期刊
MOLECULAR THERAPY-METHODS & CLINICAL DEVELOPMENT
卷 7, 期 -, 页码 132-145出版社
CELL PRESS
DOI: 10.1016/j.omtm.2017.10.003
关键词
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资金
- BioCanRx (Biotherapeutics for Cancer Treatment - Networks of Centres of Excellence)
- Genome BC
- BC Cancer Foundation
- Canadian Cancer Society [703329]
- VCH-CIHR-UBC MD/PhD Studentship Award
- CIHR Frederick Banting and Charles Best Canada Graduate Scholarship Doctoral Research Award
There is great potential for engineering cellular therapeutics by repurposing biological systems. Here, we report utilization of the granzyme-perforin pathway of cytotoxic lymphocytes as a cell-to-cell protein delivery module. We designed and constructed granzyme B-derived chaperone molecules fused to a fluorescent protein payload and expressed these constructs in natural killer (NK) cells. Using confocal microscopy and flow cytometry, we investigated the co-localization of the chaperones with lytic granules and the chaperone-mediated transfer of the fluorescent protein payload from NK to target cells in co-culture experiments. A synthetic chaperone consisting of the granzyme B ER signal peptide and a domain encompassing putative N-linked glycosylation sites in granzyme B is insufficient for payload transfer to target cells, whereas full-length granzyme B is sufficient for payload delivery. Combining our functional data with an analysis of the crystal structure of granzyme B suggests that the necessary motifs for granzyme B loading into lytic granules are dispersed throughout the primary amino acid sequence and are only functional when contiguous in the tertiary structure. These results illustrate that by using granzyme B as a molecular chaperone the granzyme-perforin pathway can be exploited as a programmable molecular delivery system for cell-based therapies.
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