Real-time quantitative PCR analysis of mouse dendritic cells treated with Ac4ManNAz or PBS

Dendritic cell (DC) vaccine was among the first FDA-approved cancer immunotherapies, but has been limited by the modest cytotoxic T lymphocyte (CTL) response and therapeutic efficacy. Here we report a facile metabolic labeling approach that enables targeted modulation of adoptively transferred DCs for developing enhanced DC vaccines. We show that metabolic glycan labeling can reduce the membrane mobility of DCs, which activates DCs and improves the antigen presentation and subsequent T cell priming property of DCs. Metabolic glycan labeling itself can enhance the antitumor efficacy of DC vaccines. In addition, the cell-surface chemical tags (e.g., azido groups) introduced via metabolic glycan labeling also enable in vivo conjugation of cytokines onto adoptively transferred DCs, which further enhances CTL response and antitumor efficacy. Our DC labeling and targeting technology provides a new strategy to improve the therapeutic efficacy of DC vaccines, with minimal interference upon the clinical manufacturing process. Overall design: Mouse dendritic cells that were treated with Ac4ManNAz or PBS (1 million cells per sample) were collected, washed, frozen at -80oC, and transferred to the sequencing Core at UIUC. The staff then extract RNA and treat it with DNAse, run it on a fragment analyzer so that you get the RIN, make individually barcoded polyA+ selected RNAseq libraries, sequence them on an SP lane with 1x100nt single-reads, which produces ~ 400 million reads