Basic Biology III
Human pluripotent stem (hPS) cells can perpetually replicate themselves and can differentiate into virtually any type of cells in the adult body (pluripotency). Human embryonic stem (hES) cells, which are derived from blastocyst embryos, are the most studied type of hPS cells. The recent technological breakthrough has paved the way to convert adult differentiated cells such as skin cells into undifferentiated cells called human induced pluripotent stem (hiPS) cells that maintain pluripotency. Because of this enormous ability, hPS cells are thought to be the potential source of the future cell transplantation therapy for the treatment of diseases such as Parkinson's disease and diabetes mellitus. Despite these promising functions of hPS cells, there are many biological questions and practical hurdles to overcome. Important issues include understanding how the unique functions of hPS cells are regulated and development of technologies to derive and grow hPS cells at high efficiency under completely animal-free conditions for future medical purposes. We have recently identified a signaling pathway (like a hormone that mediates biological information) that controls cell-cell communications and survival of PS cells. Moreover, by controlling this pathway, we have established a method to grow hPS cells under a fully animal-free condition. In this proposal, we will investigate whether this signaling pathway also controls the unique functions of hPS cells. Furthermore, by changing the activity of this signaling pathway, we will develop a new method to efficiently generate and expand new hiPS cell lines that are completely free from animal-derived materials.
Statement of Benefit to California:
A major focus of our research proposal is to develop novel technologies by which human embryonic stem (hES) and human induced pluripotent stem (hiPS) cells can be propagated under completely defined conditions. The establishment of such a new method would impact virtually all hES and hiPS cell-based application programs as it involves a common basic process required to expand undifferentiated pluripotent stem cells before turning them into any type of adult cell for future therapeutic purposes. It is therefore predictable that the new methodology will be promptly translated as an intellectual property to be commercialized, and would substantially activate the biotechnology field in the State of California. More importantly, the new methodology will be provided to the Institutes in California at the highest priority where the method will accelerate the process and continued research necessary to see the clinical application of pluripotent stem cell-based transplantation. Furthermore, the introduction of pluripotent stem cell therapies into California medical care as a direct result of our research and new methodology would further enhance the quality of medicine and medical technology available for California citizens.
Project Synopsis: The goal of this proposal is to elucidate mechanisms by which a specific molecular complex may act to regulate self-renewal and differentiation programs in human pluripotent stem cells (hPSC). In the first aim, the applicant will explore whether this complex, which was recently described as a mediator of cell-cell interactions and hPSC survival, might also regulate hPSC self-renewal mechanisms, and whether its role changes as cells differentiate. Next, the applicant will investigate whether this complex is reciprocally regulated by upstream signaling pathways involved in self-renewal. Finally, in the third aim, the applicant will determine whether manipulating the complex can enhance the efficiency of cellular reprogramming. Significance and Innovation: - The potential impact of this work is uncertain, as it will likely generate a complex array of outcomes that would be difficult to exploit in a meaningful way. Additionally, there are several other groups pursuing similar lines of research, and it is not clear whether this approach offers unique advantages. - The proposed experiments explore novel hypotheses surrounding a potential relationship between cell architecture and stem cell behavior. Experiments are clearly focused on understanding basic cellular mechanisms. - If successful, this effort could lead to original insights about the role of an interesting molecular complex in hPSC survival and behavior. It may also lead to improved methodologies for deriving and expanding induced pluripotent stem cells. Feasibility and Experimental Design: - Reviewers believed that a pivotal component of the experimental plan is the construction of engineered cell lines for manipulating the expression of the molecular complex under investigation. Failure to establish such quality lines would make it impossible to pursue remaining project goals. Development of these tools should comprise an early stop/go decision. - Reviewers firmly believed that if the hypothesis to be tested in Aim 1 proves untrue, then the goals of the remaining Aims lose their relevance. - Reviewers argued that while there is preliminary data to support the underlying premise, much of the rationale for the experimental design is based on circumstantial evidence reported by others, particularly with respect to the studies addressing integrin, ROCK signaling and autophagy. In contrast, they viewed the studies addressing the role of Wnt to be logical and feasible. - Reviewers noted that the inhibitor used for experiments described in Figure 2i-k has been reported to act on other members of the molecular family being investigated and therefore may not be as specific as suggested. They felt it would be prudent to more thoroughly characterize the effects of this inhibitor on hPSC before launching an extensive set of knockdown experiments. - The overall study design lacks focus. Reviewers suggested that an in depth analysis on a narrower set of goals would improve the likelihood of success, particularly as the sheer complexity of interactions between the various pathways investigated will make outcomes potentially difficult to interpret. Principal Investigator (PI) and Research Team: - The PI has solid expertise in the regulation of pluripotent stem cells. While the publication and funding track records are modest, he/she has a number of patents and pending grants. - The co-investigator is an expert in the molecular complex being studied and represents a major strength or the proposal. - The research team is qualified to carry out the proposed studies. The budget is appropriate and the facilities are excellent. Responsiveness to the RFA: - The proposed studies utilize hPSC and address molecular and cellular mechanisms relevant to human stem cell biology. - A small number of experiments comparing mouse and human cells were included without being justified by the applicant as groundbreaking or necessary. Reviewers considered those experiments nonresponsive.
- This application scored below the initial scientific merit funding line, no programmatic reason to fund the application was proposed, and the GWG voted to place the application in Tier 3, Not Recommended for Funding.
- Ali Brivanlou