Serum-free and xenogen-free human cardiac explant-derived stem cells and uses and methods for the production thereof
Inventors
Stewart, Duncan J. • Courtman, David • Mount, Seth • Davis, Darryl
Assignees
Publication Number
US-11083756-B2
Publication Date
2021-08-10
Expiration Date
2037-05-30
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Abstract
Methods for generating serum-free and/or xenogen-free cardiac explant-derived stem cells (EDC) are provided. These methods may include providing an initial cardiac explant, which has been minced and digested; plating the initial cardiac explant; culturing the plated cardiac explant in serum-free and xenogen-free medium; harvesting EDC cells surrounding or emerging from the plated cardiac explant; and optionally performing static expansion of harvested EDC cells in serum-free and xenogen-free media. Serum-free and/or xenogen-free cardiac EDC cells produced by these methods, as well as methods and uses thereof for the treatment of heart failure in a subject in need thereof, are also provided.
Core Innovation
The invention provides methods for generating serum-free and/or xenogen-free cardiac explant-derived stem cells (EDC). These methods involve obtaining an initial cardiac explant, optionally from atrial appendages or myocardial, atrial, or ventricular biopsy, mincing and digesting the explant with collagenase, plating the explant on a cell culture plate, culturing the plated cardiac explant in serum-free and xenogen-free medium, harvesting EDC cells emerging from the explant with trypsin, and optionally performing static expansion of harvested EDC cells in serum-free and xenogen-free media.
The problem being solved is that traditional culture media for cardiac explant-derived stem cells typically include ill-defined or xenobiotic components such as fetal bovine serum, which present variability and risks of infectious or toxic contaminants, creating barriers to clinical translation. Moreover, prior methods altering EDC culture conditions have often impaired the regenerative performance of these cells, limiting their therapeutic potential.
By transitioning to serum-free and xenogen-free culture conditions, the invention addresses the need for clinically compatible cardiac-derived cell products with improved standardization and safety. The methods provide EDC cells that are smaller, more homogeneous, with maintained or enhanced regenerative potential for treating heart failure. Furthermore, the invention includes preparations of these cells for clinical use and their administration by intra-myocardial or intra-coronary injection.
Claims Coverage
The patent includes one independent claim covering a method for transitioning cardiac explant-derived stem cells to serum-free and xenogen-free culture conditions, detailing key steps and culture conditions.
Method for transitioning cardiac explant-derived stem cells to serum-free and xenogen-free culture conditions
A method comprising providing an initial cardiac explant from atrial appendages or ventricular/atrial biopsy, mincing and digesting with collagenase; plating the explant on a cell culture plate; culturing in serum-free and xenogen-free medium; harvesting EDC cells emerging from the explant using trypsin; and performing static expansion of harvested EDC cells on fibronectin-coated cultureware in serum-free and xenogen-free medium, without sphering of cells.
Use of GMP-grade materials and specific reagents
Collagenase used is collagenase I/II, cell culture plate and cultureware are fibronectin-coated, media is serum-free and xenogen-free Nutristem XF, trypsin is TrypLE Select, and these components can be GMP-grade for clinical compatibility.
Culture conditions and supplementation
Culturing can be performed at physiological oxygen tension (~5%) or ambient oxygen (~21%); initial culture may include serum supplementation (~2%) for about 48 hours or more, followed by full replacement with serum-free and xenogen-free medium.
Cell expansion, harvesting repetition, and clinical preparation
Static expansion of harvested cells is performed for about 7 days; harvesting and expansion steps may be repeated up to five times with cells cryogenically stored and pooled for administration as single or multiple intra-myocardial or intra-coronary injections. The harvested cells may be autologous and intended for clinical use.
Use of produced serum-free and xenogen-free cardiac explant-derived cells
The serum-free and xenogen-free EDC cells produced by the method can be used for treating heart failure and for repairing or regenerating cardiac tissue in a subject by transplantation via intra-myocardial or intra-coronary injection.
Provision of kits
Kits comprising serum-free and xenogen-free human cardiac explant-derived cells along with at least one of serum-free and xenogen-free medium, tools for injection, collagenase, cell culture plate, trypsin, cultureware, vessels, differentiating factors, or instructions for culturing and injecting the cells.
In summary, the independent claim presents a comprehensive method for producing serum-free and xenogen-free cardiac explant-derived stem cells using defined GMP-grade materials and conditions. It covers culture, harvesting, optional expansion, storage, preparation for clinical use, and therapeutic application to treat heart failure by delivering these cells into cardiac tissue.
Stated Advantages
Serum-free and xenogen-free culture conditions produce a smaller, more homogeneous cell product with stable phenotypic profiles compared to serum cultures.
The method supports ex vivo proliferation of cardiac explant-derived stem cells from multiple tissue sources with negligible effects on phenotype or regenerative capacity.
Serum-free cultured cells retain paracrine activity important for cardiac repair and exhibit enhanced therapeutic efficacy in a mouse model of myocardial infarction compared to serum-cultured cells.
The approach avoids ill-defined or xenobiotic components such as fetal bovine serum, reducing variability and potential contamination risks for clinical translation.
Cells remain viable after clinical catheter-based delivery methods and brief storage, facilitating clinical implementation.
Documented Applications
Treatment of heart failure in subjects in need thereof by transplantation of serum-free and xenogen-free human cardiac explant-derived cells.
Repairing and/or regenerating cardiac tissue, specifically myocardium, in subjects by intra-myocardial or intra-coronary injection of serum-free and xenogen-free cardiac explant-derived stem cells.
Use of serum-free and xenogen-free cardiac explant-derived stem cells in clinical settings, including kits for cell delivery and compositions with culture media and differentiating factors.
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