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Assignees
Paracrine Therapeutics Pte Ltd
MemberParacrineParacrineParacrine develops regenerative therapies for chronic wound care, leveraging autologous cell-based treatments and real-time AI platforms to enhance healing precision, predictability, and efficiency. The company operates globally, focusing on scalable and cost-effective biological solutions that integrate advanced machine learning for improved patient outcomes.
Paracrine develops regenerative therapies for chronic wound care, leveraging autologous cell-based treatments and real-time AI platforms to enhance healing precision, predictability, and efficiency. The company operates globally, focusing on scalable and cost-effective biological solutions that integrate advanced machine learning for improved patient outcomes.
Publication Number
US-9018005-B2
Publication Date
2015-04-28
Expiration Date
Abstract
We disclose a method comprising: (a) providing an embryonic stem (ES) cell; and (b) establishing a progenitor cell line from the embryonic stem cell; in which the progenitor cell line is selected based on its ability to self-renew. Preferably, the method selects against somatic cells based on their inability to self-renew. Preferably, the progenitor cell line is derived or established in the absence of co-culture, preferably in the absence of feeder cells, which preferably selects against embryonic stem cells. Optionally, the method comprises (d) deriving a differentiated cell from the progenitor cell line.
Core Innovation
The invention describes deriving lineage-restricted, self-renewing progenitor cell lines from embryonic stem cells, exemplified by E-RoSH endothelial progenitor lines (mouse) and huES9.E MSC-like lines (human). The derived lines exhibit stable karyotype and long-term culture longevity exceeding forty generations, loss of pluripotency markers including alkaline phosphatase and OCT4, absence of teratoma formation, enriched lineage marker expression, and functional differentiation in lineage-specific assays.
The invention provides a method to derive lineage-restricted, self-renewing progenitor cell lines by selecting for cells that self-renew. The method biases differentiation, disaggregates embryonic stem cell derivatives, and cultures cells under conditions that select for self-renewal while excluding factors that promote embryonic stem cell pluripotency, followed by serial enrichment and cloning of progenitor populations [procedural detail omitted for safety].
The invention addresses obtaining mesenchymal progenitor cells from parental embryonic stem cells that self-renew yet are lineage restricted compared to the parental embryonic stem cell and are maintainable in cell culture for extended generations. The derived lines do not undergo transformation, display reduced or absent expression of specified pluripotency markers, maintain stable karyotype and gene expression, and retain functional lineage-associated properties with capacity for downstream differentiation and use in therapeutic and screening contexts.
Claims Coverage
The independent claim defines one method with five main inventive features.
Providing parental human embryonic stem cells or descendants
Providing the parental embryonic stem cell or descendants of the parental embryonic stem cell obtained by [procedural detail omitted for safety], wherein the parental embryonic stem cell is a human embryonic stem cell.
Feeder-free culture in rich media with serum or serum replacement
Culturing the parental embryonic stem cell in the absence of feeder cells in rich media comprising essential nutrients and serum or serum replacement.
Absence of growth regulators that promote embryonic stem cell growth
Wherein the rich media does not comprise additional growth regulators or hormones that promote growth of embryonic stem cells.
Production of mesenchymal progenitor cells which self-renew
The culturing produces mesenchymal progenitor cells which self-renew.
Establishing a non-transformed, long-term maintainable, lineage-restricted cell line
Thereby establishing in the absence of transformation a mesenchymal progenitor cell line that is maintainable in cell culture for more than 20 generations and is lineage restricted compared to the parental embryonic stem cell.
The independent claim centers on providing parental embryonic stem cells or their descendants and culturing them feeder-free in rich media lacking growth regulators to produce self-renewing mesenchymal progenitor cells and thereby establish a non-transformed, lineage-restricted progenitor cell line maintainable for more than 20 generations.
Stated Advantages
Generation of mesenchymal progenitor cells that self-renew.
Establishment of cell lines maintainable in cell culture for more than 20 generations (and examples exceeding 40 generations).
Derivation of lineage-restricted progenitor cell lines compared to the parental embryonic stem cell.
Non-transformed, non-tumorigenic cell lines with lack of substantial teratoma formation in recipient animals.
Reduced or absent expression of pluripotency markers (for example alkaline phosphatase, OCT4, BMP4, FGF5, Sox-2, Utf1).
Stable karyotype and stability of chromosome number and gene expression pattern over long-term culture.
Documented Applications
Derivation and culture of E-RoSH endothelial progenitor cell lines (mouse) and huES9.E MSC-like cell lines (human).
In vitro endothelial functional assays [procedural detail omitted for safety] as demonstrations of lineage-specific function.
In vitro MSC lineage differentiation assays including adipocyte and osteocyte differentiation as demonstrations of MSC-like potential.
Characterization and monitoring using [procedural detail omitted for safety].
Assessment of tumorigenicity by lack of teratoma formation in recipient animals [procedural detail omitted for safety] as a documented evaluation of in vivo behavior.
Drug screening
Regenerative therapy and disease treatment
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