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Optimized Protocol for Efficient Transfection of Dendritic Cells without Cell Maturation
Published on: July 8, 2011
Modification of human embryonic stem cell-derived dendritic cells with mRNA for efficient antigen presentation and
Kevin P Nishimoto1, Su-Yi Tseng, Jane S Lebkowski
1Geron Corporation, 230 Constitution Drive, Menlo Park, CA 94025, USA. knishimoto@geron.com
Aim:
Dendritic cell (DC)-based vaccines are designed to exploit the intrinsic capacity of these highly effective antigen presenting cells to prime and boost antigen-specific T-cell immune responses. Successful development of DC-based vaccines will be dependent on the ability to utilize and harness the full potential of these potent immune stimulatory cells. Recent advances to generate DCs derived from human embryonic stem cells (hESCs) that are suitable for clinical use represent an alternative strategy from conventional approaches of using patient-specific DCs. Although the differentiation of hESC-derived DCs in serum-free defined conditions has been established, the stimulatory potential of these hESC-derived DCs have not been fully evaluated.
Methods:
hESC-derived DCs were differentiated in serum-free defined culture conditions. The delivery of antigen into hESC-derived DCs was investigated using mRNA transfection and replication-deficient adenoviral vector transduction. hESC-derived DCs modified with antigen were evaluated for their capacity to stimulate antigen-specific T-cell responses with known HLA matching. Since IL-12 is a key cytokine that drives T-cell function, further enhancement of DC potency was evaluated by transfecting mRNA encoding the IL-12p70 protein into hESC-derived DCs.
Results:
The transfection of mRNA into hESC-derived DCs was effective for heterologous protein expression. The efficiency of adenoviral vector transduction into hESC-derived DCs was poor. These mRNA-transfected DCs were capable of stimulating human telomerase reverse transcriptase antigen-specific T cells composed of varying degrees of HLA matching. In addition, we observed the transfection of mRNA encoding IL-12p70 enhanced the T-cell stimulation potency of hESC-derived DCs.
Conclusion:
These data provide support for the development and modification of hESC-derived DCs with mRNA as a potential strategy for the induction of T-cell-mediated immunity.
