Regulatory T cells and methods of identifying, obtaining and using to treat immuno-based disorders

1. A method of identifying and isolating a population of immunosuppressive regulatory T-cells, the method comprising:

screening, a sample comprising a population of T-cells to detect a level of cellular expression of at least three different biomarkers, the at least three different biomarkers including a CD4 biomarker, CD25 biomarker and a CD6 biomarker; wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample and wherein detection of the CD6 low/− expression comprises fluorescent intensity of below 50%, 40%, 30%, 20% or 10% as compared to fluorescence intensity observed for all cells in the sample, and isolating the subpopulation of immunosuppressive regulatory T-cells.

2. The method of claim 1 , wherein the method further comprises enriching the population of immunosuppressive regulatory T-cells.

3. The method of claim 2 , wherein the method further comprises contacting the population of immunosuppressive regulatory T-cells with a stimulatory composition thereby expanding population of immunosuppressive regulatory T-cells, wherein the stimulatory composition comprises a T cell receptor (TCR)/cluster of differentiation 3 (CD3) activator that is antigen specific for the population of immunosuppressive regulatory T-cells.

4. The method of claim 1 , wherein the screening step further includes screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD127 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD127 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample.

5. The method of claim 1 , wherein the screening step further includes screening the sample comprising a population of T-cells to detect a level of cellular expression of a HLA-Dr biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− HLA-Dr low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample.

6. The method of claim 1 , wherein the screening step further includes screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD49d biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD49d low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD38 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD38 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD45RA biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD45RA low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a FoxP3 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− FoxP3 + or CD4 + CD25 + CD6 low/− FoxP3 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CTLA-4 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CTLA-4 + or CD4 + CD25 + CD6 low/− CTLA-4 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a GITR biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− GITR + or CD4 + CD25 + CD6 low/− GITR low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a LAG-3 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− LAG-3 + or CD4 + CD25 + CD6 low/− LAG-3 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD39 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD39 + or CD4 + CD25 + CD39 low/− CD39 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a Helios biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− Helios + or CD4 + CD25 + CD6 low/− Helios low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a FcRL3 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− FcRL3 + or CD4 + CD25 + CD6 low/− FcRL3 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CCR7 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CCR7 + or CD4 + CD25 + CD6 low/− CCR7 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CCR4 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CCR4 + or CD4 + CD25 + CD6 low/− CCR4 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CCR8 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CCR8 + or CD4 + CD25 + CD6 low/− CCR8 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD62L biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD62L + or CD4 + CD25 + CD6 low/− CD62L low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a ICOS biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− ICOS + or CD4 + CD25 + CD6 low/− ICOS low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD103 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD103 + or CD4 + CD25 + CD6 low/− CD103 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a PD-1 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− PD-1 + or CD4 + CD25 + CD6 low/− PD-1 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD134 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD134 + or CD4 + CD25 + CD6 low/− CD134 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a GARP biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− GARP + or CD4 + CD25 + CD6 low/− GARP low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD45RB biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD45RB + or CD4 + CD25 + CD6 low/− CD45RB low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD45RO biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD45RO + or CD4 + CD25 + CD6 low/− CD45RO low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD95 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD95 + or CD4 + CD25 + CD6 low/− CD95 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample;

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD122 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD122 + or CD4 + CD25 + CD6 low/− CD122 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample; and/or

screening the sample comprising a population of T-cells to detect a level of cellular expression of a CD8 biomarker, wherein detection of a subpopulation of T-cells comprising a CD4 + CD25 + CD6 low/− CD8 + or CD4 + CD25 + CD6 low/− CD8 low/− expression pattern identifies the population of immunosuppressive regulatory T-cells in the sample.

7. A method of identifying and isolating two or more distinct immunosuppressive regulatory T-cell populations, the method comprising the steps of

screening a sample comprising a population of T-cells to detect a level of cellular expression of a CD4 biomarker, a CD25 biomarker, a CD6 biomarker, and one additional biomarker, the one additional biomarker being a FoxP3 biomarker, a CTLA-4 biomarker, a CD45RA biomarker, or a HLA-Dr biomarker;

wherein detection of two distinct populations of immunosuppressive regulatory T-cells is based upon i) T-cells comprising a CD4 + CD25 + CD6 low/− FoxP3 + expression pattern and CD4 + CD25 + CD6 low/− FoxP3 low/− expression pattern, ii) T-cells comprising a CD4 + CD25 + CD6 low/− CTLA-4 + expression pattern, and CD4 + CD25 + CD6 low/− CTLA-4 low/− expression pattern, iii) T-cells comprising a CD4 + CD25 + CD6 low/− CD45RA + expression pattern and CD4 + CD25 + CD6 low/− CD45RA low/− expression pattern, or iv) T-cells comprising a CD4 + CD25 + CD6 low/− HLA-Dr + expression pattern and CD4 + CD25 + CD6 low/− HLA-Dr low/− expression pattern

wherein cells designated as CD4 + , CD25 + , FoxP3 + , CTLA-4 + , CD45RA + or HLA-Dr + exhibit 10% or more, 20% or more, 30% or more, 40% or more, or 50% or more fluorescence intensity as compared to the fluorescence intensity observed for all cells being screened;

cells designated as CD6 low , FoxP3 low , CTLA-4 low , CD45RA low or HLA-Dr low exhibit 50% or less, 40% or less, 30% or less, 20% or less, or 10% or less fluorescence intensity as compared to the fluorescence intensity observed for all cells being screened;

cells designated as CD6 − , FoxP3 − , CTLA-4 − , CD45RA − or HLA-Dr − exhibit 10% or less, 7% or less, 5% or less, 3% or less, or 2% or less fluorescence intensity as compared to the fluorescence intensity observed for all cells being screened;

and isolating the distinct populations of immunosuppressive regulatory T-cells.

8. The method of claim 7 , wherein the method further comprises enriching one or more of the two distinct maturation subsets of immunosuppressive regulatory T-cell populations.

9. The method of claim 8 , wherein the method further comprises contacting the one or more of the two distinct maturation subsets of immunosuppressive regulatory T-cell populations with a stimulatory composition thereby expanding population of immunosuppressive regulatory T-cells.