Funding opportunities

Characterizing the Molecular Mechanisms of Pluripotent Stem Cell Self-Renewal

Funding Type: 
New Faculty II
Grant Number: 
Funds requested: 
$2 430 000
Funding Recommendations: 
Not recommended
Grant approved: 
Public Abstract: 
Human pluripotent stem cells (PSC) have the potential to produce all of the different cells in the human body. There are many studies using human PSC to develop new therapies for patients with genetic disease, birth defects or injuries. One obstacle to the development of these stem cell therapies is that very often human PSC randomly differentiate (or change) into non-specific cell types in the laboratory. When the cells randomly differentiate they can not be used to make the cells or tissues of interest. It’s not clear how to prevent this random differentiation because at this time we do not completely understand the biology of these human stem cells and what factors keep them alive and expanding. A second problem with current stem cell research is that the most reliable method to keep the human stem cells alive and expanding is to culture them on cells obtained from mice. This culture with animal cells makes the currently available human stem cells unsafe for use in patients. The overall goal of these studies is the development of safe methods to produce human stem cells to make cells and tissues for patients. We are developing a completely new approach to keep human stem cells growing. We are developing a synthetic material based on a normal human protein that is known to be required for human stem cells to survive and expand without differentiation. Excitingly, our preliminary studies demonstrate that by adding our material, we can keep stem cells alive, and undifferentiated for at least three days (the longest time evaluated). The goal of these studies is to continue and further study our new approach, and evaluate its potential to maintain stem cell survival and growth. We will also study the genes and proteins involved in normal human stem cell survival, to learn more about the biology of these cells, and optimize the development of materials for maintaining human stem cell survival and expansion for clinical applications. If successful, the development of such synthetic materials that maintain the undifferentiated state of stem cells will make all aspects of human stem cell research more reliable and efficient. This in turn will increase the time that researchers have to developing therapeutic protocols, rather than spending a large amount of their time and energy to keep their cells alive and renewing. Furthermore, it will facilitate the scale-up of stem cell applications from small-scale laboratory studies. Thus, our studies have the potential to impact on all human stem cell research, and increase the rate of development of all stem cell based therapies. The development of a safe, synthetic material that can replace mouse cells, and reliably keep the stem cells alive would prove very useful to stem cell researchers, as well as increase the safety of the cultures for therapeutic purposes.
Statement of Benefit to California: 
One of the biggest hurdles in human stem cell research is that in the laboratory, the cells spontaneously and randomly change into a variety of cell types in culture by a process called differentiation. Cells that have undergone uncontrolled differentiation can not be used to make cells or tissues for use in patients. A high degree of technical training can reduce this random differentiation, but it is not preventable with currently available techniques to grow stem cells. Our project will study the biological processes that keep stem cells renewing and expanding. We will then use this information to develop new materials that can be added to cells in the laboratory to keep stem cells growing in an undifferentiated state - so that they can better be used to make cells and tissues for clinical applications. If successful, these materials will increase the efficiency of all human stem cell research, by dramatically decreasing the amount of time spent researchers spend keeping human stem cells alive and undifferentiated. In turn, this will allow researchers to spend more time and effort developing stem cell therapies for human disease. Finally, since our material will be made in a laboratory using safe procedures, it will also increase the safety of the stem cells used to make cells and tissues for patients, by reducing the need for animal or human products in the system.
Review Summary: 
The overall goal of this project is to improve our understanding of the pathways involved in human pluripotent stem cell self-renewal and to use this knowledge to develop new methods or factors to promote maintenance of pluripotency in human stem cells. A number of different transcription factors have been identified that participate in human embryonic stem cell (hESC) self-renewal, and this proposal seeks to more fully understand the specific role played by one of these factors. In Aim 1, the applicant will assess the biological role of the transcription factor in maintaining pluripotency of hESC and induced pluripotent stem (iPS) cells by analyzing effects of expressing two mutant versions of this factor - one with enhanced activity and one with dominant negative activity. The goal of Aim 2 is to identify the proteins that bind to the transcription factor using several approaches. Aim 3 is focused on the development of small protein mimics of the transcription factor that could be used to promote expansion of hESCs for clinical applications without the need for feeder cells. This application presents a very straightforward and well-structured research plan. The applicant suggests that the difficulty in efficiently culturing hESCs in an undifferentiated state is due in part to a limited understanding of the pathways that regulate self-renewal. Improving culture efficiency would assist the field, yet other recent advances have resulted in improved ease of culture, so the impact on the regenerative medicine field may be relatively low. Furthermore, the significance of the proposed research is somewhat diminished given the relative expendability of the transcription factor under investigation for reprogramming of somatic cells to iPS cells. However, new basic knowledge of the role of the transcription factor in self-renewal is likely to be gained. The research plan is well designed, and the fact that the studies will focus on hESCs and iPS cells and on comparing the two cell types is considered a relative strength of the proposal. However, the reviewers identified several limitations of the proposed studies related to availability of critical reagents, choice of markers for analysis of transfected cells, and questions about the feasibility of certain approaches. The third aim is the most speculative and least developed aspect of the proposal, but the most interesting aim, likely to have an impact on clinical applications if successful. The preliminary data are supportive of the proposed concepts and hypotheses, but the reviewers had different opinions about their scope. Overall, this proposal presents an interesting, conservative approach, lacking innovation but entirely focused on human ESCs and iPS cells. The applicant was recruited to a tenure-track Assistant Professor position at the home institution in 2004. She/he was instrumental in establishing an hESC core facility and is currently its director. In addition, she/he holds leadership roles in stem cell and gene therapy committees in national societies. One reviewer expressed concern that this many commitments to committees and directorships may diffuse his/her research time during this formative career stage. The applicant has developed many of the research tools required for the project, and has the necessary expertise to see the project to fruition. However, the reviewers expressed only moderate enthusiasm for the applicant’s productivity; the publication record is not extensive, particularly with regard to stem cells, and much of his/her published work over the last few years has been related to viral vector work. The applicant’s overall research program was judged to be somewhat diffuse. She/he has, however, attracted considerable extramural research funding. The applicant has provided a relatively well-structured career development plan with detailed goals outlined, but provides no clear timeline or plan of action for how these goals will be met. The mentoring plan identifies five scientists and clinicians, some of whom are world-class leaders in relevant fields. However, one reviewer questioned whether sufficient interactions between the applicant and the mentors were planned. The home institution provides excellent academic, intellectual and financial support for the applicant’s research and ample protected research time. The home institution has a long history of commitment to stem cell research and has recruited a number of stem cell biologists over the past few years. New faculty appointments in stem cell biology are envisaged with two of these in the same department as that of the applicant. Overall, the reviewers were not enthusiastic in their assessment of this application. Although the proposal was considered to be straightforward and have some potential significance, the reviewers felt it lacked innovation and was not sufficiently ambitious. Similarly, they commended some of the applicant’s accomplishments, but were only moderately enthusiastic about his/her productivity and scientific focus.

© 2013 California Institute for Regenerative Medicine