Tolerogenic dendritic cells, method for their production and uses therof

1. An in vitro method for generating a population of T(Tr-DC) cells having regulatory activity, the method comprising the steps of:

a) generating a tolerogenic human dendritic cell population (Tr-DC) population from a subject that comprises the marker phenotype: CD14 + , CD11c + , CD11b + , CD1a − , CD83 + , CD80 + CD86 + , HLA-DR + , and CD71 + , wherein at least about 92% of the generated human Tr-DC are CD86 + positive;

b) irradiating the Tr-DC population of a);

c) isolating peripheral blood mononuclear cells (PBMCs) or CD4 + T cells from a subject and

d) stimulating said isolated PBMCs or the CD4 + T cells, in culture, with an effective amount of said irradiated Tr-DC cell population, to generate a population of T(Tr-DC) cells;

wherein the subject of step a) is the same or different from the subject of step c).

2. The method of claim 1 , wherein the irradiation is conducted with an exposure of 6000 Rads from an X-ray source.

3. The method of claim 1 , wherein the population of Tr-DC is obtained from a subject by a process comprising the steps of:

a) collecting a sample from the subject;

b) generating Tr-DC cells from the collected sample that express: CD14 + , CD11c + , CD11b + , CD1a − , CD83 + , CD80 + , CD86 + , HLA-DR + , and CD71 + , wherein at least about 92% of the generated human Tr-DC are CD86 + positive and wherein the sample is a blood sample, a spleen sample or a lymph node sample.

4. The method of claim 3 , where the step of generating Tr-DC cells comprises the steps of:

a) collecting peripheral blood mononuclear cells (PBMCs) from a subject;

b) isolating adherent cells from collected PBMCs; and

c) exposing said isolated adherent cells at the start of culture on day 0 to an effective amount of GM-CSF, IL-4 and IL-10 to generate the Tr-DC cells.

5. The method of claim 1 , wherein the ratio of PBMC or CD4 + T cells to irradiated Tr-Dc is 1:10.

6. The method of claim 1 , wherein the isolated PBMCs or isolated CD4+ T cells are stimulated with an effective amount of the irradiated Tr-DC cell population for 10 days.

7. The method of claim 1 , wherein the isolated PBMCs are stimulated with an effective amount of the irradiated Tr-DC cell population in the presence of IL-10.

8. The method of claim 1 , wherein the Tr-DC, and the PBMCs or the CD4 + T cells originate from different subjects.

9. The method of claim 1 , wherein the generated population of T(Tr-DC) cells is anergic; and comprises cells with the following marker phenotype: IL-10 ++ , TGF-β + , IL-4 − , IFN-γ + and IL-2 negative to low.

10. A method of inducing or restoring immune tolerance in a subject in need thereof comprising administering to the subject in need thereof the population of T(Tr-DC) cells produced by the method of claim 1 .

11. The method of claim 10 , wherein the population of T(Tr-DC) cells is anergic; and

comprises cells with the following marker phenotype: IL-10 ++ , TGF-β + , IL-4 − , IFN-γ + and IL-2 negative to low.

12. A method for treating a disease in a subject, wherein the disease is selected from the group consisting of graft versus host disease, organ graft rejection, autoimmune disease, allergies, asthma, and chronic inflammatory disease in a subject in need thereof, the method comprising administering to the subject the population of T(Tr-DC) cells produced by the method of claim 1 .

13. The method of claim 12 , where the autoimmune disease is selected from the group consisting of type 1 diabetes mellitus, autoimmune entheropathy, rheumatoid arthritis, systemic lupus erythematosus, multiple sclerosis and psoriasis.

14. The method of claim 12 , where the chronic inflammatory disease is selected from the group consisting of inflammatory bowel disease, Crohn's disease and vasculitis.

15. The method of claim 12 , wherein the allergy is atopic dermatitis.

16. The method of treating a disease according to claim 12 , wherein the disease is graft versus host disease or organ graft rejection.

17. The method of claim 12 , wherein the population of T(Tr-DC) cells is anergic; and

comprises cells with the following marker phenotype: IL-10 ++ , TGF-β + , IL-4 − , IFN-γ + and IL-2 negative to low.

18. A method for treating a disease in a subject, wherein the disease is selected from the group consisting of: immune dysfunction, Polyendocrinopathy Enteropathy X-linked (IPEX) syndrome, Autoimmune Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy (APECED) syndrome, and OMENN's syndrome, the method comprising administering to the subject the population of T(Tr-DC) cells produced by the method of claim 1 .

19. The method of claim 18 , wherein the population of T(Tr-DC) cells is anergic; and comprises cells with the following marker phenotype: IL-10 ++ , TGF-β + , IL-4 − , IFN-γ + and IL-2 negative to low.