Human embryonic stem cells (hESCs) can undergo unlimited reproduction and retain the capability to differentiate into all cell types in the body. Therefore, as a renewable source of various cell types, hESCs hold great promise for human cell replacement therapy. Significant progress has been made in establishing the conditions to differentiate hESCs into cells of therapeutic value. However, a major obstacle to the clinical application of these promising hESC-based therapies is the immune-mediated rejection of hESC-derived cells by the recipient because these cells express antigens that differ from those of the recipient patients. While rejection of grafts expressing different antigens can be delayed for a period of time if the recipient’s immune system is persistently suppressed, most grafts are rejected eventually. In addition, persistent immune suppression also increases the risk for cancer and infection. Therefore, to develop hESC-based therapy, it is critical to develop effective approaches to induce immune tolerance to hESC-derived cells.
Extensive studies indicate that transplantation of donor's hematopoietic stem cells (HSCs) into the recipient prior to graft transplantation can induce tolerance of the recipient to the graft. Therefore, one promising approach that may be used to induce tolerance to hESC-derived cells is to transplant hESC-derived HSCs into the recipient. However, several bottlenecks hinder the evaluation of this promising approach. For example, due to the lack of specific markers for human HSCs, the differentiation and identification of hESCs into HSCs remains to be optimized. In addition, the animal model to study the human immune responses to hESC-derived cells remains to be established and validated.
To address these challenges, I have assembled a team of researchers with complementary expertise. We will genetically modified hESCs to improve our ability to identify and purify hESC-derived HSCs. In addition, we will develop humanized mouse models with functional human immune system to test whether transplantation of hESC-derived HSC and/or hESC-derived dendritic cells are sufficient to induce tolerance to hESC-derived cells such as cardiomyocytes. The ability to identify hESC-derived HSCs would facilitate the effort to provide a renewable cell resource for therapeutic bone marrow transplantation. In addition, our research could optimize the conditions to transplant hESC-derived HSCs in order to induce immune tolerance to differentiated cells derived from matched hESCs.
Limited therapeutic options are available for several devastating and costly diseases such as diabetes and heart diseases in California and our nation. In the case of diabetes, 1 of every 10 Californians (2.7 million) were afflicted with diabetes in 2007, costing the State $24.5 billion annually. Heart diseases remain the number one cause of death in California and nation, costing California even more than diabetes. Therefore, these diseases have devastating consequences on both those afflicted and on State/National healthcare costs. There remains an urgent and critical need for a cell-based cure of these diseases.
While significant progress has been made in the derivation of functional beta cells and cardiomyocytes from human ES cells, these allogenic cells will be rejected by the recipient upon transplantation unless the immune system of the recipient is persistently suppressed. However, immune suppression itself has severe consequences with significantly increased risk of cancer and infection. Therefore, it is critical to develop effective approaches to induce immune tolerance to hESC-derived cells. Our proposed research is aimed to develop approaches to induce immune tolerance to a wide range of hESC-derived cell types such as cardiomyocytes. The millions spent now on research is nominal when compared to the billions that will be saved in treatment costs and the improved quality of life for patients.
The Principal Investigator (PI) proposes to induce tolerance to human embryonic stem cell (hESC)-derived tissues by establishing hematopoietic chimerism using engraftable hematopoietic stem cells (HSCs) derived from the same hESC. In Aim 1, the PI proposes to generate a knock-in of marker genes into the locus for a hematopoietic transcription factor, and to use the modified hESC to optimize the conditions for HSC differentiation and identification. The derived HSC will then be validated by in vivo transfer studies. The goal of Aim 2 is to utilize a state of the art mouse model to determine whether hematopoietic chimerism with hESC-derived HSC and/or immature dendritic cells will induce tolerance to teratomas and cardiomyocytes derived from the same hESC.
Overall, the reviewers judged the proposed approaches to be innovative and creative, they appreciated the knock-in strategy, as it would facilitate both tracking and purification of hESC-derived HSCs. Reviewers acknowledged that the proposal has the potential for significant impact by advancing the clinical applicability of stem cell-based therapies and by improving derivation strategies for engraftable hESC-derived HSCs, an important bottleneck in the field.
Reviewers felt that in general, the rationale for the proposed tolerance approach and the chosen experimental strategy is strong. The aims are logically constructed and well supported by preliminary data, which include convincing evidence that the PI can achieve homologous recombination in hESC. However, reviewers had several concerns about this proposal. Aim 2 depends on the team’s ability to differentiate HSC from hESCs, but there is no preliminary data showing their ability to do that. That differentiation that has been challenging for the field even in mouse, so the lack of preliminary data is a serious concern. Also, reviewers saw no reason to believe that the HSCs obtained in the proposed studies would engraft any better than other hESC-derived populations, and further pointed out that achieving hematopoietic chimerism will require that the derived HSCs compete with fetal liver-derived HSC, which may also be problematic. A reviewer suggested that repopulating with the hESC-derived HSCs alone or with less competitive populations should be included as another experimental group in the plan. Reviewers expressed concern that the PI did not address the potential consequences of the marker gene knock-in on the expression or function of the targeted key hematopoietic transcription factor. They further pointed out that additional markers may have to be included in the HSC identification strategy, since the targeted gene is expressed in cells other than HSC. Alternate plans are not well developed.
The reviewers praised the PI’s expertise and outstanding track record. The PI is an expert immunologist, is experienced in homologous recombination methods, and allocates 10% commitment to this project. The research team is strong, and the collaborating PIs provide complementary expertise relevant to the proposed aims, but reviewers were concerned by the lack of expertise in obtaining engraftable HSC from hESC, a critical part of the proposal.
In summary, this application is focused on the use of hESC-derived HSCs to induce tolerance to hESC-derived grafts. The proposal is innovative and addresses a significant problem, and the PI and the team have a strong track record, but the reviewers felt that the problems with the research plan and its feasibility reduce the likelihood that the proposal would have any real impact.