Genes that are up- or down-regulated during differentiation of human embryonic stem cells
First Claim
1. A method for assessing a culture of undifferentiated primate pluripotent stem (pPS) cells or their progeny, comprising detecting or measuring expression of two or more of the markers in any of Tables 5 to 9, other than hTERT or Oct 3/4.
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Accused Products
Abstract
Genes that are up- or down-regulated during differentiation provide important leverage by which to characterize and manipulate early-stage pluripotent stem cells. Over 35,000 unique transcripts have been amplified and sequenced from undifferentiated human embryonic stem cells, and three types of differentiated progeny. Statistical analysis of the assembled transcripts identified genes that alter expression levels as differentiation proceeds. The expression profile provides a marker system that has been used to identify particular culture components for maintaining the undifferentiated phenotype. The gene products can also be used to promote differentiation; to assess other relatively undifferentiated cells (such as cancer cells); to control gene expression; or to separate cells having desirable characteristics. Manipulation of particular genes can be used to forestall or focus the differentiation process, en route to producing a specialized homogenous cell population suitable for human therapy.
158 Citations
48 Claims
- 1. A method for assessing a culture of undifferentiated primate pluripotent stem (pPS) cells or their progeny, comprising detecting or measuring expression of two or more of the markers in any of Tables 5 to 9, other than hTERT or Oct 3/4.
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7. A method for assessing a culture of undifferentiated primate pluripotent stem (pPS) cells or their progeny, comprising detecting or measuring:
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a marker from the following list;
Cripto, gastrin-releasing peptide (GRP) receptor, and podocalyxin-like protein; and
either hTERT and/or Oct 3/4, or a second marker from the list. - View Dependent Claims (8, 9, 10, 20)
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- 21. A method for assessing the growth characteristics of a cell population, comprising detecting or measuring expression of two or more of the markers in any of Tables 5 to 9, at least one of which is neither hTERT nor Oct 3/4.
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27. A method for maintaining pPS cells in a pluripotent state, comprising causing them to express one of the following markers at a higher level:
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Forkhead box O1A (FOXO1A);
Zic family member 3 (ZIC3);
Hypothetical protein FLJ20582;
Forkhead box H1 (FOXH1);
Zinc finger protein, Hsal2;
KRAB-zinc finger protein SZF1-1;
or Zinc finger protein of cerebellum ZIC2;
orany other marker listed in Table 5 with the symbol “
{circle over (x)}”
. - View Dependent Claims (28)
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29. A method for causing pPS cells to differentiate into a particular tissue type, comprising causing them to express one of the following markers at a lower level:
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Forkhead box O1A (FOX01A);
Zic family member 3 (ZIC3);
Hypothetical protein FLJ20582;
Forkhead box H1 (FOXH1);
Zinc finger protein, Hsal2;
KRAB-zinc finger protein SZF1-1;
or Zinc finger protein of cerebellum ZIC2;
orany other maker listed in Table 5 with the symbol “
{circle over (x)}”
;
or by causing them to express one of the markers listed in Table 6 with the symbol “
{circle over (x)}”
at a higher level. - View Dependent Claims (30)
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31. A method for maintaining pPS cells in a pluripotent state, comprising culturing pPS cells or their progeny in the presence of a normally secreted protein that is encoded by a gene listed in Table 2, 5, 7, or 9.
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32. A method for causing pPS cells to differentiate, comprising culturing pPS cells or their progeny in the presence of a normally secreted protein that is encoded by a gene listed in Table 3, 6, or 8.
- 33. A method for causing an encoding sequence to be preferentially expressed in undifferentiated pPS cells, comprising genetically altering pPS cells with the encoding sequence under control of a promoter for one of the markers listed in Table 2, 5, or 7.
- 36. A method for causing an encoding sequence to be preferentially expressed in differentiated cells, comprising genetically altering the pPS cells with the encoding sequence under control of a promoter for one of the markers listed in Table 3, 6, or 8.
- 39. A method for sorting differentiated cells from less differentiated cells, comprising separating cells expressing a surface marker in any of Tables 5 to 9 from cells not expressing the marker.
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42. A method for causing pPS cells to proliferate without differentiation, comprising culturing them with mesenchymal stem cells.
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43. A method for identifying genes that are up- or down-regulated during differentiation of pPS cells, comprising:
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a) sequencing transcripts in an expression library from undifferentiated pPS cells;
b) sequencing transcripts in one or more expression libraries from one or more cell types that have differentiated from the same line of pPS cells;
c) determining the frequency of transcripts from each gene sequenced in each of the libraries; and
d) identifying the gene as being up- or down-regulated during differentiation of the pPS cells if the frequency of transcripts in the library from the undifferentiated pPS cells is statistically different from the frequency of transcripts in one or more libraries from the differentiated cell types. - View Dependent Claims (44)
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Specification