Live Cell Surface-Based One-Step FucT Conjugate Technology

Employing living cells as a therapeutic drug is the development direction of future drugs. Cell-based therapies (i.e., CAR-T, TCR-T) have become attractive treatments for advanced therapeutic strategies due to their secretory function, specific homing effects, unique therapeutic potential, and ex vivo expandability. However, these methods still face multiple challenges: a) exposure of cells to harsh conditions; b) utilization of viral gene transfer; c) possibility of endogenous gene disruption; and d) permanent genetic modification.

Therefore, a simple and powerful method that directly modifies the cell surface to introduce novel functions is highly desirable. Here, Creative Biolabs developed a single-step, chemical enzyme-based approach that can quickly engineer cell surfaces in minutes. Through this fast, simple and cost-effective technique, biomacromolecules including proteins (antibodies, antigens) and nucleic acids, as well as small molecule probes such as fluorescent and biophysical probes, are efficiently conjugated without the need for genetic modification of host cells.

CellFace™ Conjugate Technology – "One Step In Minutes"

Cell Surface glycans are universal to living cells

Glycan is an important substance that constitutes a living body. There are abundant monosaccharide and oligosaccharide modifications on almost all cell surfaces and cells. They are catalyzed by a variety of glycosyltransferases.

Empowered by years of experience and expertise in glycan biochemistry, our CellFace™ conjugate technology focuses on a class of universal glycosyls that are expressed on almost all cell surfaces, which allows the wide applicability of our cell surface modification technology.

Cell face glycans

Glycosyltransferase catalyzed single-step reaction

Glycosyltransferases are glycan-processing machines that recognize different monosaccharide / oligosaccharide derivatives (conjugates donors) with activated groups and link them to specific substrates. Utilizing the substrate specificity of glycosyltransferase and its compatibility with sugar donors, it is possible to modify and functionally study specific glycosylation modifications in vivo.

e used FucT, a fucose transferase that utilizes GDP-Fucose as a sugar donor, which specifically recognizes glycosyl fragments on the cell surface and covalently attaches fucose to the ends. The enzyme has very good compatibility with the donor, and even if a sterically hindered group is introduced, it does not affect the catalytic efficiency of the enzyme. Since many cells have glycocalyx on their surface, our technique can be a powerful tool for attaching large molecules to the cell surface.

Conjugate broad substrate scope

Our CellFace™ conjugate technology utilizes fucosyltransferases (Fuc T) to transfer biomacromolecules such as IgG antibodies (MW~150 KD) to glycocalyx on the surface of living cells, as the antibody is conjugated to natural donor substrate GDP-fucose of Fuc T. This highly efficient one-step approach requires no genetic modification and hardly interferes with the endogenous function of a cell.

Given the unprecedented substrate tolerance of Fuc T, this one-step FucT-based chemoenzymatic method provides a versatile method for engineering cell surface and research applications.

Notably, IgG signals bound to the cell surface are detected after incubation with FucT and IgG. Labeling efficiency is concentration dependent, and under saturation conditions, approximately 2.5 x 105 rIgG molecules were introduced into the cell surface.

Confocal microscopy images Fig.1 Confocal microscopy images of cells treated with or without FucT when incubated with IgG.

Live Cell Surface-based Polyvalent Display Conjugate Technology Activated Alkynes-based Diverse Level Conjugation Technology FucoID-based Tumor Antigen-specific T Cell Surface Engineering Technology Live Cell Surface-based N-Glycan Editing Technology