Tumor antigen-encoding
mRNA for dendritic cell (DC)-based vaccination has gained increasing popularity in recent years. Within this context, two main strategies have entered the clinical trial stage: the use of
mRNA for ex vivo
antigen loading of DCs and the direct application of
mRNA as a source of
antigen for DCs in vivo. DCs transfected with
mRNA-encoding
Wilms' tumor 1 (
WT1) protein have shown promising clinical results. Using a stepwise approach, we re-engineered a WT1
cDNA-carrying transcription vector to improve the translational characteristics and immunogenicity of the transcribed
mRNA. Different modifications were performed: (i) the WT1 sequence was flanked by the lysosomal targeting sequence of dendritic cell
lysosomal-associated membrane protein to enhance cytoplasmic expression; (ii) the nuclear localization sequence (NLS) of WT1 was deleted to promote shuttling from the nucleus to the cytoplasm; (iii) the WT1 DNA sequence was optimized in silico to improve translational efficiency; and (iv) this WT1 sequence was cloned into an optimized
RNA transcription vector. DCs electroporated with this optimized
mRNA showed an improved ability to stimulate WT1-specific T-cell immunity. Furthermore, in a murine model, we were able to show the safety, immunogenicity, and therapeutic activity of this optimized
mRNA. This work is relevant for the future development of improved
mRNA-based
vaccine strategies K.Molecular
Therapy-
Nucleic Acids (2013) 2, e134; doi:10.1038/mtna.2013.54; published online 19 November 2013.