Patient-specific stem cell lines derived from human parthenogenetic blastocysts
First Claim
1. An isolated human stem cell line derived from parthenogeneic blastocysts, wherein at least one cell comprising the cell line isa) heterozygous for one or more single nucleotide polymorphisms (SNPs);
- orb) homozygous for one or more HLA alleles;
orc) a combination of (a) and (b).
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Abstract
Methods are disclosed for generating HLA homozygous parthenogenetic human stem cell (hpSC-Hhom) lines from both HLA homozygous and HLA heterozygous donors. These hpSC-Hhom lines demonstrate typical human embryonic stem cell morphology, expressing appropriate stem cell markers and possessing high levels of alkaline phosphatase and telomerase activity. Additionally, injection of these cell lines into immunodeficient animals leads to teratoma formation. Furthermore, in the case of HLA heterozygous donors, the hpSC-Hhom lines inherit the haplotype from only one of the donor'"'"'s parents. SNP data analysis suggests that hpSC-Hhom lines derived from HLA heterozygous oocyte donors are homozygous throughout the genome as assessed by single-nucleotide polymorphism (SNP) analysis. The protocol as disclosed minimizes the use of animal-derived components, which makes the stem cells more practical for clinical application.
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Citations
30 Claims
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1. An isolated human stem cell line derived from parthenogeneic blastocysts, wherein at least one cell comprising the cell line is
a) heterozygous for one or more single nucleotide polymorphisms (SNPs); - or
b) homozygous for one or more HLA alleles;
orc) a combination of (a) and (b). - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12)
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- 13. A library of stem cells comprising autologous or allogenic stem cells, wherein the stem cells are derived from parthenogenetically activated oocytes from one or more human donors, and wherein the stem cells are HLA homozygous stem cells.
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26. A method of generating HLA homozygous stem cells comprising:
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a) screening oocyte donors for HLA-haplotypes found commonly in a given population group; b) incubating human metaphase II oocytes in in vitro fertilization (IVF) media; c) incubating the cells of step (b) in IVF media comprising an ionophore; d) incubating the cells of step (c) in IVF media comprising a puromycin; and e) incubating the cells of step (d) in fresh IVF medium under low O2 tension, wherein the incubating steps (b)-(d) are carried out under high O2 tension and wherein inner cell masses (ICM) obtained from cells at step (d) produce culturable stem cells. - View Dependent Claims (27)
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- 28. A cell bank comprising HLA-homozygous stem cells, wherein the stem cells are derived from parthenogenetically activated oocytes from HLA-homozygous or HLA-heterozygous human oocyte donors.
Specification