Our Work

Research in our lab is broadly focused on understanding mechanisms regulating T cell activation. Our work has centered on altering the positive and negative co-stimulatory signals that are delivered in conjunction with signals from the T cell receptor during T cell activation. By manipulating positive co-stimulatory ligands, such as B7-1 or B7-2, or negative regulatory receptors, such as CTLA-4 or PD-1, we revealed new mechanisms to promote immunotolerance. In addition, we are studying an immunosuppressive population of T cells known as Tregs. Tregs are essential for preventing most forms of autoimmunity and we are developing strategies to utilize these cells to treat Type 1 Diabetes and other autoimmune diseases.

The breakdown of tolerance has been attributed to an imbalance of effector function and immune regulation, specifically defective regulation due to defects in the T regulatory cells (Treg) subset. Thus, multiple efforts have been forged to re-instate that balance in setting such as autoimmune disease and organ transplantation or disrupt it as a means to promote anti-tumor immunity. Recent investigations have focused on Treg instability in the autoimmune and cancer settings, and targeting of the FOXP3 pathway to selectively enhance Treg function. We have also focused attention on novel approaches to understanding FOXP3 activity and delivering specific signals to Tregs to promote Treg stability and function, including the use of novel IL-2 and anti-IL-2 approaches. Finally, we have initiated early clinical trials translating the insights gained from mouse studies to deliver Tregs and IL-2 therapeutically to promote rebalancing of effector and Treg function in autoimmunity and transplantation.

History of the Lab

In the mid-80s, we were the first lab to develop an anti-murine CD3 mAb (145-2C11) that has been used by hundreds of investigators to murine T cells. Moreover, this antibody has been used to address everything from thymic development, tolerance, T cell receptor signaling, and regulatory T cell development. We showed that the first dose toxicity caused by OKT3 was due to Fc receptor (FcR) binding and crosslinking. Thus, we developed a novel engineered anti-CD3 mAbs that eliminated FcR binding and as a consequence eliminated the cytokine storm induced by the activating anti-CD3. We demonstrated that the Fc receptor non-binding anti-CD3 mAbs have selective tolerogenic effects on Th1 T cells and promotes Tregs. 

a. Bluestone JA et al. Characterization of murine thymocytes possessing T3-associated T cell receptor structure. Nature 1987, 326(6108):82-84. (PMID: 3102972)
b. Leo O… and Bluestone JA. Identification of a monoclonal antibody specific for murine T3 polypeptide. Proc Natl Acad Sci 1987, 84:1374-78. (PMCID: PMC304432)
c. Smith JA…. and Bluestone JA. Nonmitogenic anti-CD3 monoclonal antibodies deliver a partial TCR cell signal and induce clonal anergy. J Exp Med 1997, 185:1413-22. (PMCID: PMC2196281)
d. Penaranda C… and Bluestone JA. Anti-CD3 therapy promotes tolerance by selectively depleting pathogenic cells while preserving regulatory T cells. J Immunol 2011, 187:2015-22. (PMCID: PMC3150219)

OKT3 was the first monoclonal antibody approved for clinical use. However, the toxicity was problematic for further drug development. We worked with J&J to develop the first non-mitogenic anti-CD3 mAbs to treat organ transplant rejection and autoimmunity.  We showed efficacy in kidney transplantation, psoriatic arthritis and type 1 diabetes, modeled the function in humanized models and are involved in Phase III trials.

a. Herold KC…. and Bluestone JA. Anti-CD3 Monoclonal Antibody in New Onset Type 1 Diabetes. N Eng J Med 2002, 346(22):1692-98. (PMID: 12037148)
b. Herold KC.... Bluestone JA, and the AbATE Study Team. Teplizumab (anti-CD3 mAb) treatment preserves C-peptide responses in patients with new onset type 1 diabetes: Metabolic and immunologic features at baseline that identify clinical responders. Diabetes 2013;62(11):3766-74. (PMCID: PMC3806618)
c. Lebastchi J….Bluestone JA, and Herold KC. Immune Therapy and b-Cell Death in Type 1 Diabetes. Diabetes 2013, 62(5):1676-80. (PMCID: PMC3636605)

In the late 1990s it became clear that T cells required a second signal to be fully activated to proliferate, produce IL-2 and differentiate into fully activated effector cells. We pursued the development of a potent CD28 antagonist, CTLA-4Ig. We were the first to show that this soluble receptor blocked CD28-mediated activation and importantly prevented graft rejection in an animal transplantation model. We performed a number of additional first and seminal studies showing that co-stimulatory blockade promotes T cells tolerance, identified a second CD28 ligand, B7-2. Our efforts were instrumental in the development of the first CTLA-4Ig drug, Abatacept.

a. Lenschow DJ…. and Bluestone, JA. Long-term survival of xenogeneic pancreatic islet grafts induced by CTLA4Ig. Science 1992, 257(5071):789-92. (PMID: 1323143)
b. Lenschow, DJ…. and Bluestone, JA. Expression and functional significance of a novel ligand for CTLA-4. Proc Natl Acad Sci 1993, 90(23):11054-8. (PMCID: PMC47920)
c. Levisetti, MG….and Bluestone, JA. Immunosuppressive effects of CTLA4Ig in a non-human primate model of allogeneic pancreatic islet transplantation. J Immunol 1997, 159(11):5187-91. (PMID: 9548454)
d. Szot, G…. and Bluestone, JA. Tolerance Induction and Reversal of Diabetes in Mice Transplanted with Human Embryonic Stem Cell-Derived Pancreatic Endoderm. Cell Stem Cell 2015, 16(2):148-57. (PMID: 25533131)

In the course of defining co-stimulation pathways, several investigators suggested that a CD28-related molecule, CTLA-4, also functioned as a co-stimulator. In a major paradigm shift, we showed that CTLA-4 was a negative regulator of T cell activation. This broke open a new field, termed checkpoint regulators, that led to the first approved CTLA-4, and subsequently PD-1 checkpoint blockers in humans, Ipilimumab and lambrolizumab, respectively. We went on to define the biochemical bases for CTLA-4 function and utilize this function in vivo to alter alloantigen recognition and Treg function.

a. Walunas, TL…. and Bluestone, JA. CTLA-4 can function as a negative regulator of T cell activation. Immunity 1994, 1(5):405-13. (PMID: 7882171)
b. Walunas, TL, and Bluestone, JA. CTLA-4 ligation blocks CD28-dependent T cell activation. J Exp Med 1996, 183(6):2541-50. (PMCID: PMC2192609)
c. Lee, K-M…. and Bluestone, JA. Molecular basis of T cell inactivation by CTLA-4. Science 1998, 282(5937):2263-6. (PMID: 9856951)
d. Bour-Jordan, H….and Bluestone, JA. CTLA-4 regulates the requirement for cytokine-induced signals in T(H)2 lineage commitment. Nat Immunol 2003, 4(2):182-8. (PMID: 12524538)

The 2000s brought to light the critical importance of immune regulation by a population of cells, regulatory T cells (Tregs), identified by a combination of cell surface markers and a master transcription factor, Foxp3. We were the first to demonstrate the critical importance of Tregs in a murine model of type 1 diabetes. We went on to demonstrate the impact of antigen-specific Tregs, the role of IL-2 deficiency in compromising Treg survival and function and Treg instability as a key driver of autoimmune susceptibility in mice and humans. These studies led the identification of key cell surface markers that enabled the isolation and expansion of these cells for the first human studies of Tregs in T1D and organ transplantation.

a. Salomon, B….and Bluestone, JA. B7/CD28 costimulation is essential for the homeostasis of the CD4+CD25+ immunoregulatory T Cells that control autoimmune diabetes. Immunity 2000, 12(4):431-40. (PMID: 10795741)
b. Tang, Q….and Bluestone, JA. Central role of a defective interleukin-2 production in the triggering islet autoimmune destruction. Immunity 2008, 28(5):687-97. (PMCID: PMC2394854)
c. Bailey-Bucktrout SL… and Bluestone JA.  Self-antigen-driven activation induces instability of regulatory T cells during an inflammatory autoimmune response. Immunity 2013, 39(5):949-62. (PMCID: PMC3912996)
d. DuPage M.… and Bluestone, JA. The chromatin-modifying enzyme Ezh2 is critical for the maintenance of regulatory T cell identity after activation. Immunity 2015, 42(2):227-38. (PMCID: PMC4347854)
e. Bluestone JA et al., Type 1 diabetes immunotherapy using polyclonal regulatory T cells. Sci. Transl. Med. 7:315ra189, 2015.