Dendritic cells pulsed with RNA are potent antigen-presenting cells in vitro and in vivo.

Abstract

SulTlnlaryImmunization with defined tumor antigens is currently limited to a small number of cancers where candidates for tumor rejection antigens have been identified. In this study we investigated whether pulsing dendritic cells (DC) with tumor-derived P,.NA is an effective way to induce CTL and tumor immunity. DC pulsed with in vitro synthesized chicken ovalbumin (OVA) P, NA were more effective than OVA peptide-pulsed DC in stimulating primary, OVA-specific CTL responses in vitro. DC pulsed with unfractionated P, NA (total or polyA +) from OVA-expressing tumor cells were as effective as DC pulsed with OVA peptide at stimulating CTL responses. Induction of OVA-specific CTL was abrogated when polyA + P,.NA from OVA-expressing cells was treated with an OVA-specific antisense oligodeoxynucleotide and P,.Nase H, showing that sensitization of DC was indeed mediated by OVA P,.NA. Mice vaccinated with DC pulsed with P, NA from OVA-expressing tumor cells were protected against a challenge with OVA-expressing tumor cells. In the poorly immunogenic, highly metastatic, B16/F10.9 tumor model a dramatic reduction in lung metastases was observed in mice vaccinated with DC pulsed with tumor-derived P, NA (total or polyA +, but not polyA-P,.NA). The finding that RNA transcribed in vitro from cDNA cloned in a bacterial plasmid was highly effective in sensitizing DC shows that amplification of the antigenic content from a small number of tumor cells is feasible, thus expanding the potential use of P,.NA-pulsed DCbased vaccines for patients bearing very small, possibly microscopic, tumors.

Selective uptake of naked vaccine RNA by dendritic cells is driven by macropinocytosis and abrogated upon DC maturation

“…Although it is known for more than one decade that antigenencoding RNA co-incubated with APCs, is internalized and translated, 21 surprisingly few data are published on the underlying mechanisms in vitro and in vivo. The only study that investigated the uptake of RNA in vivo proposed a saturable Ca +2 ion-dependent process without defining the exact mechanism and cell type.…”

Route of administration influences the antitumor effects of bone marrow-derived dendritic cells engineered to produce interleukin-12 in a metastatic mouse prostate cancer model

“…In addition, it has been shown that DC pulsed with peptide of tumorassociated antigens, whole cell lysates, or RNA can trigger tumor-specific responses. [30][31][32] These approaches are currently limited in prostate cancer due to the lack of defined tumor antigens; the technical difficulties in obtaining sufficient amounts of mRNA; and limitations The T-cell stimulatory capacity of DC correlates with increased surface expression of MHC II and costimulatory molecules. 33 The upregulation of costimulatory molecules and MHC I/II of DC could enhance the immunogenicity of tumor antigens in poorly immunogenic cancers such as prostate cancer that are otherwise not recognized by immune system.…”

Toll‐like receptor‐dependent activation of several human blood cell types by protamine‐condensed mRNA

“…Similarly, in the context of immunostimulation by CpG motifs, the persistence leads to overstimulation and toxicity [5,6]. Taking all these drawbacks into account, our group as well as others developed genetic vaccines using messenger RNA (mRNA) instead of plasmid DNA [7][8][9][10][11] (for a review on mRNA vaccines, see Pascolo [12]). mRNA are easy to manufacture in high amounts, to purify to homogeneity and to characterize.…”