Dataset Information

Expression Kinetics and Innate Immune Response after Electroporation and LNP-Mediated Delivery of a Self-Amplifying mRNA in the Skin.

ABSTRACT: In this work, we studied the expression kinetics and innate immune response of a self-amplifying mRNA (sa-RNA) after electroporation and lipid-nanoparticle (LNP)-mediated delivery in the skin of mice. Intradermal electroporation of the sa-RNA resulted in a plateau-shaped expression, with the plateau between day 3 and day 10. The overall protein expression of sa-RNA was significantly higher than that obtained after electroporation of plasmid DNA (pDNA) or non-replication mRNAs. Moreover, using IFN-? reporter mice, we elucidated that intradermal electroporation of sa-RNA induced a short-lived moderate innate immune response, which did not affect the expression of the sa-RNA. A completely different expression profile and innate immune response were observed when LNPs were used. The expression peaked 24 h after intradermal injection of sa-RNA-LNPs and subsequently showed a sharp drop. This drop might be explained by a translational blockage caused by the strong innate immune response that we observed in IFN-? reporter mice shortly (4 h) after intradermal injection of sa-RNA-LNPs. A final interesting observation was the capacity of sa-RNA-LNPs to transfect the draining lymph nodes after intradermal injection.

SUBMITTER: Huysmans H

PROVIDER: S-EPMC6722285 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Expression Kinetics and Innate Immune Response after Electroporation and LNP-Mediated Delivery of a Self-Amplifying mRNA in the Skin.

Huysmans Hanne H Zhong Zifu Z De Temmerman Joyca J Mui Barbara L BL Tam Ying K YK Mc Cafferty Séan S Gitsels Arlieke A Vanrompay Daisy D Sanders Niek N NN

Molecular therapy. Nucleic acids 20190807

In this work, we studied the expression kinetics and innate immune response of a self-amplifying mRNA (sa-RNA) after electroporation and lipid-nanoparticle (LNP)-mediated delivery in the skin of mice. Intradermal electroporation of the sa-RNA resulted in a plateau-shaped expression, with the plateau between day 3 and day 10. The overall protein expression of sa-RNA was significantly higher than that obtained after electroporation of plasmid DNA (pDNA) or non-replication mRNAs. Moreover, using IF ...[more]

PMID: 31472371

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Project description:Dengue virus (DENV) is the most common vector-borne viral disease, with nearly 400 million worldwide infections each year concentrated in the tropical and subtropical regions of the world. Severe dengue complications are often associated with a secondary heterotypic infection of one of the four circulating serotypes. In this scenario, humoral immune responses targeting cross-reactive, poorly neutralizing epitopes can lead to increased infectivity of susceptible cells via antibody-dependent enhancement (ADE). In this way, antibodies produced in response to infection or vaccination are capable of contributing to enhanced disease in subsequent infections. Currently, there are no available therapeutics to combat DENV disease, and there is an urgent need for a safe and efficacious vaccine. Here, we developed a nucleotide-modified mRNA vaccine encoding the membrane and envelope structural proteins from DENV serotype 1 encapsulated in lipid nanoparticles (prM/E mRNA-LNP). Vaccination of mice elicited robust antiviral immune responses comparable to viral infection, with high levels of neutralizing antibody titers and antiviral CD4+ and CD8+ T cells. Immunocompromised AG129 mice vaccinated with the prM/E mRNA-LNP vaccine were protected from a lethal DENV challenge. Vaccination with either a wild-type vaccine or a vaccine with mutations in the immunodominant fusion loop epitope elicited equivalent humoral and cell-mediated immune responses. Neutralizing antibodies elicited by the vaccine were sufficient to protect against a lethal challenge. Both vaccine constructs demonstrated serotype-specific immunity with minimal serum cross-reactivity and reduced ADE in comparison to a live DENV1 viral infection.IMPORTANCE With 400 million worldwide infections each year, dengue is the most common vector-borne viral disease. Forty percent of the world's population is at risk, with dengue experiencing consistent geographic spread over the years. With no therapeutics available and vaccines performing suboptimally, the need for an effective dengue vaccine is urgent. Here, we develop and characterize a novel mRNA vaccine encoding the dengue serotype 1 envelope and premembrane structural proteins that is delivered via a lipid nanoparticle. Our DENV1 prM/E mRNA-LNP vaccine induces neutralizing antibody and cellular immune responses in immunocompetent mice and protects an immunocompromised mouse from a lethal DENV challenge. Existing antibodies against dengue can enhance subsequent infections via antibody-dependent enhancement (ADE). Importantly our vaccine induced only serotype-specific immune responses and did not induce ADE.

Project description:The skin is an attractive tissue for vaccination in a clinical setting due to the accessibility of the target, the ease of monitoring and most importantly the immune competent nature of the dermal tissue. While skin electroporation offers an exciting and novel future methodology for the delivery of DNA vaccines in the clinic, little is known about the actual mechanism of the approach and the elucidation of the resulting immune responses. To further understand the mechanism of this platform, the expression kinetics and localization of a reporter plasmid delivered via a surface dermal electroporation (SEP) device as well as the effect that this treatment would have on the resident immune cells in that tissue was investigated. Initially a time course (day 0 to day 21) of enhanced gene delivery with electroporation (EP) was performed to observe the localization of green fluorescent protein (GFP) expression and the kinetics of its appearance as well as clearance. Using gross imaging, GFP expression was not detected on the surface of the skin until 8 h post treatment. However, histological analysis by fluorescent microscopy revealed GFP positive cells as early as 1 h after plasmid delivery and electroporation. Peak GFP expression was observed at 24 h and the expression was maintained in skin for up to seven days. Using an antibody specific for a keratinocyte cell surface marker, reporter gene positive keratinocytes in the epidermis were identified. H&E staining of treated skin sections demonstrated an influx of monocytes and granulocytes at the EP site starting at 4 h and persisting up to day 14 post treatment. Immunological staining revealed a significant migration of lymphocytic cells to the EP site, congregating around cells expressing the delivered antigen. In conclusion, this study provides insights into the expression kinetics following EP enhanced DNA delivery targeting the dermal space. These findings may have implications in the future to design efficient DNA vaccination strategies for the clinic.

Dataset Information

Expression Kinetics and Innate Immune Response after Electroporation and LNP-Mediated Delivery of a Self-Amplifying mRNA in the Skin.

Publications

Expression Kinetics and Innate Immune Response after Electroporation and LNP-Mediated Delivery of a Self-Amplifying mRNA in the Skin.

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