Journal of Autoimmune Disorders

Editorial

The immune system is operated on the basis of checks and balances. Inflammatory Th1 and Th17 cells, in the process of fighting off pathogens, may hyper-react and cause damages to tissues. Regulatory T cells (Tregs), on the other hand, function to suppress exuberant immune reactivity and promote peripheral tolerance [1]. Tregs characteristically express high and stable level of the IL-2 receptor alpha chain (IL-2Rα, CD25^high) on the cell surface and the transcription factor fork-head box protein 3 (Foxp3) [2]. Disease ensues when there is an imbalance between the inflammatory and the regulatory T cells. It has been shown that patients with multiple sclerosis (MS), type 1 diabetes (IDDM), rheumatoid arthritis (RA) and other autoimmune diseases are deficient in Tregs frequency and function [3-5]. As such, replenishing the stock of Tregs in these patients seems to be a reasonable strategy. One approach is to design protocols to induce the body's own endogenous Treg pool. Given our current limited knowledge of the mechanisms of Treg functions, not much success of this approach has been reported. Another approach which has gained popularity in recent years is to provide the body with exogenously generated and expanded Tregs. Two types of Tregs are commonly used in adoptive Treg therapy. Natural Tregs (nTregs) are derived and develop in the thymus and use a diverse T cell receptor repertoire [6]. Induced Tregs (iTregs) populate the periphery and are induced by TGF-β to express Foxp3 after encountering antigen [7]. Initial investigations of the concept of adoptive Treg therapy in animal use natural Tregs (nTregs) and were mostly studied in the mouse model of bone marrow transplantation. Freshly isolated nTregs or ex vivo expanded donor Tregs transferred into recipients were found to ameliorate graft versus host disease (GVHD) and facilitate engraftment [8,9]. Others had also demonstrated utilities in preventing the rejection of pancreatic islets [10] and organ allografts [11] and the development of autoimmune diseases.

Based on these animal studies, several phase I and phase I/II clinical trials on adoptive Treg therapy in stem cell transplantation were launched [12-14]. These studies were valuable in establishing certain parameters in experimental design and in pointing out the limitations and challenges of the trade. Among the prominent challenges is the inability to collect sufficient number of Tregs for adoptive therapy as nTregs only constitute 3-5% of the peripheral circulating CD4+ T cells [15]. To remedy this, in vitro expansion methods have been introduced [16]. These methods depend on the use of high concentration of IL-2 and other pharmaceuticals such as rapamycin to expand Tregs. These approaches, by themselves, have inherent risks in that their effects on the long term stability of the expanded Tregs in vivo are not known. It has been shown that unstable Tregs could be easily converted into effector T cells in vivo [17]. In addition, depending on the separation methods, Tregs may be contaminated with small number of effector cells, which in the presence of the expansion pharmaceuticals may also proliferate and increase in numbers, eventually causing complications in vivo. Another prominent challenge is the lack of antigen specificities of the nTregs. Not only that the frequency of Tregs specific for given antigen is low, these polyclonal Tregs might cause cumulative global suppression of the host's immune system including protective immunity against infection and tumor growth. As a corollary, the question is whether using antigen-specific Tregs for adoptive Treg therapy is a better choice than nTregs. In the GVHD model, use of alloantigenspecific Tregs did not seem to improve the results too much better than nTregs [18]. However, in the IDDM NOD model, in vitro expanded antigen-specific Tregs suppressed prediabetic and diabetic mice significantly better than polyclonal nTregs [19]. Developing novel protocols to generate large quantities of antigen-specific Tregs for adoptive Treg therapy is now an active area of research activities.

In view of the limitations and challenges of adoptive Treg therapy for treatment of immunological diseases, scientists are trending towards resolving some of these issues through induced pluripotent stem cells (iPSC). In 2006 and 2007, Takahashi and Yamanaka [20,21] provided evidence that adult mouse and human fibroblasts could be reprogrammed to become pluripotent stem cells by transducing them with retrovirus carrying certain stem cell transcription factors. These so-called iPSCs possessed all the properties of embryonic stem cells. Yamanaka and co-workers defined minimally 4 stem cell factors, Oct3/4, Sox2, Klf4 and c-Myc, for successful cell reprogramming. This discovery revolutionized the field of stem cell research because stem cells can be generated without the ethical difficulties regarding the use of human embryos.

Because Takahashi and Yamanaka [20] initially used retroviral vector-mediated delivery of the 4 reprogramming factors, there were concerns that the integrating retroviral material might cause insertional mutagenesis in the host genome. Investigators began to look for safer non-integrating vectors or non-vector delivery systems [22-24]. Zhou et al. [25] first fused the 4 reprogramming protein with a poly-arginine protein transduction domain and generated iPSCs from mouse embryonic fibroblast (MEF) cells. Recently, Hou et al. [26] took a totally different approach and used a chemical reprogramming strategy of combining 7 small-molecule chemical that also generate iPSCs from MEF cells. These non-vector designs, however, often suffer from low reprogramming efficiency. An Israeli group of scientists discovered that by depleting a deacetylation repressor Mbd3, they were able to improve the efficiency of Oct4, Sox2, Klf4 and Nanog (OSKN) transduction to100% [27].

In conclusion, the iPSC technology is moving very fast and holds the future for generating therapeutic reagents that serve to modulate a wide spectrum of diseases.

References

1. Takahashi T, Tagami T, Yamazaki S, Uede T, Shimizu J, et al. (2000) Immunologic self-tolerance maintained by CD25+CD4+ regulatory T cells constitutively expressing cytotoxic T lymphocyte associated antigen 4. J Exp Med 192: 303-310.
2. Sakaguchi S, Sakaguchi N, Asano M, Itoh M, Toda M (1995) Immunologic self-tolerance 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 155: 1151-1164.
3. Ehrenstein MR, Evans JG, Singh A, Moore S, Warnes G, et al. (2004) Compromised function of regulatory T cells in rheumatoid arthritis and reversal by anti-TNF therapy. J Exp Med 200: 277-285.
4. Lindley S, Dayan CM, Bishop A, Roep BO, Peakman M, et al. (2005) Defective suppressor function in CD4+CD25+ T-cells from patients with type 1 diabetes.Diabetes 54: 92-99.
5. Viglietta V, Baecher-Allan C, Weiner HL, Hafler DA (2004) Loss of functional suppression by CD4+CD25+ regulatory T cells in patients with multiple sclerosis. J Exp Med 199: 971-979.
6. Bluestone JA, Abbas AK (2003)  Natural versus adaptive regulatory T cells. Nature Rev Immunol 3: 253-257.
7. Schmitt EG, Williams CB (2013)Generation and function of induced regulatoryT cells. Frontiersin 4:152.
8. Hanash AM, Levy RB (2005) Donor CD4+CD25+ T cells promote engraftment and tolerance following MHC-mismatched hematopoietic cell transplantation.  Blood 105:1828-1836.
9. Cohen JL, Trenado A, Vasey D, Klatzmann D, Salomon BL (2002) CD4+CD25+ immuno-regulatory T cells: new therapeutics for graft-versus-host disease. J Exp Med 196:401-406.
10. Sanchez-Fueyo A, Weber M, Domenig C, Strom T, Zheng XX (2002) Tracking the immuno-regulatory mechanisms active during allograft tolerance. J Immunol 168: 2274-2281.
11. Joffre O, Santolaria T, Calise D, Al Saati T, Hudrisier D, et al. (2008)  Prevention of acute and chronic allograft rejection with CD4+CD25+Foxp3+ regulatory T-lymphocytes.  Nat Med 14:88-92.
12. Trzonkowski P, Bieniaszewska M, Juscinska J (2009) First-in-man clinical results of the treatment of patients with graft versus host disease with human ex vivo expanded CD4+CD25+CD127- T regulatory cells. ClinImmunol 133: 22-26.
13. Brunstein CG, Miller JS, Cao Q(2011) Infusion of ex vivo expanded T regulatory cells in adults transplanted with umbilical cord blood: safety profile and detection kinetics. Blood 117: 1061-1070.
14. DiIanni M, Falzetti F, Carotti A (2011) Tregs prevent GVHD and promote immune reconstitution in HLA-haploidentical transplantation.Blood 117: 3921-3928.
15. Safinia N, Sagoo P, Lechler R, Lombardi G (2010) Adoptive regulatory T cell therapy: challenges in clinical transplantation. CurrOpin Organ Transplant 15:427-434.
16. Hoffmann P, Eder R, Kunz-Schughart LA, Andreesen R, Edinger M (2004) Large-scale in vitro expansion of polyclonal human CD4+CD25high regulatory T cells. Blood 104: 895-903.
17. Zhou X, Bailey-Bucktrout SL, Jeker LT, Penaranda C, Martínez-Llordella M, et al.(2009)  Instability of the transcription factor Foxp3 leads to the generation of pathogenic memoryT cells in vivo. Nat Immunol 10, 1000-1007.
18. Trenado A, Charlotte F, Fisson S et al. (2003) Recipient-type specific CD4+CD25+regulatory T cells favor immune reconstitution and control graft-versus-host disease while maintaining graft-versus-leukemia. J Clin Invest 112: 1688-1696.
19. Tang Q, Henriksen KJ, Bi M (2004) In Vitro–expanded Antigen-specific Regulatory T Cells Suppress Autoimmune Diabetes.J Exp Med 199: 1455-1465.
20. Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126: 663-676.
21. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka MT, et al. (2007)Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors.  Cell 131: 861–872,
22. Stadtfeld M, Nagaya M, Utikal J, Weir G, Hochedlinger K (2008) Induced pluripotent stem cells generated without viral integration. Science 322: 945-949.
23. Okita K, Nakagawa M, Hyenjong H, Ichisaka T, Yamanaka S (2008) Generation of mouse induced pluripotent stem cells without viral vectors. Science 322: 949-953.
24. Yu J, Hu K, Smuga-Otto K, Tian S, Stewart R, et al. (2009) Human induced pluripotent stem cells free of vector and transgene sequences. Science 324: 797-801.
25. Zhou H, Wu S, Joo JY, Zhu S, Han DW, et al. (2009) Generation of induced pluripotent stem cells using recombinant proteins. Cell Stem Cell 4: 381-384.
26. Hou P, Li Y, Zhang X, Liu C, Guan J, et al. (2013) Pluripotent stem cells induced from mouse somatic cells by small-molecule compounds. Science 341: 651-654.
27. Rais Y, Zviran A, Geula S, Gafni O, Chomsky E, et al. (2013) Deterministic direct reprogramming of somatic cells to pluripotency.  Nature 502: 65-70.