Safety profile of the intravenous administration of brain-targeted stable nucleic acid lipid particles.

Mariana Conceição,Célia Gomes,Luís Pereira De Almeida,Hirokazu Hirai,N Manjunath,Pedro Costa,Clévio Nóbrega,Maria C Lima,João Nuno Moreira,Liliana Mendonça

doi:10.1016/j.dib.2016.01.017

Abstract

In a clinical setting, where multiple administrations of the therapeutic agent are usually required to improve the therapeutic outcome, it is crucial to assess the immunogenicity of the administered nanoparticles. In this data work, we investigated the safety profile of the repeated intravenous administration of brain-targeted stable nucleic acid lipid particles (RVG-9r-targeted SNALPs). To evaluate local activation of the immune system, we performed analysis of mouse tissue homogenates and sections from cerebellum. To investigate peripheral activation of the immune system, we used serum of mice that were intravenously injected with RVG-9r-targeted SNALPs. These data are related and were discussed in the accompanying research article entitled “Intravenous administration of brain-targeted stable nucleic acid lipid particles alleviates Machado–Joseph disease neurological phenotype” (Conceição et al., in press) [1].

Highlights

In a clinical setting, where multiple administrations of the therapeutic agent are usually required to improve the therapeutic outcome, it is crucial to assess the immunogenicity of the administered nanoparticles
Results were analyzed using GraphPad Prism software Analyzed Mice were intravenously injected with brain-targeted stable nucleic acid lipid particles Mouse serum, mouse brain sections and mouse cerebellar homogenates were analyzed to evaluate the immune response Center for Neurosciences and Cell Biology, University of Coimbra, Coimbra, Portugal Data is supplied in this article
To investigate whether intravenous administration of RVG-9r-targeted SNALPs would contribute to inflammation, which could preclude repeated administration, we first evaluated the mRNA levels of proinflammatory cytokines (IL-1β, IL-6 and TNF-α) and a microglia-related gene (Cebpb) in the cerebella of these animals

Summary

Detection of the cerebellar mRNA levels of inflammatory mediators

5-weeks-old C57 BL/6 ataxin-3 [Q69] transgenic mice were intravenously injected on 3 consecutive days with 2.5 mg/kg of siRNA (siCTR or siMutAtax3) encapsulated in RVG-9r-targeted liposomes or HEPES-buffered saline solution (HBS). To analyze mutant ataxin-3 levels, mouse cerebella were harvested 48 h after the third injection. To detect cerebellum mRNA levels, total RNA was extracted using Qiazol solution (Qiagen) followed by purification of the RNA product using the Nucleospin RNA kit (Macherey-Nagel). CDNA synthesis was performed using the iSCRIPT cDNA synthesis kit (Bio-Rad). To determine the levels of IL-1β, IL-6 and TNF-α we used 2.5 mL of cDNA diluted 4 times (for Cebpb cDNA was diluted 10 times) and annealing temperature of 61 °C. QRT-PCR was performed using SsoAdvancedTM Universal SYBRs Green Supermix (Bio-Rad), in a Step One Plus Real-Time PCR System (Applied Biosystems). The mRNA relative quantification was performed with the Pfaffl method relatively to control samples

Fluorescence immunohistochemistry

Quantification of IL-6 serum levels by ELISA

Findings

Statistical analysis