The goal of our research is to develop efficient methods for making a particular class of immune-system cells known as regulatory T cells (Tregs). Tregs have the potential to be useful in a wide variety of clinical situations. For instance, they could be used to control the harmful immune responses seen in patients with autoimmune diseases such as childhood (Type I) diabetes, rheumatoid arthritis, multiple sclerosis and inflammatory bowel disease; and to suppress rejection of transplanted organs in patients given heart, liver or kidney transplants. These patients are normally treated with toxic immunosuppressive drugs to prevent transplant rejection, but nevertheless tend to lose the organs and need to get on a long waiting list all over again. Treating them with Tregs might preserve the transplant, possibly indefinitely, and is expected to be much less toxic because it would decrease or eliminate the need for the immunosuppressive drugs.
Bone marrow transplants are a special case. Stem cells present in the bone marrow give rise to all types of blood cells, including red blood cells which carry oxygen, platelets which are necessary for blood clotting so that one does not bleed to death from a minor injury, and a large variety of cells which fight off bacterial and viral infection. Aging patients tend to develop bone marrow failure spontaneously, and patients who have been given chemotherapy for cancer also almost invariably lose bone marrow function. When these patients are treated with bone marrow from a different donor, the bone marrow (graft) itself can start attacking the patient (host), in a life-threatening scenario known as graft-versus-host disease. Again, Tregs can help to prevent this disease, thus realizing the promise of transplantation with bone marrow stem cells.
We plan to develop efficient ways to make Tregs from different types of stem cells. For patients receiving transplanted organs, we hope to take their own normal T cells and turn them into Tregs. For patients suffering from autoimmune diseases, it might be more useful to make Tregs artificially from their bone marrow stem cells, whereas for cancer patients who have been given chemotherapy followed by a bone marrow transplant from a different person, it might be possible to make Tregs from the same bone marrow cells that the patient receives, in the hope that these Tregs can suppress graft-versus-host disease. And finally, there are clinical situations in which it might be useful to use the very new technique of induced pluripotent stem cells to make stem cells from a patient’s skin, then turn those stem cells into Tregs. We believe that all these approaches are feasible in principle, given recent discoveries from our own and other labs. Although our research will be done mostly in animals (mice), we believe that it will be possible to translate it quickly into humans, and that if successful, it will address a pressing clinical need.
In this application we propose to develop efficient methods for making cells of the immune system known as regulatory T cells (Tregs). As described in the proposal, Tregs have the potential to be outstandingly useful to many different types of patients: people receiving solid organ transplants, bone marrow transplants and stem cell transplants, as well as people with autoimmune diseases of various kinds. Our research is therefore aimed at improving the health of the citizens of the State of California and the United States. Any clinical trials that result from the research would be performed in hospitals in California and would be of benefit to patients in the state.
Aside from the purely medical importance of our research, however, our project will benefit the State of California from an economic point of view as well. The research institute and the core facilities where the research is to be performed are located in the State of California and will be led by a California-based research team, and all technology licensing will benefit the state directly. We intend to hire and train at least one research technician, two postdoctoral fellows and one Ph.D. graduate student who will all live and work in the State of California and by buying goods and services, will contribute to the economic health of the state. California-based businesses and vendors will be used as suppliers of all needed equipment, services and supplies. Any meetings that involve external speakers and collaborators will be held in California, even though other locations could be chosen. Thus there would be substantial long term employment in the State of California if this research were funded.
This proposal focuses on the generation of stable T regulatory (Treg) cells from CD4+ T cells and from pluripotent and multipotent stem cell (SC) populations. The applicants have identified a set of transcription factors (TFs) involved in DNA demethylation and propose that these TFs play a role in stabilizing expression of a specific gene essential to the maintenance of the Treg phenotype. Treg cells have strong immunomodulatory properties that could be used clinically to facilitate graft acceptance and induction of donor-specific tolerance. However, Treg cells generated in vitro through the addition of TGF-beta do not maintain a stable Treg cell phenotype and lose expression of the critical gene and Treg cell functionality in vivo. The applicants propose 3 aims designed to first determine the importance of these TFs in stabilizing expression of Treg gene and function of Treg cells generated in culture; then to develop efficient methods to reprogram murine multipotent and pluripotent SCs into Treg cells; and, finally, to use the generated Treg cells for therapeutic application in mouse models of transplant rejection and autoimmune disease.
Reviewers unanimously termed this project "high risk/high gain". Identifying new ways to increase the stability of the Treg cell phenotype and function is important not only for the clinical applicability of SC-derived therapies and achieving long-term tolerance, but also in utilizing the cell population for treatment of autoimmune disease. If successful, this proposal would impact all of these areas and was thus considered "high gain". Reviewers thought that the focus on one particular TF was both innovative and creative and incorporated state-of-the-art molecular and cellular biology.
Despite this high impact and a scientific rationale that was for the most part well justified and strong, there were several aspects of the research plan that reviewers considered "high risk". Reviewers were particularly concerned about the Treg differentiation portion of the application. They commented that expression of these TFs might not be sufficient to induce commitment and adequate differentiation of multipotent stem cell precursors to the Treg lineage, an issue that was not addressed by alternative plans. Reviewers also agreed that the study plan provided limited details on how the specific TFs that impacted Treg cell fate would be identified, which cellular subsets would be compared, and what type of output would be used. Finally, reviewers commented that the applicants do not discuss the issue of antigen-specificity of their in vitro generated Tregs. Some reviewers thought that generation of antigen-specific Tregs might be necessary for the in vivo testing of Treg function proposed in Aim 3, and one reviewer noted that antigen-specificity is an essential parameter in achieving sufficient and long-lasting tolerance to the targeted auto-antigen(s) in autoimmunity and/or donor-specific allo-antigen(s) in tissue transplantation. Other reviewers, however, thought that polyclonally-activated cells, which contain a small number of auto or crossreactive Tregs, might be sufficient for this purpose. Despite these feasibility concerns, the reviewers agreed that the proposal was carefully designed to give meaningful results and that the timelines were reasonable.
The reviewers considered the principal investigator (PI) to be outstanding. The PI has knowledge in immunology, transcription factor regulation, and T cells in development that would be highly beneficial in the embryonic stem cell and induced pluripotent stem cell field. Reviewers did note that the team lacked adequate SC expertise, particularly in the area of cellular reprogramming and thought the project would benefit from the addition of someone with this expertise. However, the PI’s track record as an innovative and successful investigator, combined with the excellent environment, is likely to bring success.
Overall the reviewers were highly enthusiastic about this "high risk/high gain" proposal and thought their feasibility concerns could be successfully addressed by this outstanding PI.