta T cell, or a gamma/delta T cell.
15 . A method according to claim 1 wherein said cell is transdifferentiated to a CD8+ T cell.
16 . A method according to claim 1 wherein said cell is transdifferentiated to a CD4+ T cell.
17 . A method according to claim 1 wherein the vertebrate somatic cell is selected from the group consisting of: a mesenchymal stem cell (MSC), a skin fibroblast, an endothelial cell, a keratinocyte, and a hepatocyte.
18 . (canceled)
19 . A method according to claim 17 wherein the vertebrate somatic cell is a mesenchymal stem cell.
20 . A method according to claim 17 wherein the vertebrate somatic cell is a skin fibroblast.
21 . A method according to claim 17 wherein the vertebrate somatic cell is a human mesenchymal stem cell (hMSC), and wherein said hMSC is derived from bone marrow.
22 . A method according to claim 1 wherein the immune effector cell of (c) is a cell which expresses one or more gene(s) selected from the group consisting of: CD3, CD45, CD2, CD5, CD7, CD4, and CD8.
23 . A method according to claim 1 wherein the immune effector cell of (c) is a cell which expresses one or more gene(s) selected from the list consisting of: CD105 and CD73 at a lower level compared to the expression of the same gene(s) in a vertebrate somatic cell of (a).
24 . A method according to claim 1 wherein the immune effector cell of (c) is a cell which does not express one or more gene(s) selected from the list consisting of: CD105 and CD73.
25 . (canceled)
26 . A method according to claim 1 wherein inducing expression of at least three transcription factors comprises:
providing nucleic acid(s) comprising three nucleotide sequences, each nucleotide sequence encoding a transcription factor selected from said at least three transcription factors, each nucleotide sequence being operatively linked to a promoter sequence capable of directing expression of said transcription factor; and
introducing said nucleic acid into said cell.
27 . A method according to claim 26 , wherein introducing said nucleic acid into said cell comprises electroporation of said cell.
28 . A cell obtained by the method according to claim 1 .
29 . An isolated nucleic acid comprising nucleotide sequences encoding Tbet, Ets1 and Tcf7, each nucleotide sequence being operatively linked to a promoter sequence capable of directing expression of said nucleotide sequence encoding Tbet, Ets1 and Tcf7.
30 . An isolated nucleic acid according to claim 29 , wherein each said nucleotide sequence encoding Tbet, Ets1 and Tcf7 is operatively linked to a respective promoter sequence.
31 . An isolated nucleic acid according to claim 29 , wherein said nucleotide sequences encoding Tbet, Ets1 and Tcf7 are all operatively linked to a single promoter sequence.
32 . An isolated nucleic acid according to claim 29 , wherein said nucleotide sequences are arranged in the order 5′-promoter sequence-nucleotide sequence encoding Tbet-nucleotide sequence encoding Ets1-nucleotide sequence encoding Tcf7-3′.
33 - 35 . (canceled)
36 . A method of treating a subject comprising administering to said subject a cell according to claim 28 .