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RS1-00187-1: Precursors isolated from human embryonic stem cell culture: potential use for kidney regeneration

Recommendation: Not recommended for funding

Public Abstract (provided by applicant)

End stage renal disease has reached epidemic proportions in the United States. In the U.S., more than 400,000 people are on chronic dialysis every year and another 20,000 patients have undergone a kidney transplant. Currently dialysis and transplantation remain the only viable treatment despite significant complications with both. This along with an increasing shortage of organs has heightened interest in developing novel methods of therapy for kidney replacement. A great deal of attention has recently been focused on stem cell technology as an alternative modality for the treatment of a variety of diseases. This is particularly true in the fields of tissue engineering and regenerative medicine, because stem cells have the capacity to differentiate into a variety of cell types and represent a potentially in-exhaustible source of cells that can be used to regenerate new tissue for transplantation. A great deal of effort is focused in our laboratory to researching novel sources for stem cell lines. We have demonstrated that in culture, a new population of stem cells derived from human amniotic fluid exhibit the capacity to differentiate into normal organ structures in developing embryonic kidneys. These results encouraged us to pursue live animal models which have demonstrated similar outcomes after the induction of injury to the kidney. This is an important concept because this may represent a novel method for cell therapy in those patients with End Stage Renal Disease where recovery of the organ could be made possible. In our efforts of continuing to examine other stem cell lines we would like to isolate another novel population of stem cells derived from human embryonic stem cell cultures and apply these cells to regenerative applications in the laboratory to determine their potential value in tissue regeneration. At the edge of human embryonic stem cell colonies, there exists a heterogeneous population of cells both differentiated and partially differentiated. These cells do not have the same pluripotential capacity as the undifferentiated stem cells near the center of the colony but also do not form teratomas (tumors), which affords them greater advantage for future clinical applications. We plan to isolate these precursors cells, establish their optimal culture conditions and characterize them. In addition we would like to show that these cells can differentiate into kidney cells and confirm that they do not form teratomas in live animals. The isolation of these precursor cells from human embryonic cultures may prove helpful in understanding some of the mechanisms of development and cell differentiation. However, most important characteristic of these novel precursor cells is that they may have viable future clinical applications, since they can be derived in virtually an unlimited amount and likely avoid the risk of tumor formation when applied into biological systems.

Statement of Benefit to California (provided by applicant)

The State of California is home of some of the nation’s best organ transplant centers. Having our laboratory next to one of California’s busiest pediatric renal transplant programs in the country, {REDACTED}, we became acutely aware of the growing shortage of organs for our patients. While the need for donor organs continues to rise every year, emerging technologies such as those offered by stem cell research may assist our patients and the rest of the citizens in California with alternative technologies that perhaps can make a significant impact in regenerative medicine and transplantation therapies. Although human embryonic stem cells show the capacity for multiple differentiation and the prospect for new medical cures, they also have an inherent propensity to form teratomas ( benign tumors) in culture which may actually limit their use for future clinical applications. However, a very novel population of precursor cells exist on the periphery of a typical human embryonic stem cell colony that maintain the ability for multiple cell line differentiation but will not form teratomas. These cells could possibly be a safer alternative for regenerative applications aimed at evolving into human clinical therapies that would help those citizens of California in need of organ replacement therapies in the future. We intend to characterize the potential these cells have for differentiation into various cell lines and test their capacity to differentiate into renal cells in the laboratory, a process by which we have extensive experience and success in performing with other precursor cells.

Review

SYNOPSIS OF PROPOSAL: The goal of this proposal is to use a unique subpopulation of human ES cells to generate renal-like structures in vitro and in vivo. This idea is based on the fact that ES cells often form complex teratomas after transplantation in vivo. It is proposed that some human ES cells are more restricted in their developmental potential and therefore tend to differentiate along specific lineages rather than into complex teratomas. This proposal aims to test this hypothesis and the proposed studies are expected to form the basis for future clinical applications.

INNOVATION AND SIGNIFICANCE: The applicants propose a new approach toward the use of human ES cells in the treatment of end stage renal disease. The apparent goal of this proposal is to identify cells that might be potentially useful for kidney generation. This proposal has two specific aims. The first is to identify the relevant cells (“preferred cells”) from hESC colonies in culture and characterize these cells to determine their potential for mesodermal tissue derivation. The second specific aim is to determine the capacity of cells derived from hESC cultures to survive, replicate and integrate themselves into whole embryonic kidneys in culture. Their potential to survive and replicate without forming teratomas in vivo in animals will also be investigated. The significance is high, because generation of cells that are useful for treating end stage renal disease represents a significant endeavor.

STRENGTHS OF PROPOSAL:In addition to the innovative nature of the proposal, the applicants have previously shown that some harvested cells can recapitulate some essential steps in nephrogenesis. Therefore, the applicants are highly committed to the use of regenerative methods in the treatment of renal disease and have demonstrated the ability to successfully develop innovative approaches to this problem.

A strength of this proposal is the experience of the PI in his/her field, as well as the PI’s productivity and dedication to solving the problem of kidney failure.

WEAKNESSES OF THE PROPOSAL: This proposal unfortunately suffers from a number of weaknesses.

It is repeatedly asserted that some ES cells do not form teratomas in vivo. Indeed, this constitutes the main justification for the proposal. Remarkably, however, this claim is not backed up by any citation or unpublished data. In fact, a major component of one of the aims is to show that teratomas do not form when the “preferred cells” are introduced. They can’t have it both ways.

Other experiments are poorly explained and poorly justified. The applicants suggest that they might try using cells derived from embryoid bodies if their primary approach is not successful. At no point is it explained what are the relative merits of choosing to the “preferred cells” versus embryoid body cells. It is a bit disappointing that the applicant omitted obvious relevant experiments, although this is not a major problem and might be too much to expect at this early stage.

The PI spends a lot of time describing experiments that are relevant neither to the specific aims or the goals of this RFA.

It is not clear at all why the author prioritizes, as defined in the hypothesis, the use of the “preferred cells”. Why is this strategy better than the conventional approach?

Additional specific weaknesses include the following. The roles of proposed collaborators are not justified; it is not explained why their advice requires a bi-annual visit to California. The applicants ask for funds for animal experiments in the first year of the proposal but the time line indicates that no animals will be used until year two. It is not clear why the applicant chose to refer to the “preferred cells” in the way s/he did. The last thing the field needs is novel jargon not supported by scientific evidence. In addition, there seems to be a misunderstanding as to the definition of a hESC.

This grant could be funded by the NIH, and does not require CIRM funding.

DISCUSSION: Concern was voiced about the premise of the proposed studies, the claim that the “preferred cells” do not form teratomas. Discussion focused on whether the applicant understood the definition of hESC.The proposal could have benefited from the inclusion of more detail about embryoid body experiments that were mentioned as a backup strategy.

The following Working Group members had a conflict of interest with this application and were therefore recused from participating in review of, discussion of, and voting on the application:

• None