WS05.01Lipid nanoparticles and vesicles delivery tools to deliver mRNA and ribonucleoprotein to the lungs for the development of genome editing in cystic fibrosis

IF 5.4 2区医学 Q1 RESPIRATORY SYSTEM

Journal of Cystic Fibrosis Pub Date : 2025-06-01 DOI:10.1016/j.jcf.2025.03.516

G. Maule , S. Saxena , A. Colliva , S. Vodret , J. Parot , A. Molska , E. Gurrieri , M. Stancampiano , S.E. Borgos , D. Guidone , A. Borrelli , L.J.V. Galietta , S. Hak , S. Zacchigna , A. Cereseto

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引用次数: 0

Abstract

Objectives

Genome editing technologies hold great for the treatment of cystic fibrosis (CF). However, current delivery methods remain inefficient, posing a major barrier to their clinical translation.

The GenDel-CF project aims to develop in vivo delivery tools to transfer 1) mRNA and 2) ribonucleoprotein (RNP) expressing CRISPR-Cas9.

Methods and Results

For the first goal our consortium aims to develop novel lipid nanoparticle (LNP) formulations. We recently developed LNP-GD19 that were tested by encapsulating Cre-mRNA and delivered to reporter mice. LNPs were delivered via either intratracheal (IT) administration in adult mice or intravenous (IV) injection in neonates. We obtained high levels of Cre-mediated recombination resulting in mGFP expression throughout the lung parenchyma upon both administration routes. IT administration resulted nearly 80% of GFP-positive in secretory cells, 48% in epithelial cells from the airways and submucosal glands, and 75% in bronchial epithelial and alveolar type 1 cells. This pointed to IT delivery as the best delivery route for targeting epithelial cells critical for CF therapy.

For the second aim, consisting in CRISPR-Cas delivery as RNP we focused on engineered vesicles as promising delivery systems for genome editing. We exploited base editors as editing systems, specifically ABE8e-SpCas9, due to their proven efficiency in precisely and effectively repairing CFTR mutations. The GE-vesicles were produced using membrane anchoring motifs to capture maximal amounts of ABE8e-SpCas9 and sgRNA. Particles were characterized for size, particle number and editor content showing overall homogenous size in the range of 100-130 nm and efficient encapsulation of the ABE8e-SpCas9 resulting in up to 60% of base conversion.

Conclusions

In conclusion, our GD19 LNP formulation GE-vesicles lay the foundation for editing tools delivery to repair CF mutations.

查看原文本刊更多论文

脂质纳米颗粒和囊泡递送工具将mRNA和核糖核蛋白递送到肺部，用于囊性纤维化基因组编辑的发展

目的基因组编辑技术在囊性纤维化（CF）的治疗中具有重要意义。然而，目前的递送方法仍然效率低下，对其临床转化构成了主要障碍。GenDel-CF项目旨在开发体内递送工具来转移表达CRISPR-Cas9的1)mRNA和2)核糖核蛋白（RNP）。方法和结果第一个目标是开发新型脂质纳米颗粒（LNP）制剂。我们最近开发了LNP-GD19，通过封装Cre-mRNA并传递给报告小鼠进行了测试。LNPs通过成年小鼠气管内（IT）给药或新生儿静脉注射（IV）给药。我们获得了高水平的cre介导重组，导致mGFP在两种给药途径下在整个肺实质中表达。给药后，分泌细胞中近80%的gfp阳性，气道和粘膜下腺上皮细胞中48%的gfp阳性，支气管上皮和肺泡1型细胞中75%的gfp阳性。这表明IT递送是靶向对CF治疗至关重要的上皮细胞的最佳递送途径。对于第二个目标，包括CRISPR-Cas作为RNP的递送，我们专注于工程囊泡作为基因组编辑的有前途的递送系统。我们利用碱基编辑器作为编辑系统，特别是ab8e - spcas9，因为它们在精确和有效地修复CFTR突变方面具有效率。利用膜锚定基序产生ge囊泡，以捕获最大量的ab8e - spcas9和sgRNA。对颗粒的大小、颗粒数和编辑器含量进行了表征，表明在100-130 nm范围内的颗粒尺寸总体均匀，并且对ABE8e-SpCas9的有效封装导致高达60%的碱基转化率。总之，我们的GD19 LNP制剂ge囊泡为编辑工具递送修复CF突变奠定了基础。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Journal of Cystic Fibrosis 医学-呼吸系统

CiteScore

10.10

自引率

13.50%

发文量

1361

审稿时长

50 days

期刊介绍： The Journal of Cystic Fibrosis is the official journal of the European Cystic Fibrosis Society. The journal is devoted to promoting the research and treatment of cystic fibrosis. To this end the journal publishes original scientific articles, editorials, case reports, short communications and other information relevant to cystic fibrosis. The journal also publishes news and articles concerning the activities and policies of the ECFS as well as those of other societies related the ECFS.