Description

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.