Blood and immune cells originate and mature in the bone marrow. Bone marrow cells are mixtures of blood cells at different stages of development, and include rare populations of blood-forming stem cells. These stem cells are the only cells capable of generating the blood system for the life of an individual. Bone marrow transplants (BMT) have been performed > 50 years, to replace a diseased patient’s blood system with that of a donor. Unfortunately, BMT have associated dangers which make the procedure high risk. Major risks include a syndrome called graft-versus-host disease (GvHD) which results when the donor’s mature blood cells attack the organs of the host, and toxicity from the treatments (radiation and chemotherapy) required to permit the donor cells to take in the recipient. These risk factor limit the use of BMT to only immediate life-threatening diseases.
If made safer, BMT could cure many other debilitating diseases. In addition to being curative of blood cancers and non-malignant blood diseases (such as sickle-cell anemia), these transplants can cure autoimmune diseases, such as juvenile (type I) diabetes and multiple sclerosis. In addition, simultaneous BMT with organ transplants induces “tolerance” to the new organ, meaning the recipient will not reject the graft because the new blood system provides continuous proteins to re-train the recipient immune system not to attack it. This establishment of tolerance eliminates the need for drugs that suppress the immune system.
In efforts to make BMT safer, our research has focused on isolating the blood stem cells away from the other bone marrow cells because transplants of pure stem cells do not cause GvHD. We developed the methods to purify the blood stem cells from mouse and human blood forming sources and showed in mice that transplants of blood stem cells can cure autoimmune disease and induce tolerance to solid organ transplants. However, this technology has not been tested in human clinical trials because safer methods must be developed that permit the stem cells to engraft in recipients.
Our studies in mice show that we can replace the toxic drugs and radiation used to prepare recipients for BMT with non-toxic proteins that target the cells responsible for rejection of blood stem cells. The goal of this study is to translate this technology from mice to patient clinical trials. If successful, the studies will open the door to the use of blood stem cell transplants to the many thousands of patients who could benefit from this approach. The science behind achieving blood stem cell engraftment by the methods we propose look toward the future when blood stem cells and other tissues will be developed from pluripotent stem cells (ES, NT and iPS). We envision that the blood stem cells will induce tolerance to tissues derived from the same pluripotent stem cell line, in the same way that adult blood stem cells induce tolerance to organs from the same living donor.
Statement of Benefit to California:
The science and the preclinical pathway to induce human immune tolerance in patients with degenerative diseases so that new blood and tissue stem cells can regenerate their lost tissues: For stem cell biology to launch the era of regenerative medicine, stem cells capable of robust and specific regeneration upon transplantation must be found, and methods for safe patient administration must be developed. In the cases where cell donation cannot come from the host, immune responses will reject the donor stem cells. Successful transplants of blood-forming stem cells (HSC) leads to elimination immune cells that reject organ grafts from donors. While bone marrow or cord blood transplants contain immune cells called T cells that will attack the host in a potentially lethal graft against host immune reaction, purified HSC do not do this. Pluripotent stem cells (ES, NT, iPS) can make all cell types in the body and provide a shortcut to find tissue and organ stem cells. Just as co-transplants of adult HSC prevent rejection of organs from the same donor, co-transplants of HSC derived from pluripotent cells should protect tissues derived from the same pluripotent line. Attack by a patient's blood system against one’s own organs cause the syndromes of autoimmune disease including juvenile diabetes, multiple sclerosis, and lupus. Transplanted HSC from donor mice genetically resistant to these diseases end the autoimmune attack permanently. We have in mice, substituted minimally toxic antibodies for toxic chemoradiotherapy to prepare the host for HSC transplants. Now it is time to take these advances to humans, with human immune cell and HSC-targeting antibodies.
Long-term potential benefits to the state of California and its residents: The justification for Proposition 71 was to establish in California centers of research not funded adequately in the areas of stem cell biology and regenerative medicine. This research, if successful, is the platform for the application of stem cell biology to regenerative medicine. The costs for long-term immune suppression to patients who receive organ transplants are enormous, both in terms of quality of life, even survival, and healthcare resources. Add to that the lifetime costs of insulin to treat juvenile diabetes, with the inevitable premature diseases of compromised blood vessels and organs, and the shortened lifespan of patients. Add to that the costs to lives and the healthcare system of lupus, of multiple sclerosis, of other autoimmune diseases like juvenile and adult rheumatoid arthritis and scleroderma, and of muscular dystrophy, to mention a few, and the value to Californians and people everywhere is obvious. If our studies are successful, and if the clinical trials were first done in California, our citizens will have the first chance at successful treatment. Further, if these studies are successful - new antibodies, if produced by CIRM funds, will generate royalties which eventually will return to the state.
Year 1The successful transplantation of blood forming stem cells from one person to another can alter the recipient immune system in profound ways. The transplanted blood forming cells can condition the recipient to accept organs from the original stem cell donor without the need for drugs to suppress their immune system; and such transplantations can be curative of autoimmune diseases such as childhood diabetes and multiple sclerosis. Modification of the immune system in these ways is called immune tolerance induction.
Unfortunately, the current practice of blood stem cell transplantation is associated with serious risks, including risk of death in 10-20% of recipients. It has been a long-standing goal of investigators in this field to make transplantations safer so that patients that must undergo this procedure have better outcomes, and so that patients who need an organ graft or that suffer from an autoimmune disorder can be effectively treated by this powerful form of cellular therapy. The major objectives of our proposal are to achieve this goal by developing methods to prepare patients to accept blood forming stem cell grafts with reagents that specifically target cell populations in recipients that constitute the barriers to engraftment, and to transplant only purified blood forming stem cells thereby avoiding the potentially lethal complication call graft-vs-host disease.
The proposal has four Specific Aims. Aims 1 and 2 focus on development of biologic agents that specifically target recipient barrier cells. Aims 3 and 4 propose to test the reagents and approaches developed in the first two aims in mouse models to induce tolerance to co-transplanted tissues and to cure animals with Type 1 diabetes mellitus or multiple sclerosis. These aims have not changed in this reporting period.
One parameter of success in this project is the development of one or more biologic reagents that can replace toxic radiation and chemotherapy that can be used in human clinical trials by the end of the third year of funding (Aim 2). In this regard, significant progress has been made in the last year. A reagent critical to the success of donor blood forming stem cell engraftment is one that targets and eliminates the stem cells that already reside in the recipients. Recipient blood stem cells block the ability of donor stem cells to take. In our prior mouse studies we determined that a protein (antibody) that specifically targets a molecule on the surface of blood forming stem cells called CD117 is capable of eliminating recipient blood stem cells thus opening up special niches and allowing donor stem cells to engraft. This antibody was highly effective in permitting engraftment of purified donor blood stem cells in mice that lack a functional immune system. In this application we proposed to develop and test reagents that could target and eliminate human blood forming stem cells by targeting human CD117. This year we have identified and tested such an antibody which is manufactured by a third party. This anti-CD117 antibody has been evaluated in early clinical trials for an indication separate from our proposed use and appears to be non-toxic. In mice that we generated to house a human blood system, the antibody was capable eliminating the human blood forming stem cells. We plan to pursue the use of this reagent in a clinical trial as a non-toxic way to prepare children with a disease called severe combined immunodeficiency (SCID) for transplantation. Without a transplant children with SCID will die. The use of the anti-CD117 antibody and transplantation of purified blood forming stem cells has the potential to significantly reduce the complications of such transplants and improve the outcomes for these patients. The trial will be the first step to using this form of targeted therapy and serve as a pioneering study for all indications for which a blood forming stem cell transplant is needed, including the induction of immune tolerance.
Year 2The transplantation of blood forming stem cells from one individual to another can alter the recipient immune system in profound ways. Transplanted blood forming cells can condition the recipient to accept organs from the original stem cell donor without the need for drugs to suppress their immune system. Such transplantations can also be curative of autoimmune diseases such as childhood diabetes and multiple sclerosis. Modification of the immune system in these ways is called immune tolerance induction.
The major goal of this project is to enable the use of blood forming stem cell transplantation for the purpose of immune tolerance induction without unwanted side effects. The current practice of blood stem cell transplantation is associated with serious risks, including risk of death in 10-20% of recipients due to complications of transplant conditioning and graft-versus-host disease. We aim to abolish or reduce the risks of these transplantations so that this curative form of stem cell therapy can safely treat patients who need an organ graft or who suffer from an autoimmune disorder. To achieve our goals, we proposed the development of methods to prepare patients to accept blood forming stem cell grafts with reagents that specifically target recipient cell populations that constitute the barriers to engraftment, and to transplant only purified blood forming stem cells, thereby avoiding graft-versus-host disease.
The proposal has four Specific Aims. Aims 1 and 2 focus on development of biologic agents that specifically target recipient barrier cells. Aims 3 and 4 propose testing the reagents and approaches developed in the first two aims in mouse models to induce tolerance to co-transplanted tissues and to cure animals with muscular dystrophy, Type 1 diabetes mellitus and multiple sclerosis. These aims have not changed in this reporting period.
In this reporting period, significant progress has been made in the first three aims. In prior years we identified a biologic reagent that has the potential to replace toxic radiation and chemotherapy. Radiation and chemotherapy are used in transplantation to eliminate the blood forming stem cells of recipients because recipient stem cells block the ability of donor cells to take. The novel reagent we have studied is a protein, called a monoclonal antibody, which differs from radiation and chemotherapy because it specifically targets and eliminates recipient blood stem cells. This antibody reagent recognizes a molecule on the surface of blood stem cells called CD117. In years 1 and 2 we began testing of an anti-human CD117 (anti-hCD117) antibody in mice. Mice were engrafted with human blood cells and we showed that this antibody safely and specifically eliminated the human blood forming cells. These studies were proof-of-concept that the antibody is appropriate for use in human clinical trials.
This last year we were awarded a CIRM Disease Team grant to move the testing of this anti-hCD117 from the experimental phase in mice to a clinical trial for the treatment of children with a disease call severe combined immunodeficiency (SCID), also known as the “bubble boy” disease. Children with SCID are missing certain types of white blood cells (lymphocytes) so they cannot defend themselves from infections. Without a transplant, children with SCID will die. The use of the anti-CD117 antibody and transplantation of purified blood forming stem cells has the potential to significantly reduce the complications of such transplants and improve the outcomes for these patients. The use of the anti-CD117 antibody and transplantation of purified blood forming stem cells has the potential to significantly reduce the complications of such transplants and improve the outcomes for these patients. The trial will be the first step to using this form of targeted therapy and serve as a pioneering study for all indications for which a blood forming stem cell transplant is needed, including the induction of immune tolerance.
In the last year we have moved forward with the purification of skeletal muscle stem cells based upon labeling and sorting of primitive muscle cells that express an array of molecules on the cell surface. We have also transplanted a special strain of mice (mdx) that are a model for muscular dystrophy with blood forming stem cells from normal mouse donors. In the coming year we will perform simultaneous transplants of blood forming stem cells and skeletal muscle stem cells from normal donor mice into the mdx mice. We will determine if the blood stem cells permit the long-term survival of the muscle stem cells in recipients transplanted across histocompatibility barriers. Our ultimate goal is to achieve long-term recovery of muscle cell function in the recipients of these co-transplantations.