Funding opportunities

In vitro differentiation of hESCs into corticospinal motor neurons

Funding Type: 
SEED Grant
Grant Number: 
RS1-00170
Principle Investigator: 
Funds requested: 
$500 000
Funding Recommendations: 
Recommended if funds allow
Grant approved: 
Yes
Public Abstract: 
Statement of Benefit to California: 
Review Summary: 
SYNOPSIS: In this proposal an attempt will be made in-vitro to guide hESC towards corticospinal motor neuron (CSMN) differentiation. A connection between the mouse work from a few labs to human CSMN biology will also be tested. Three questions will be addressed in this proposal: 1) Is Fez-like biology functional in humans, 2) Are focused markers of differentiation pathways turned on when hESC are differentiated into CSMN, and 3) Can new neurons be formed in Fez-like knockout mice that themselves do not form them? SIGNIFICANCE AND INNOVATION: Relatively little attention has been given to characterizing the differentiation and development of CSMNs compared to spinal motor neurons, perhaps at least partly because of the absence of markers for these cells. A greater knowledge concerning these cells and ways to derive them from hESCs may increase our knowledge of the motor system as well as our understanding of the pathogenesis and treatment of various motor neuron disease, such as amyotrophic lateral sclerosis. For these reasons, the topic of the proposal is a significant one. The focus on methods of deriving CSMNs from hESCs and the planned transplantation of these cells into Fezl-/- mice to look at their functional capacities are novel directions and approaches. This is important yet risky work, that will provide important foundations if successful, and will likely give useful negative data even if not fully achieved. STRENGTHS: This proposal is based on strong biology, and attempts to be at the forefront of the field of directed differentiation of ES cells. The efficient differentiation of hESCs into CSMNs is an important goal, and the PI, a newly appointed Assistant Professor in the Department of Molecular, Cellular, and Developmental Biology at UCSC, has experience in transplantation procedures into rodents as well as issues related to CSMNs. The plan of the experiments is logical, progressing from the identification of markers to differentiation to transplantation. The PI also addresses some anticipated problems/pitfalls related to the planned experiments. The preliminary data of the PI with respect to Fezl and the generation of the Fezl-/- mouse are interesting and lend support to the planned studies. WEAKNESSES: This is a risky proposal, and the PI would have benefited from collaborations with others who are expert in the differentiation of ESCs. A failure to identify appropriate markers for CSMNs will make Specific Aim II difficult to achieve, and a failure to differentiate CSMNs will make Specific Aim III impossible to achieve. If the three markers listed by the PI under Specific Aim I are not relevant to human CSMNs, what will the PI do? The description of the roles of Fezl and others are a bit oversimplified from what is known about the need for combinatorial controls. Further details related to the transplantation and other experimental details would have been valuable. For example, how were the times of sacrifice (3 weeks to 3 months) determined? Also, in Specific Aim III it would be valuable to stain the transplanted knockout animals for Fezl. DISCUSSION: ALS is an important disease focus, and focusing on one of the two neuron types that die is important. A key drawback to this proposal is the lack of collaborators with a track record in the embryonic stem cell field and culture. The PI is well trained, and the primary author on a very relevant publication in PNAS, but has only produced one paper since receiving a PhD in 2000. The approach is somewhat naïve in that the description of differentiation into CSMN does not include a discussion of possible combinatorial approaches (Fezl and other genes). Also, there is human homology for Fez-like, but whether it will be a marker is unclear. Aim 3 would produce a very nice knockout mouse model, but it will take a lot to get there. One reviewer commented that the applicant should go slower and more stepwise, and stick to the fundamental biology. PROGRAMMATIC REVIEW: In programmatic review, it was noted that this is a young investigator working in the area of coticospinal motor neurons which degenerate in ALS. It was also noted that based on the scientific score, this investigator was near the top of the 'Recommended if funds available' pool. It was agreed that this investigator be moved to a 'Special funding pool' that would have priority for funding if additional monies became available.
Conflicts: 

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