Funding opportunities

WNT signaling in neural stem cell self renewal and neural crest cell specification and migration

Funding Type: 
Basic Biology III
Grant Number: 
RB3-02087
Funds requested: 
$1 375 854
Funding Recommendations: 
Not recommended
Grant approved: 
No
Public Abstract: 
Human pluripotent stem cells (hPSCs) and their derivatives represent the only research tool to study human development. As such, these cells allow us to study the progression of diseases at the cellular level in a dish, probe how specific genetic defects contribute to the myriad of developmental and birth defects, and generate the “raw material” for the development of cell-based therapies of presently incurable diseases, such as cancer, cardiovascular disease, and neurodegenerative disorders. However, our understanding of the basic mechanisms underlying stem cell biology is incomplete, and the processes by which individual cells organize each other to give rise to the complexity of multi-cellular life remain mysterious. At the heart of embryonic development lies an intricate process of cell communication. Individual cells within the developing organism produce and release signals, known as growth factors, that instruct neighboring cells to assume specific behaviors and properties. Unique combinations of such growth factors regulate a multitude of developmental processes, including the growth and differentiation of hPSC. Wnt proteins represent a major class of growth factors with potent effects on stem cells and developmental processes. However, despite 30 years of research on these proteins with over 1,800 publications in 2010 alone, the mechanisms by which Wnt proteins elicit specific cellular responses and regulate stem cell biology remain poorly understood. One reason for the slow progress in the study of Wnt proteins is that their manipulation in biological systems has been difficult and even impossible. We have developed powerful technologies that allow us to systematically dissect the role of Wnt signaling in regulating the behavior of hPSC. By developing the means to isolate Wnt proteins we are now able to examine their effects on stem cell growth and differentiation. We will specifically examine the role of Wnt signaling in neural stem cells, a cell type that can generate many of the cells found in the central nervous system (CNS). In addition, in collaboration with our research partners [REDACTED] we propose to examine the effects of Wnt signaling in Axolotl, a newt with remarkable regenerative potential in the CNS. By performing a comparative analysis between human and Axolotl neural stem cells we will be able to identify the unique properties that allow efficient regeneration of the Axolotl CNS and apply this knowledge to design strategies for stimulating regeneration of the human CNS. By identifying the mechanisms by which Wnt proteins act we will contribute valuable tools and protocols for the manipulation and specific differentiation of hPSC and neural stem cells into mature cell types that can be utilized in cell replacement therapies. Additionally, these studies will pave the road to designing strategies that stimulate cells in the central nervous system to repair damage.
Statement of Benefit to California: 
The rise in life expectancy to over 80 years will likely lead to an increase in the number of people suffering from age-related diseases, such as cancer, heart disease and neurodegenerative disorders. Current medical treatments can control, but not cure, such diseases. Recent advances in the study of human pluripotent stem cells (hPSCs) have provided the opportunity to develop novel cell replacement therapies for the treatment of many such diseases. Development of novel cell based therapies will also overcome the inadequacy of conventional drug-based treatments. Several scientific obstacles need to be overcome before the full potential of hPSC-based therapies can be realized. First, sufficiently large numbers of clinical grade hPSC that can be thoroughly tested and characterized need to be derived. Second, robust protocols for the directed differentiation of hPSC into functionally mature cell types suitable for transplantation need to be developed. To address these challenges we propose a set of experiments that will significantly expand our understanding of basic biology of hPSCs and one of their descendants, neural stem cells. One powerful approach to affect hPSC behavior is through the manipulation of the extracellular environment. We are particularly focused on one class of potent stem cell factors, called Wnt proteins. We and others have shown that Wnt proteins can dramatically affect the growth of stem cells so that Wnt-stimulated cells in some cases grow more robustly or in other cases produce specific mature cell populations. These experiments will enable the development of cost-effective protocols for the large-scale production of undifferentiated hPSC and functionally mature cell types. Our proposed research is fundamental to applications of hPSC in regenerative medicine and has broad benefits to researchers with a wide spectrum of scientific interests. This research will not only benefit the health of Californians, but also the California economy by developing new reagents, protocols and technologies that will be adopted by existing companies as well as seed and complement novel business ideas. The outcome of this project will contribute to the development of a biotechnology platform that can provide great benefits to the advancement of California biotechnology. The patents, royalties and licensing fees that result from the advances in the proposed research will provide California tax revenues. Thus, the current proposed research provides not only the essential foundation for the scientific advances in regenerative medicine to improve health and quality of life, but also potential technology advancement and financial profit for the people in California.
Review Summary: 
Project Synopsis: The overall goal of this proposal is to define the role of Wnt signaling in development and differentiation of human neural stem and progenitor cells (hNPC) derived from human embryonic stem cells (hESC). The properties of hNPCs and their derivatives will be compared with those of their in vivo counterparts from the axolotl, an amphibian with a remarkable capacity to regenerate its central nervous system (CNS). As the human CNS displays very little regeneration following damage, the applicant reasons that such comparisons may provide insight into mechanisms that could lead to improved strategies for regenerating human CNS. In the first Aim, the effects of Wnt signaling perturbation on NPCs derived from hESC and axolotl spinal cords will be characterized and compared at the cellular and molecular level. The mechanism of Wnt action will be further explored in Aim 2 by investigating the contribution of a specific downstream effector. For the third Aim, the applicant will examine the role of Wnt signaling in human neural crest (NC)-like cells, a transient subpopulation derived from hNPCs. Significance and Innovation: - Although comparisons between human and axolotl NPCs might provide insights into CNS regeneration in axolotls, numerous comparative studies between humans and model regenerative species have not led to major advances in regenerative medicine. - If successful, these studies may offer insights into the activation of endogenous repair pathways in humans or enhanced ability to produce robust hNPC populations for therapeutic applications. Such results have the potential to be transformative. - The rationale for choosing to examine hESC-derived neural crest cells is unclear and poorly connected to the rest of the proposal. - While the focus on Wnt signaling is logical, the proposed studies are largely descriptive and not organized around clear hypotheses. Feasibility and Experimental Design: - Although the overall rationale for the study is strong, the description of the experiments to be undertaken was not organized in a manner to logically test the hypotheses described. - Reviewers found serious deficiencies in the experimental details, particularly with regard to the phenotypic analysis and the predicted effects of the different treatments. - No rationale for the inclusion of experiments on two additional Wnts, Wnt7a and Wnt11, was provided. - The proposed experiments represent logical extensions of the applicant’s previous and ongoing efforts. Many of the necessary tools and reagents have already been developed. - Reviewers were not convinced that some of the proposed experiments were designed appropriately to yield meaningful and interpretable results. The proposal would have benefitted from a more thorough discussion of the scientific questions to be addressed and a clear explanation of how the resulting data would be interpreted. - The focus of Aim 3 on neural crest-like cells is discordant with the rest of the proposal, which concerns the biology of NPCs. Inclusion of this aim detracts from the overall feasibility of achieving project goals within a three-year timeline. PI and Research Team: - The principal investigator (PI) has a very strong background in Wnt signaling as well as the culture and handling of hESC. - Appropriate co-investigators have been recruited to assist with neural progenitor derivation and bioinformatics analysis. - The partner PI is a well-regarded expert on the axolotl nervous system and a world leader in the use of this organism to study regeneration. - Reviewers had mixed impressions of the level of integration between the collaborating teams. While some admired the synchrony, others considered the link between them to be tenuous. Responsiveness to the RFA: - The proposed research utilizes hESC and addresses fundamental questions in human stem cell biology. - The axolotl studies were not viewed as particularly responsive, and reviewers were not convinced that studies of Wnt signaling in axolotls would readily translate into fundamental mechanistic insights about human stem cell biology.
Programmatic review: 
  • 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.
Conflicts: 

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