Efficient delivery of mRNA to dendritic cells using in vitro reconstituted Virus-Like Particles (VLPs)
Habtom H. Habte1, Adam Biddlecome2, Katherine M. McGrath1, Sharmila Sambanthamoorthy1, Charles M. Knobler2, John B. Leppard1, Marcio Lasaro1, William M. Gelbart2, Knut Elbers3.
1Boehringer Ingelheim Pharmaceuticals Inc., Ridgefield, USA; 2University of California, Los Angeles, USA; 3Boehringer Ingelheim Pharmaceuticals Inc., Ingelheim, Germany
Dendritic cells (DCs) are highly specialized antigen presenting cells of the immune system that play a key role in inducing effective CD4+ and CD8+ T cell responses. Efficient delivery of vaccines to DCs is a critical step towards developing a protective vaccine against infectious diseases and cancers. Cowpea Chlorotic Mottle Virus (CCMV) is a single-stranded (+)-sense RNA virus whose capsid protein can self-assemble in vitro with cognate or heterologous RNA into VLPs. Here, we evaluated the use of CCMV VLPs as an RNA-based vaccine delivery platform, including whether antibodies against the viral capsid protein can modulate the RNA uptake and activation of DCs. For these studies monocyte-derived DCs generated from human PBMCs were incubated with either unpackaged/”naked” mRNA or with the mRNA in CCMV-VLP form. To evaluate the effect of anti-capsid antibodies, the CCMV-VLPs were pre-incubated with anti-CCMV antibodies. After incubating DCs with the VLPs for 24 hours, we determined the expression of maturation markers – CD80, CD86 and MHC-II – by flow cytometry, and the mRNA levels by quantitative PCR. We observed an increased expression of maturation markers and mRNA levels in DCs treated with mRNA-containing CCMV-VLPs compared to mRNA treatment alone. Remarkably, an enhanced DC activation and high mRNA uptake was detected in a dose-dependent manner when the CCMV-VLPs were pre-incubated with anti-CCMV antibodies. In all experiments expression of maturation markers correlates with the mRNA levels of the DCs. Overall, these studies demonstrate that VLP-protection is able to enhance mRNA uptake by DCs and that the presence of anti-CCMV antibodies significantly enhances mRNA levels and activation of DCs in vitro.
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