Enhanced dendritic cell antigen presentation in RNA-based immunotherapy

Matthew F. Kalady, Mark W. Onaitis, Karen M. Padilla, Sirisha Emani, Douglas Tyler, Scott K. Pruitt

Research output: Contribution to journalArticle

51 Citations (Scopus)

Abstract

Background. Dendritic cells pulsed with mRNA provide a unique approach to tumor immunotherapy. We hypothesized that increased mRNA transfection efficiency and dendritic cell maturation would improve antigen processing and presentation as well as T-cell costimulation, resulting in enhanced induction of antimelanoma immune responses. Methods. Immature monocyte-derived dendritic cells were transfected with mRNA by passive pulsing, lipofection, or electroporation. Dendritic cells were either left untreated or matured using the double-stranded RNA poly(I:C). T-Cell cultures were generated by stimulation of naïve T-cells with each set of dendritic cells. Specific antigen presentation and specific effector T-cell generation were analyzed by an IFN-γ release Elispot assay. Results. Greatest intracellular green fluorescent protein was observed by flow cytometry following dendritic cell electroporation with green fluorescent protein mRNA. DC presentation of Mart-1/Melan A peptide, as measured by Elispot assay using a specific T-cell clone, was greatest following transfection with Mart-1/Melan A mRNA by electroporation. Maturation of dendritic cells further improved antigen presentation regardless of transfection technique. Specific Mart-1/Melan A effector T cells were produced after culture of naïve T cells with dendritic cells that were electroporated with Mart-1/Melan A mRNA and then matured, but not for dendritic cells that remained immature. Conclusions. Efficient mRNA transfection by electroporation as well as dendritic cell maturation results in increased levels of Mart-1/Melan A antigen presentation and enhanced production of antigen-specific effector T cells. This combination of strategies may be used to enhance immune responses to RNA-based dendritic cell vaccines.

Original languageEnglish
Pages (from-to)17-24
Number of pages8
JournalJournal of Surgical Research
Volume105
Issue number1
DOIs
StatePublished - 2002
Externally publishedYes

Fingerprint

Antigen Presentation
Immunotherapy
Dendritic Cells
RNA
MART-1 Antigen
T-Lymphocytes
Electroporation
Messenger RNA
Transfection
Green Fluorescent Proteins
Double-Stranded RNA
Monocytes
Flow Cytometry
Vaccines
Clone Cells
Cell Culture Techniques
Antigens
Peptides

Keywords

  • Antigen presentation
  • Dendritic cells
  • Immunotherapy
  • mRNA-based vaccines

ASJC Scopus subject areas

  • Surgery

Cite this

Kalady, M. F., Onaitis, M. W., Padilla, K. M., Emani, S., Tyler, D., & Pruitt, S. K. (2002). Enhanced dendritic cell antigen presentation in RNA-based immunotherapy. Journal of Surgical Research, 105(1), 17-24. https://doi.org/10.1006/jsre.2002.6435

Enhanced dendritic cell antigen presentation in RNA-based immunotherapy. / Kalady, Matthew F.; Onaitis, Mark W.; Padilla, Karen M.; Emani, Sirisha; Tyler, Douglas; Pruitt, Scott K.

In: Journal of Surgical Research, Vol. 105, No. 1, 2002, p. 17-24.

Research output: Contribution to journalArticle

Kalady, MF, Onaitis, MW, Padilla, KM, Emani, S, Tyler, D & Pruitt, SK 2002, 'Enhanced dendritic cell antigen presentation in RNA-based immunotherapy', Journal of Surgical Research, vol. 105, no. 1, pp. 17-24. https://doi.org/10.1006/jsre.2002.6435
Kalady, Matthew F. ; Onaitis, Mark W. ; Padilla, Karen M. ; Emani, Sirisha ; Tyler, Douglas ; Pruitt, Scott K. / Enhanced dendritic cell antigen presentation in RNA-based immunotherapy. In: Journal of Surgical Research. 2002 ; Vol. 105, No. 1. pp. 17-24.
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AU - Pruitt, Scott K.

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N2 - Background. Dendritic cells pulsed with mRNA provide a unique approach to tumor immunotherapy. We hypothesized that increased mRNA transfection efficiency and dendritic cell maturation would improve antigen processing and presentation as well as T-cell costimulation, resulting in enhanced induction of antimelanoma immune responses. Methods. Immature monocyte-derived dendritic cells were transfected with mRNA by passive pulsing, lipofection, or electroporation. Dendritic cells were either left untreated or matured using the double-stranded RNA poly(I:C). T-Cell cultures were generated by stimulation of naïve T-cells with each set of dendritic cells. Specific antigen presentation and specific effector T-cell generation were analyzed by an IFN-γ release Elispot assay. Results. Greatest intracellular green fluorescent protein was observed by flow cytometry following dendritic cell electroporation with green fluorescent protein mRNA. DC presentation of Mart-1/Melan A peptide, as measured by Elispot assay using a specific T-cell clone, was greatest following transfection with Mart-1/Melan A mRNA by electroporation. Maturation of dendritic cells further improved antigen presentation regardless of transfection technique. Specific Mart-1/Melan A effector T cells were produced after culture of naïve T cells with dendritic cells that were electroporated with Mart-1/Melan A mRNA and then matured, but not for dendritic cells that remained immature. Conclusions. Efficient mRNA transfection by electroporation as well as dendritic cell maturation results in increased levels of Mart-1/Melan A antigen presentation and enhanced production of antigen-specific effector T cells. This combination of strategies may be used to enhance immune responses to RNA-based dendritic cell vaccines.

AB - Background. Dendritic cells pulsed with mRNA provide a unique approach to tumor immunotherapy. We hypothesized that increased mRNA transfection efficiency and dendritic cell maturation would improve antigen processing and presentation as well as T-cell costimulation, resulting in enhanced induction of antimelanoma immune responses. Methods. Immature monocyte-derived dendritic cells were transfected with mRNA by passive pulsing, lipofection, or electroporation. Dendritic cells were either left untreated or matured using the double-stranded RNA poly(I:C). T-Cell cultures were generated by stimulation of naïve T-cells with each set of dendritic cells. Specific antigen presentation and specific effector T-cell generation were analyzed by an IFN-γ release Elispot assay. Results. Greatest intracellular green fluorescent protein was observed by flow cytometry following dendritic cell electroporation with green fluorescent protein mRNA. DC presentation of Mart-1/Melan A peptide, as measured by Elispot assay using a specific T-cell clone, was greatest following transfection with Mart-1/Melan A mRNA by electroporation. Maturation of dendritic cells further improved antigen presentation regardless of transfection technique. Specific Mart-1/Melan A effector T cells were produced after culture of naïve T cells with dendritic cells that were electroporated with Mart-1/Melan A mRNA and then matured, but not for dendritic cells that remained immature. Conclusions. Efficient mRNA transfection by electroporation as well as dendritic cell maturation results in increased levels of Mart-1/Melan A antigen presentation and enhanced production of antigen-specific effector T cells. This combination of strategies may be used to enhance immune responses to RNA-based dendritic cell vaccines.

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