Early post-transfection isolation of cells (EPIC) for biologics production
AssigneeGenzyme Corporation
InventorsVictor R. Cairns, Christine DeMaria, Jason Vitko
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Abstract
Provided herein are methods for selecting a population of cells expressing a target polypeptide. In some aspects, the disclosure provides methods for sorting and selecting populations of transfected host cells based on their early expression of a selectable polypeptide. In certain embodiments, the sorting is performed using fluorescence-activated cell sorting or magnetic-activated cell sorting based on the selectable polypeptide. Such selection methods can be further utilized to generate clonal populations of producer cells, e.g. for large-scale manufacturing of a target polypeptide of interest.
Claims21
A method of producing a population of producer cells expressing a target polypeptide, the method comprising:
- (a) transfecting host cells with a vector that encodes one or more mRNAs, wherein the one or more mRNAs encode a selectable polypeptide and the target polypeptide;
- (b) isolating from the transfected host cells, between two and four days after transfection, a highly positive transient expression sub-population of early-expressing transfected host cells which express the selectable polypeptide, wherein the highly positive transient sub-population of early-expressing transfected host cells comprises about the top 10 percent of transfected host cells in terms of their expression of the selectable polypeptide;
- and
- (c) expanding the isolated highly positive transient expression sub-population of early-expressing transfected host cells, thereby producing a population of producer cells expressing the target polypeptide, wherein steps (b) and (c) are each performed in drug-selection-free medium.
Dependencies are inferred from the claim text.