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Design of an optimized Wilms' tumor 1 (WT1) mRNA construct for enhanced WT1 expression and improved immunogenicity in vitro and in vivo

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Abstract
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.
Keywords
ACUTE MYELOID-LEUKEMIA, MHC CLASS-I, ELECTROPORATED DENDRITIC CELLS, CYTOTOXIC T-LYMPHOCYTES, THERAPEUTIC VACCINATION, ANTIGEN PRESENTATION, MELANOMA PATIENTS, STIMULATORY CAPACITY, NUCLEAR-LOCALIZATION, DIRECT-INJECTION

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Chicago
Benteyn, Daphné, Sébastien Anguille, Sandra Van Lint, Carlo Heirman, An MT Van Nuffel, Jurgen Corthals, Sebastian Ochsenreither, et al. 2013. “Design of an Optimized Wilms’ Tumor 1 (WT1) mRNA Construct for Enhanced WT1 Expression and Improved Immunogenicity in Vitro and in Vivo.” Molecular Therapy-nucleic Acids 2.
APA
Benteyn, D., Anguille, S., Van Lint, S., Heirman, C., Van Nuffel, A. M., Corthals, J., Ochsenreither, S., et al. (2013). Design of an optimized Wilms’ tumor 1 (WT1) mRNA construct for enhanced WT1 expression and improved immunogenicity in vitro and in vivo. MOLECULAR THERAPY-NUCLEIC ACIDS, 2.
Vancouver
1.
Benteyn D, Anguille S, Van Lint S, Heirman C, Van Nuffel AM, Corthals J, et al. Design of an optimized Wilms’ tumor 1 (WT1) mRNA construct for enhanced WT1 expression and improved immunogenicity in vitro and in vivo. MOLECULAR THERAPY-NUCLEIC ACIDS. 2013;2.
MLA
Benteyn, Daphné, Sébastien Anguille, Sandra Van Lint, et al. “Design of an Optimized Wilms’ Tumor 1 (WT1) mRNA Construct for Enhanced WT1 Expression and Improved Immunogenicity in Vitro and in Vivo.” MOLECULAR THERAPY-NUCLEIC ACIDS 2 (2013): n. pag. Print.
@article{6963162,
  abstract     = {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.},
  articleno    = {e134},
  author       = {Benteyn, Daphn{\'e} and Anguille, S{\'e}bastien and Van Lint, Sandra and Heirman, Carlo and Van Nuffel, An MT and Corthals, Jurgen and Ochsenreither, Sebastian and Waelput, Wim and Van Beneden, Katrien and Breckpot, Karine and Van Tendeloo, Viggo and Thielemans, Kris and Bonehill, Aude},
  issn         = {2162-2531},
  journal      = {MOLECULAR THERAPY-NUCLEIC ACIDS},
  keyword      = {ACUTE MYELOID-LEUKEMIA,MHC CLASS-I,ELECTROPORATED DENDRITIC CELLS,CYTOTOXIC T-LYMPHOCYTES,THERAPEUTIC VACCINATION,ANTIGEN PRESENTATION,MELANOMA PATIENTS,STIMULATORY CAPACITY,NUCLEAR-LOCALIZATION,DIRECT-INJECTION},
  language     = {eng},
  pages        = {9},
  title        = {Design of an optimized Wilms' tumor 1 (WT1) mRNA construct for enhanced WT1 expression and improved immunogenicity in vitro and in vivo},
  url          = {http://dx.doi.org/10.1038/mtna.2013.54},
  volume       = {2},
  year         = {2013},
}

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