Funding opportunities

Investigating the mechanisms of oocyte-based and factor-based reprogramming

Funding Type: 
Basic Biology IV
Grant Number: 
RB4-06192
Funds requested: 
$1 336 382
Funding Recommendations: 
Not recommended
Grant approved: 
No
Public Abstract: 
The use of induced Pluripotent Stem Cells (iPSCs) genetically identical to patients has great promise for treating many diseases. However, iPSCs generated by current techniques, even iPSC-derivatives genetically identical to the recipient, express proteins that cause them to be rejected, presenting a major problem for their use for regenerative medical applications. Prior research demonstrated eggs (called oocytes) have the potential to change (reprogram) skin cells in a way that may eliminate this problem. However, use of human oocytes is limited by practical, ethical and legal considerations. Identification of the factors in human oocytes that provide this optimized reprogramming will improve the efficacy of human iPSC-based patient-specific cellular therapies. By identifying, cloning and using these factors to generate fully reprogrammed iPSCs, we will eliminate the requirement for donor human oocytes and permit reprogramming that will eliminate the immune rejection problem. The ultimate objective for this proposal is to develop a method that will fully reprogram human skin cells to an Embryonic Stem Cell (ESC)-like state that will not express proteins that cause iPSCs and their derivatives to be rejected by the patient’s immune system. If successful, the ability to transplant patient-specific reprogrammed skin cells, without immune rejection problems, will significantly advance the use of iPSCs for a wide variety of regenerative medical applications.
Statement of Benefit to California: 
Diseases (such as Parkinson’s, diabetes and heart disease), injuries (such as spinal cord injury) and age-related tissue degeneration afflict many people both inside and outside the State of California, resulting in physical, emotional and financial burdens on individuals and on society as a whole. The generation of induced Pluripotent Stem Cells (iPSCs) that are genetically identical to patients has significant promise for future patient-specific cellular therapies for these diseases, injuries and aliments. Effective iPSC therapies have the potential to cure or alleviate the symptoms for literally millions of individuals in the future. However, a major unsolved problem for the use of iPSCs for patient-specific cellular therapies is the expression of immunogenicity genes by the iPSCs differentiated derivatives, leading to their immune rejection, even when using genetically identical cells. This immunogenicity problem fundamentally limits the use of iPSC-based therapies. This proposed research seeks to address this immunogenicity problem by identifying the key factors in human eggs (oocytes) that enable the generation of iPSCs that are not rejected following autologous transplantation. Finally, solving the immunogenicity issue would also provide a significant boost to the stem cell biotechnology industry in the State of California, providing jobs and an additional source of revenue through potential patents and licensing agreements.
Review Summary: 
A study in the mouse system has shown that induced pluripotent stem (iPS) cells and their derivatives cause an immune reaction when transplanted into a genetically identical host. Since embryonic stem (ES) cells did not trigger this immune response, the applicant seeks to overcome this problem by identifying factors that will enable the generation of iPS cells that more closely resemble ES cells than current iPS cells do. To achieve this goal, the applicant proposes 1) to compare the immunogenic profile of ES cells that were generated following somatic cell nuclear transfer into oocytes to that of iPS cells, utilizing an animal model, 2) to identify a combination of genes, shown to be highly expressed in oocytes but not in fibroblasts, that can reprogram fibroblasts to pluripotency, and then 3) to determine whether the oocyte factor reprogrammed human and animal model iPS cells, and their derivatives, show evidence of being less immunogenic than iPS cells derived using conventional reprogramming factors. Significance and Innovation - The overall rationale for this proposal is not well conceived, as it is based almost exclusively on a single report that iPS cells are rejected in a genetically identical host. - The rationale for the choice and number of oocyte-expressed factors to be tested is not convincing and several of the proposed factors have been previously tested and demonstrated to be involved in pluripotency, raising doubt whether new factors that improve reprogramming outcomes will be identified. - Reviewers were not persuaded by the hypothesis that iPS cells produced by the proposed method would be non-immunogenic. - While the oocyte biology-based approach for iPS cell generation is innovative, it is also flawed. It is built on the assumption that reprogramming factors acting in the oocyte can induce an ES cell-like (pluripotent) state, whereas in vivo oocytes reprogram the genome to totipotency. - The proposed epigenetic analysis to examine reprogramming fidelity is innovative. Feasibility and Experimental Design - Although the pursuit of questions relating to immunogenicity of human iPS cell lines is commendable, the proposed method to evaluate the various cell lines is insufficient; more sophisticated immunological analyses are needed for this project to successfully advance current understanding of immunogenicity of iPS cells. - The number of cell lines to be analyzed is not sufficiently high to yield conclusive data. - The preliminary data provide evidence that the applicant is competent to work with ES cells, iPS cells, and the chosen animal model but much additional data is needed to support the project rationale. - The proposed approaches for the generation and analysis of iPS cells are feasible in principle. Principal Investigator (PI) and Research Team - The PI is a new investigator who is well positioned to do the proposed research. - The collaborators are a strength comprising an excellent team to generate vectors and iPS cells, and to perform epigenetic studies. Responsiveness to the RFA - The application is responsive to the RFA; it is focused on basic aspects of stem cell biology. - Although most of the work is to be done in non-human cells rather than human cells, the rationale for this choice is strong.
Conflicts: 
  • Shoukhrat Mitalipov
  • William Matsui

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