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RL1-00671-1: Derivation of clinical grade HLA-homozygous human stem cell lines using parthenogenetic activation of oocytes

Recommendation: Not recommended for funding

Public Abstract (provided by applicant)

The goal of these studies is to develop a bank of special stem cell lines from which therapeutic cells may be created that will not be immune rejected by millions of patients of different ages, sexes and racial groups. Researchers and clinicians may use these stem cells as source material to develop new treatments of previously untreatable degenerative and hereditary diseases, including; paralysis, blindness, Alzheimer’s disease, diabetes, hemophilia, Huntington’s disease, muscular dystrophy, Parkinson’s disease, sickle cell anemia and many others.

One of the most serious problems of cell transplantation is immune rejection of the cells. The success of transplantation depends on creating cells that are immune matched with the patient. These cell lines carry the special property called “HLA-homozygosity” that makes them easier to immune-match with large segments of the human population for therapeutic use.

Human stem cells known as “embryonic stem cells,” can be changed into any cell found in the body. These cells are usually obtained from fertilized eggs and this is an ethical problem for many people who consider destroying a fertilized egg to be the same as destroying a human life. We address this problem by using unfertilized human eggs to create the stem cell lines. The process of deriving these cell lines is called parthenogenesis. These cell lines divide infinitively and represent an inexhaustible source of derivatives and have been shown to have the same ability as embryonic stem cells to become any cell in the body. For this work it is necessary to obtain a limited number of human eggs from informed donors under conditions and review dictated by state and federal regulations.

The method of parthenogenesis has already proven to produce several HLA-homozygous stem cell lines. One parthenogenetic HLA homozygous stem cell line has already been created that is immune-matched with hundreds of millions of people of differing sexes, ages and racial groups. Nuclear transfer has yet to produce a human patient matched embryonic stem cell line. A method of reprogramming of the patient’s somatic cells and its nuclei has not resulted in the creation of clinically useful patient matched stem cell lines. Moreover, all cells of patients with hereditary diseases carry the corresponding genetic defect. In these situations it would be important to use pluripotent stem cell lines obtained from healthy donors, not from the patient.

It is necessary to create additional {REDACTED} lines matched with other large segments of the human population. Establishing a bank of {REDACTED} lines and tissue repository from these lines may potentially eliminate the concern for immune rejection and be used to treat millions of persons worldwide. In addition, the cell bank will result in intellectual property that will bring increased valuation for California research organizations, increased investment, and increased employment and tax revenues to California.

Statement of Benefit to California (provided by applicant)

We propose to build a bank of HLA-homozygous human parthenogenetic stem cell lines that carry different common immune types (called HLA haplotypes) allowing the cells to be immune-matched to millions of people of differing sexes and racial backgrounds. The cell bank will be a California-based resource used in research and therapeutic applications worldwide in cases where stem cells or their derivatives are studied or needed to cure disease.

This project will benefit California through the use of proven technology to create a unique bank of stem cells to solve one of the most serious problem in regenerative medicine, that of immune rejection. This California-based resource will have the potential to relieve the suffering of millions of people worldwide because it will allow clinicians to match cells to their patients and treat a large patient base for many different diseases. Intellectual property will be created for California through this resource, resulting in increased valuation for California research organizations and the associated increase in investment, employment and tax revenues. The bank will provide cells that will be more ethically acceptable because they do not require the use of fertilized embryos. Finally, this bank will reduce the need to obtain human oocytes in the long run because once established, it will reduce the need to constantly obtain human oocytes for the creation of “patient-specific” stem cells though methods such as nuclear transfer.

California scientists now working intensively with human stem cells because of their great potential for use in regenerative medicine to cure many intractable diseases including diabetes, liver disease, blindness, Alzheimer’s disease, muscular dystrophy, Parkinson’s disease, to name but a few. The goal is the derivation of human parthenogenetic HLA homozygous stem cell lines with the different HLA haplotypes found commonly within the U.S. Several of these lines have been created and proof of principle is established. One of these lines carries the most common HLA haplotype within the U.S. population. This line has a haplotype that can be immune-matched to tens of millions of Americans across different racial groups. We propose to derive additional HLA-homozygous lines with a goal of covering the entire population with special attention to being sure all racial groups are included.

Established informed consent procedures will be used and all regulatory requirements for HHS Policy for Protection of Human Research Subjects (45CFR46) and the National Academy of Sciences Guidelines for the Human Embryonic Stem Cell Research will be followed in obtaining oocytes. All facilities will be inspected by the appropriate agencies as required. All HLA-homozygous lines will be created and banked under cGMP conditions in a manner that minimizes exposure to animal components and will be tested to establish them as clinical grade.

Review

The overall aim of this project is the derivation of clinical grade HLA-homozygous human stem cell lines using parthenogenetic activation of oocytes. With the proper selection of oocyte donors with regard to human leukocyte antigen (HLA) haplotypes, it should be possible to generate a bank of human parthenogenetic HLA-homozygous stem cell (hpSC-Hhom) lines with different haplotypes. Significance of the creation of hpSC-Hhom lines lies in the possibility of enhancing immune-matching with higher proportions of the population, thereby potentially facilitating the transplantation of cellular derivatives of such lines by reducing the potential for rejection by immune-mediated mechanisms.

The principal investigator (PI) and his/her team are experts in deriving human parthenogenetic stem cell lines as evidenced by their publications. They propose to derive two or three hpSC-Hhom lines carrying different common HLA haplotypes every year over the period of three years. These lines will be derived from normal oocytes from donors undergoing in vitro fertilization (IVF). The investigators propose to carry out derivation, expansion and storage of HLA-homozygous stem cells under current Good Manufacturing Practices (cGMP) conditions. The cell lines will be characterized for embryonic stem cell markers. Their differentiation ability will be assessed by formation of embryoid bodies and their teratoma formation ability will be assessed in immunodeficient mice. However, reviewers expressed concern that based on murine studies the differentiation potential of parthenogenetic cells is rather limited, and the analyses proposed in the application to assess cell line differentiation potential were considered rudimentary.

Reviewers felt that the PI overestimated the value of HLA matching in avoiding graft rejection. The PI has not considered the contribution of minor histocompatibility antigens in tissue rejection. For example, it was pointed out that even genotypically HLA identical bone marrow transplants are rejected due to the presence of multiple minor histoincompatibilities between the donor and recipient unless immunosuppression is given. Moreover, the HLA complex is extremely polymorphic and even the most common haplotypes are shared by only a small fraction of the population. The applicant did not propose any proof of concept studies in the proposal to test if cell lines derived from parthenogenetic embryos will overcome problems associated with HLA mismatch. From a feasibility point of view there was also a concern whether the PI will be able to consent enough volunteers needed to screen for common HLA haplotypes and who would have sufficient excess oocytes for the study. Finally, there was concern that inadequate resources were allocated to the most labor intensive aspects of the proposal, line derivation and cell culture. Because of these concerns, overall enthusiasm for this proposal was low.

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:

• Heimfeld, Shelly
• Strom, Stephen