Funding opportunities
Functional Significance of hESC Glycotype Diversity: Fate Specification & Immune Tolerance
Funding Type:
Basic Biology II
Grant Number:
RB2-01549
Funds Committed:
$1,354,926
Funding Recommendations:
Not recommended
Public Abstract:
Biological molecules, the building blocks of cells, exist in many forms. Commonly, scientists study the role of proteins in normal and disease processes. As a result, the functions of other types of molecules are less well studied. Consequently, significant gaps in knowledge exist. For example, the role of sugars is often ignored. Until now, this has been largely the case in the human embryonic stem cell (hESC) field. Why is this a significant omission? There is a great deal of evidence that suggests sugars could play important roles in the basic biology of these cells and their suitability for patient therapies. Some of these data come from the field of medicine that deals with blood transfusions. Attempts to treat the illness of one individual using donated blood has been documented for hundreds of years, but prior to the early 1900s, there was no explanation for why this procedure benefited some patients and not others. In 1901, a physician described a code based on sugar structures that distinguishes groups of individuals. Matching the code of the donor and the recipient cells prevented rejection and enabled routine blood transfusions. It is now understood that the biosynthetic machinery for producing these determinants is heritable. As a result, various groups within the population express different codes (blood types), the reason that blood banks need many donors. Subsequent work has shown that these structures are present on many other types of cells. Preliminary experimental evidence from our group shows that hESCs also have this sugar code. As with blood cells, they can be grouped according to type. Additionally, the work of others suggests that the expression patterns of these determinants change as the cells differentiate. In this context, we propose two sets of studies. The first focuses on undifferentiated hESCs that are in a constant state of self-renewal. We want to create a bank of cells that is representative of the sugar codes of United States citizens. We also want to precisely define the nature of the sugar determinants and the proteins that carry them. In addition, we will focus on hESCs that have been induced to differentiate into cells that make up the pancreas, heart, and nervous systems. We think that the distribution of the sugars and the protein scaffolds that present these structures change during the differentiation process. The second study is comprised of functional analyses. In these experiments, we will test the hypothesis that the sugar code influences the basic biological properties of hESCs. Specifically, we will determine if these structures play a role in their ability to self-renew or differentiate. We will also determine if, like blood cells, the body is able to recognize and destroy stem cells based on mismatches in the sugar code. Thus, at the conclusion of these experiments we will know whether it will be important to consider these sugar structures in differentiation and/or transplantation procedures.
Statement of Benefit to California:
It is envisioned that human embryonic stem cells (hESCs) will be the foundation on which regenerative medicine therapies will be built. Investigative teams are working hard to translate research findings into cell-based treatments for patients with a wide variety of chronic and/or fatal diseases. While the importance of these efforts cannot be overstated, it is essential to realize that hESC biology is a very young field, slightly more than a decade old. In general, it takes many years for new discoveries to be fully understood. Accordingly, it is important to keep learning about the basic properties of these cells as gaps in our knowledge could create major delays in developing and implementing clinical applications.
The project we propose in this application benefits the citizens of California by investigating early on, during the development phases when treatment strategies are being devised, an issue that might become a confounding factor when clinical trials begin in earnest. Specifically, we are investigating the role of sugar structures that are matched between donors and recipients in blood transfusions. We think that these molecular signatures might play important roles in governing basic aspects of hESC biology. This phenomenon could account for, at least in part, the differences in behavior that have been observed among the lines. Additionally, their sugar codes could play a role in the response of patients’ immune systems to cell replacement therapies that utilize hESCs. If these determinants are mismatched in blood transfusions, the donor cells are often destroyed, which illustrates the possible hazards.
What can we do with the information that emerges from this study? We can take preventative measures now so that, in the future, therapeutic applications do not encounter costly delays in both human and financial terms. For example, if we know that it is important to match the sugar type of the donor and the recipient, then we can take this information into consideration. This would allow investigators to use the strategies that have been developed for blood banking to create the collections of cells that will be needed for hESC-based therapies. Since the development of lines that grow continuously in culture and differentiate in predictable ways is a long process, these efforts should begin as soon as possible.
By sponsoring projects that are designed to advance stem cell science, the California Institute for Regenerative Medicine is one of the most exciting ventures in state history. Citizens voted for this initiative because they believe that it is important to put tax dollars into research that has the potential to develop novel therapies and cures for major human diseases. Accordingly, the benefits of this work will accrue not only locally, within the state, but to all patients who have medical conditions that are amenable to these types of therapies.
Review Summary:
EXECUTIVE SUMMARY
The goal of this proposal is to assess the significance of blood group antigen expression (blood type) on human embryonic stem cell (hESC) pluripotency, differentiation and immunogenicity. Blood group antigens can be expressed on a variety of cell types, including stem cells, and are important determinants of immune response. In Aim 1, the applicant proposes to profile the repertoire of blood group antigens expressed in ten hESC lines and their differentiated progeny. In Aim 2, the applicant proposes to examine the function of blood group antigens in regulating the pluripotency, differentiation and immunogenicity of hESCs.
Reviewers found the rationale for much of the proposal to be unclear. They noted that undifferentiated hESCs will not be transplanted to human patients and thus better justification is required for characterizing blood group antigens on these cells rather than focusing on their differentiated progeny. In addition, the hypothesis that these antigens influence hESC self-renewal is questionable, given that the lines to be studied presumably express very different combinations of antigens, yet they all self-renew. Given the emerging field of induced pluripotent stem cell (iPSC) research and its potential to address issues of immunogenicity, one reviewer suggested that a more useful study might examine the stability of blood group antigen expression on iPSCs and their differentiated progeny compared to their source. Reviewers also questioned the rationale for specific aspects of the research plan. In Aim 1, blood group antigen carrier proteins will be profiled, but the relevance of these proteins is not explained. In addition, the applicant focuses on one type of glycosylation of blood group protein antigens while ignoring more predominant types of glycosylation and blood group antigens on glycolipids. The full complement of these epitopes must be considered to understand cellular and immune response. Reviewers did not find the proposal to be particularly novel or innovative and commented that the proposed methodologies are fairly standard in proteomics.
The reviewers found the preliminary data sparse and raised a number of concerns about the research plan. With regard to Aim 1, they noted that the proposed generation of differentiated cell types is poorly described and not supported by relevant preliminary data. It is not clear how time points will be chosen for analysis or how many will be assessed. With regard to Aim 2, reviewers expressed concern that no data supporting the efficacy of the function-perturbing antibodies on hESC derivatives is included. Reviewers also commented that the focus, in the second part of Aim 2, on antibody dependent complement-mediated lysis (ADCML) seems too narrow and relatively uninformative regarding the fate of cells in vivo. In addition to eliciting ADCML, blood group antigens may influence cell homing, signaling and engraftment.
Reviewers praised the applicant’s track record of productivity in proteomics and mass spectrometric characterization of protein complexes. However, they were uncertain about the experience of the applicant in directing the project in the capacity of a principal investigator. Furthermore, they were concerned about the effort commitment from the key personnel proposed for the study, which will likely limit its productivity.
Overall, reviewers raised significant doubts about the rationale for this proposal and did not find the preliminary data to be sufficient to support its hypotheses.
Conflicts:
