Use of PI3K M-TOR and AKT inhibitors to induce FOXP3 expression and generate regulatory T cells
The invention relates to a method of inducing Foxp3 expression in a T cell comprising (i) stimulating a T cell (ii) inhibiting signalling via PI3K alpha or PI3K delta or m-TOR or Akt in said T cell, wherein said inhibition is commenced 10 to 22 hours after the stimulation of (i). The invention also relates to certain uses of PI3K inhibitors, PI3K inhibitors for particular uses, and kits.
1. A method for generating a regulatory T-cell from a naïve T cell, comprising
(i) stimulating the naïve T cell with a T cell receptor (TCR) stimulant;
(ii) removing the TCR stimulant;
(iii) contacting the stimulated naïve T cell with an exogenous inhibitor of PI3K alpha, PI3K delta, m-TOR, or Akt, wherein said contacting is commenced 10 to 22 hours after the stimulation of (i), and after the removal of the TCR stimulant.
2. A method for generating a regulatory T-cell comprising treating a stimulated CD8− T cell with phosphatidyl inositol 3 kinase (PI3K) inhibitor wherein said inhibitor inhibits PI3K alpha and/or PI3K delta.
3. A method of inducing de novo Foxp3 expression in a naïve T cell comprising:
(i) stimulating the naïve T cell;
(ii) halting the stimulating of (i) after about 10 to 22 hours of stimulation;
(iii) contacting the stimulated naïve T cell with an exogenous inhibitor of PI3K alpha, PI3K delta, m-TOR, or Akt, wherein the contacting is commenced 10 to 22 hours after the stimulation of (i), and after the halting of (ii); and
(iv) determining that Foxp3 expression has been induced in said cells wherein said T cells do not comprise or have been depleted of CD8 + T cells and are CD4 + .
4. A method of inducing de novo Foxp3 expression in a naïve T cell comprising:
(i) stimulating the naïve T cell with a T cell receptor (TCR) stimulant for about 10 to 22 hours;
(ii) contacting the stimulated naïve T cell with an exogenous inhibitor of PI3K alpha, PI3K delta, m-TOR, or Akt, wherein the contacting is commenced 10 to 22 hours after the stimulation of (i); and
(iii) determining that Foxp3 expression has been induced in said cells wherein said T cells do not comprise or have been depleted of CD8 + T cells and are CD4 + .
5. A method of inducing Foxp3 expression in a previously stimulated naïve T cell which does not express Foxp3, comprising contacting said previously stimulated naïve T cell with an inhibitor of PI3K alpha, PI3K delta, m-TOR, or Akt, wherein said contacting is commenced 10 to 22 hours after the stimulation, and wherein determining that Foxp3 expression has been induced in said contacted T cells and wherein said T cells to not comprise or have been depleted of CD8 + T cells and are CD4 + .
6. A method of treating a subject in need of regulatory T cell(s) comprising
(i) removing a sample comprising a T cell from a subject
(ii) stimulating said T cell
(iii) optionally withdrawing said stimulation
(iv) inhibiting signalling via PI3K alpha or PI3K delta or m-TOR or Akt in said T cell, wherein said inhibition is commenced 10 to 22 hours after the stimulation of (i); and
(v) reintroducing said T cell to said subject.
7. The method according to claim 4 wherein said inhibition is commenced about 17 to 19 hours after the stimulation.
8. The method according to claim 7 wherein said inhibition is commenced about 18 hours after the stimulation.
9. The method according to claim 4 wherein the inhibiting signalling step comprises inhibiting signalling via PI3K alpha or PI3K delta.
10. The method according to claim 9 wherein inhibiting signalling via PI3K alpha or PI3K delta comprises contacting said cell with PI3K inhibitor, and wherein said inhibitor inhibits PI3K alpha and/or PI3K delta.
11. The method according to claim 4 further comprising withdrawing said stimulation no later than at the time of inhibiting signalling.
12. The method according to claim 4 wherein stimulating said T cell comprises stimulating said T cell via the T cell receptor (TCR).
13. A method of inducing differentiation of a naïve T cell into a regulatory T cell comprising contacting said naïve T cell with a PI3K inhibitor wherein said inhibitor inhibits PI3K alpha and/or PI3K delta.
14. The method according to claim 9 wherein said PI3K alpha and/or PI3K delta inhibitor comprises LY294002.
15. A kit comprising
(i) a TCR stimulant; and
(ii) an inhibitor of PI3K alpha or delta.
16. The kit according to claim 15 wherein said TCR stimulant comprises anti-TCR and anti-CD28 antibody.
17. The kit according to claim 15 further comprising an m-TOR inhibitor.
18. The kit according to claim 17 wherein said m-TOR inhibitor is rapamycin.
19. The kit according to claim 15 further comprising an Akt inhibitor.