Compositions and methods for production of exofucosylated cells for clinical applications
OwnerRobert Sacksteininventor
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Abstract
The present disclosure provides, inter alia, compositions and methods for detecting changes in level of expression of cell-surface Type 2 terminal lactosamines on a population of cultured cells propagated under different conditions. The disclosure also provides compositions and methods for enforcing stably expressed glycans on human cells. In certain embodiments, the compositions and/or methods utilize one or more members of the α(1,3)-fucosyltransferase family. In certain embodiments, glycoengineered CD44 glycosylated product (e.g. HCELL) is stable for at least 48 hours at 4° C., with retained expression after cell cryopreservation.
Claims18
A method of detecting a change of expression in cell-surface Type 2 terminal lactosamines on a population of cultured cells comprising the steps of:
- (a) contacting the cells with a glycosyltransferase and a donor nucleotide sugar, wherein the glycosyltransferase and donor nucleotide sugar are effective to enforce expression of a glycan;
- and
- (b) detecting the product glycan on the cells,
- wherein the contacting of step (a) is performed before and/or after any culture condition modification.
A method of detecting differences in level of expression of cell-surface Type 2 terminal lactosamines on a population of cultured cells propagated under different conditions comprising the steps of:
- (a) culturing a first population of cells under a first culture condition;
- (b) culturing a second population of cells under a second (different) culture condition;
- (c) contacting the first and second population of cells with a glycosyltransferase and a donor nucleotide sugar, wherein the glycosyltransferase and donor nucleotide sugar are effective to enforce expression of a glycan;
- and
- (d) detecting the glycan on the first and second population of cells.
A process for producing GMP-grade exofucosylated cells comprising:
- (a) providing cells with a culture medium comprising a supplement, wherein the cells comprise cell surface CD44 and the supplement is effective to maintain or increase the amount of a CD44 glycoform comprising sialylated Type 2 lactosamines;
- (b) expanding the cells in the culture medium;
- and
- (c) contacting the cells with a glycosyltransferase and a donor nucleotide sugar that are effective to enforce expression of the HCELL glycoform of CD44 on the cells.
Dependencies are inferred from the claim text.