- Email: [email protected]
CD4+ CD25+ regulatory T cells approach the clinic
Cytotherapy (2008) Vol. 10, No. 7, 655656
COMMENTARY
CD4 CD25 regulatory T cells approach the clinic M Edinger In 1995, Sakaguchi et al. [1] (CIT) described that CD4 T cells constitutively expressing the interleukin-2 receptor alpha chain (CD25) are potent suppressors of autoreactive T-cell responses. In their experimental system, they showed that the adoptive transfer of CD4 CD25 T cells inhibited the autoimmunity that is regularly observed in mice after early post-natal thymectomy. These experiments suggested that the suppressive T-cell population is thymus-derived and exported from the thymus only after birth and that there is no other peripheral tolerance mechanism compensating for the lack of these ‘regulatory’ T cells (Treg). A few years later, the transcription factor Foxp3 was identified as a master regulator for the development and peripheral function of those thymus-derived Treg cells, and it was shown that loss of function mutations in the foxp3 gene results in a lack of this natural Treg cell compartment [26]. In mice and humans, such mutations cause severe autoimmunity (scurfy and IPEX, respectively). Interestingly, boys with IPEX syndrome (immune dysregulation, polyendocrinopathy, eteropathy, X-linked) suffer from severe colitis and develop unusual immune responses, for example against nutritional antigens. In experimental models of colitis, Mottet et al. [7] showed that adoptively transferred Treg cells not only inhibit colitis induction but, more importantly, even cure established disease. Thus the adoptive transfer of natural Treg cells seems a promising strategy for the treatment of chronic colitis. In this issue of Cytotherapy, Sumida et al. [8] present methods for the enrichment of natural Treg cells from leukapheresis products of patients with ulcerative colitis (UC). For this purpose, they have adopted a strategy described previously for the isolation of Treg cells from healthy donors in allogeneic stem cell transplantation
[9,10]. This isolation protocol was developed for the prevention of graft-versus-host disease after allogeneic stem cell transplantation, where the co-transplantation of conventional and Treg cells at a 1:1 ratio is envisaged. Sumida et al. [8] have added a CD8-depletion step in addition to the CD19-depletion and repetitive CD25enrichment cycles to get rid of the 25% of CD8 T cells that otherwise contaminate a Treg-enriched cell product. Overall, they demonstrate an approximately 55% enrichment of natural Treg cells from leukapheresis products of colitis patients, based on the proportion of FOXP3 cells in the target cell fraction. However, the final cell product does not contain solely Treg cells but also recently activated CD4 T cells expressing CD25 and probably also some non-regulatory T cells that transiently express FOXP3 after stimulation. Whether such transient FOXP3 expression confers suppressive activity is a matter of debate and doubted by several investigators in the field. Nevertheless, exclusive surface markers for the unequivocal identification and separation of Treg cells from activated conventional T cells have not yet been identified and the protocol presented by Sumida et al. [8] is the best currently available approach for the enrichment of Treg cells under good manufacturing practice (GMP) conditions. Although additional depletion of CD127 cells or enrichment of naive Treg cells (CD45RA ) have been suggested to improve Treg cell purities [1113], such strategies remain to be evaluated at large scales with GMP-compatible reagents. Consequently, as long as we lack a technology that permits the generation of pure Treg cell products, we have to define the necessary degree of purity for each potential clinical application. Although leukocyte depletion from the peripheral blood of patients with UC is not yet a standard treatment world-wide [14], it
Correspondence to: PD Dr M. Edinger, Klinikum der Universita¨t Regensburg, Abt. fu¨r Ha¨matologie & Int. Onkologie, Franz-Josef-Strauss-Allee 11, 93053 Regensburg. E-mail: [email protected]. – 2008 ISCT
DOI: 10.1080/14653240802492705
656
M Edinger
is well accepted and frequently performed in Japan [15]. Thus phase I clinical trials combining leukapheresis for the depletion of leukocytes with the enrichment and reinfusion of autologous Treg cells seems feasible in this setting and no matter of ethical concern. Furthermore, shifting the balance from pro-inflammatory T cell responses to Treg-mediated suppression is expected to be beneficial in this patient group. Beyond safety and feasibility trials, however, the evaluation of efficacy will be a major challenge because large numbers of UC patients undergoing leukapheresis with and without additional Treg cell administrations will be required. In addition, long observation times will be necessary because of the remitting nature of the disease. Finally, the superiority of Treg cell transfers with regard to efficacy and safety will also have to be evaluated in comparison with standard pharmacologic immunosuppression (5-aminosalicylic acid, sulphasalazine, steroids, azathioprine or 6-mercaptopurine and cyclosporine) as well as recently introduced biologicals, such as the anti-tumor necrosis factor antibody infliximab. Thus, although it will be a cumbersome path to prove the efficacy of Treg cell therapies in this clinical situation, the re-infusion of freshly isolated or in vitroexpanded natural Treg cells is a promising strategy for UC patients and the first clinical trials are now in preparation. Declaration of interest: The author reports no conflicts of interest. The author alone is responsible for the content and writing of the paper.
References 1
2
Sakaguchi S, Sakaguchi N, Asano M et al. Immunologic selftolerance maintained by activated T cells expressing IL-2 receptor alpha-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J Immunol 1995;155:115164. Bennett CL, Christie J, Ramsdell F et al. The immune dysregulation, polyendocrinopathy, enteropathy, X-linked syn-
3
4
5
6 7 8
9
10 11
12
13
14
15
drome (IPEX) is caused by mutations of FOXP3. Nat Genet 2001; 27:201. Brunkow ME, Jeffery EW, Hjerrild KA et al. Disruption of a new forkhead/winged-helix protein, scurfin, results in the fatal lymphoproliferative disorder of the scurfy mouse. Nat Genet 2001;27:6873. Fontenot JD, Gavin MA, Rudensky AY. Foxp3 programs the development and function of CD4 CD25 regulatory T cells. Nat Immunol 2003;4:3306. Hori S, Nomura T, Sakaguchi S. Control of regulatory T cell development by the transcription factor Foxp3. Science 2003;299: 105761. Khattri R, Cox T, Yasayko SA et al. An essential role for Scurfin in CD4 CD25 T regulatory cells. Nat Immunol 2003;4:33742. Mottet C, Uhlig HH, Powrie F. Cutting edge: cure of colitis by CD4 CD25 regulatory T cells. J Immunol 2003;170:393943. Sumida Y, Nakamura K, Kanayama K, et al. Preparation of functionally preserved CD4 CD25high regulatory T Cells from leukapheresis products from ulcerative colitis patients, applicable to regulatory T-Cell transfer therapy. Cytotherapy 2008;10:698710. Hoffmann P, Boeld TJ, Eder R et al. Isolation of CD4()CD25() regulatory T cells for clinical trials. Biol Blood Marrow Transplant 2006;12:26774. Hoffmann P, Edinger M. CD4 CD25 regulatory T cells and graft-versus-host disease. Semin Hematol 2006;43:629. Hoffmann P, Eder R, Boeld TJ et al. Only the CD45RA subpopulation of CD4 CD25high T cells gives rise to homogeneous regulatory T-cell lines upon in vitro expansion. Blood 2006;108:42607. Seddiki N, Santner-Nanan B, Martinson J et al. Expression of interleukin (IL)-2 and IL-7 receptors discriminates between human regulatory and activated T cells. J Exp Med 2006;203: 1693700. Liu W, Putnam AL, Xu-Yu Z et al. CD127 expression inversely correlates with FoxP3 and suppressive function of human CD4 T reg cells. J Exp Med 2006;203:170111. Sands BE, Sandborn WJ, Feagan B et al. A randomized, doubleblind, sham-controlled study of granulocyte/monocyte apheresis for active ulcerative colitis. Gastroenterology 2008;135:4009. Hanai H. Positions of selective leukocytapheresis in the medical therapy of ulcerative colitis. World J Gastroenterol 2006;12: 756877.
COMMENTARY
CD4 CD25 regulatory T cells approach the clinic M Edinger In 1995, Sakaguchi et al. [1] (CIT) described that CD4 T cells constitutively expressing the interleukin-2 receptor alpha chain (CD25) are potent suppressors of autoreactive T-cell responses. In their experimental system, they showed that the adoptive transfer of CD4 CD25 T cells inhibited the autoimmunity that is regularly observed in mice after early post-natal thymectomy. These experiments suggested that the suppressive T-cell population is thymus-derived and exported from the thymus only after birth and that there is no other peripheral tolerance mechanism compensating for the lack of these ‘regulatory’ T cells (Treg). A few years later, the transcription factor Foxp3 was identified as a master regulator for the development and peripheral function of those thymus-derived Treg cells, and it was shown that loss of function mutations in the foxp3 gene results in a lack of this natural Treg cell compartment [26]. In mice and humans, such mutations cause severe autoimmunity (scurfy and IPEX, respectively). Interestingly, boys with IPEX syndrome (immune dysregulation, polyendocrinopathy, eteropathy, X-linked) suffer from severe colitis and develop unusual immune responses, for example against nutritional antigens. In experimental models of colitis, Mottet et al. [7] showed that adoptively transferred Treg cells not only inhibit colitis induction but, more importantly, even cure established disease. Thus the adoptive transfer of natural Treg cells seems a promising strategy for the treatment of chronic colitis. In this issue of Cytotherapy, Sumida et al. [8] present methods for the enrichment of natural Treg cells from leukapheresis products of patients with ulcerative colitis (UC). For this purpose, they have adopted a strategy described previously for the isolation of Treg cells from healthy donors in allogeneic stem cell transplantation
[9,10]. This isolation protocol was developed for the prevention of graft-versus-host disease after allogeneic stem cell transplantation, where the co-transplantation of conventional and Treg cells at a 1:1 ratio is envisaged. Sumida et al. [8] have added a CD8-depletion step in addition to the CD19-depletion and repetitive CD25enrichment cycles to get rid of the 25% of CD8 T cells that otherwise contaminate a Treg-enriched cell product. Overall, they demonstrate an approximately 55% enrichment of natural Treg cells from leukapheresis products of colitis patients, based on the proportion of FOXP3 cells in the target cell fraction. However, the final cell product does not contain solely Treg cells but also recently activated CD4 T cells expressing CD25 and probably also some non-regulatory T cells that transiently express FOXP3 after stimulation. Whether such transient FOXP3 expression confers suppressive activity is a matter of debate and doubted by several investigators in the field. Nevertheless, exclusive surface markers for the unequivocal identification and separation of Treg cells from activated conventional T cells have not yet been identified and the protocol presented by Sumida et al. [8] is the best currently available approach for the enrichment of Treg cells under good manufacturing practice (GMP) conditions. Although additional depletion of CD127 cells or enrichment of naive Treg cells (CD45RA ) have been suggested to improve Treg cell purities [1113], such strategies remain to be evaluated at large scales with GMP-compatible reagents. Consequently, as long as we lack a technology that permits the generation of pure Treg cell products, we have to define the necessary degree of purity for each potential clinical application. Although leukocyte depletion from the peripheral blood of patients with UC is not yet a standard treatment world-wide [14], it
Correspondence to: PD Dr M. Edinger, Klinikum der Universita¨t Regensburg, Abt. fu¨r Ha¨matologie & Int. Onkologie, Franz-Josef-Strauss-Allee 11, 93053 Regensburg. E-mail: [email protected]. – 2008 ISCT
DOI: 10.1080/14653240802492705
656
M Edinger
is well accepted and frequently performed in Japan [15]. Thus phase I clinical trials combining leukapheresis for the depletion of leukocytes with the enrichment and reinfusion of autologous Treg cells seems feasible in this setting and no matter of ethical concern. Furthermore, shifting the balance from pro-inflammatory T cell responses to Treg-mediated suppression is expected to be beneficial in this patient group. Beyond safety and feasibility trials, however, the evaluation of efficacy will be a major challenge because large numbers of UC patients undergoing leukapheresis with and without additional Treg cell administrations will be required. In addition, long observation times will be necessary because of the remitting nature of the disease. Finally, the superiority of Treg cell transfers with regard to efficacy and safety will also have to be evaluated in comparison with standard pharmacologic immunosuppression (5-aminosalicylic acid, sulphasalazine, steroids, azathioprine or 6-mercaptopurine and cyclosporine) as well as recently introduced biologicals, such as the anti-tumor necrosis factor antibody infliximab. Thus, although it will be a cumbersome path to prove the efficacy of Treg cell therapies in this clinical situation, the re-infusion of freshly isolated or in vitroexpanded natural Treg cells is a promising strategy for UC patients and the first clinical trials are now in preparation. Declaration of interest: The author reports no conflicts of interest. The author alone is responsible for the content and writing of the paper.
References 1
2
Sakaguchi S, Sakaguchi N, Asano M et al. Immunologic selftolerance maintained by activated T cells expressing IL-2 receptor alpha-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J Immunol 1995;155:115164. Bennett CL, Christie J, Ramsdell F et al. The immune dysregulation, polyendocrinopathy, enteropathy, X-linked syn-
3
4
5
6 7 8
9
10 11
12
13
14
15
drome (IPEX) is caused by mutations of FOXP3. Nat Genet 2001; 27:201. Brunkow ME, Jeffery EW, Hjerrild KA et al. Disruption of a new forkhead/winged-helix protein, scurfin, results in the fatal lymphoproliferative disorder of the scurfy mouse. Nat Genet 2001;27:6873. Fontenot JD, Gavin MA, Rudensky AY. Foxp3 programs the development and function of CD4 CD25 regulatory T cells. Nat Immunol 2003;4:3306. Hori S, Nomura T, Sakaguchi S. Control of regulatory T cell development by the transcription factor Foxp3. Science 2003;299: 105761. Khattri R, Cox T, Yasayko SA et al. An essential role for Scurfin in CD4 CD25 T regulatory cells. Nat Immunol 2003;4:33742. Mottet C, Uhlig HH, Powrie F. Cutting edge: cure of colitis by CD4 CD25 regulatory T cells. J Immunol 2003;170:393943. Sumida Y, Nakamura K, Kanayama K, et al. Preparation of functionally preserved CD4 CD25high regulatory T Cells from leukapheresis products from ulcerative colitis patients, applicable to regulatory T-Cell transfer therapy. Cytotherapy 2008;10:698710. Hoffmann P, Boeld TJ, Eder R et al. Isolation of CD4()CD25() regulatory T cells for clinical trials. Biol Blood Marrow Transplant 2006;12:26774. Hoffmann P, Edinger M. CD4 CD25 regulatory T cells and graft-versus-host disease. Semin Hematol 2006;43:629. Hoffmann P, Eder R, Boeld TJ et al. Only the CD45RA subpopulation of CD4 CD25high T cells gives rise to homogeneous regulatory T-cell lines upon in vitro expansion. Blood 2006;108:42607. Seddiki N, Santner-Nanan B, Martinson J et al. Expression of interleukin (IL)-2 and IL-7 receptors discriminates between human regulatory and activated T cells. J Exp Med 2006;203: 1693700. Liu W, Putnam AL, Xu-Yu Z et al. CD127 expression inversely correlates with FoxP3 and suppressive function of human CD4 T reg cells. J Exp Med 2006;203:170111. Sands BE, Sandborn WJ, Feagan B et al. A randomized, doubleblind, sham-controlled study of granulocyte/monocyte apheresis for active ulcerative colitis. Gastroenterology 2008;135:4009. Hanai H. Positions of selective leukocytapheresis in the medical therapy of ulcerative colitis. World J Gastroenterol 2006;12: 756877.