Optimized microfluidic formulation and organic excipients for improved lipid nanoparticle mediated genome editing,Lab on a Chip

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Optimized microfluidic formulation and organic excipients for improved lipid nanoparticle mediated genome editing
Lab on a Chip ( IF 6.1 ) Pub Date : 2024-07-22 , DOI: 10.1039/d4lc00283k
Rohan Palanki _{1,

2} , Emily L Han ₁ , Amanda M Murray ₁ , Rohin Maganti _{1,

2} , Sophia Tang ₁ , Kelsey L Swingle ₁ , Dongyoon Kim ₁ , Hannah Yamagata ₁ , Hannah C Safford ₁ , Kaitlin Mrksich ₁ , William H Peranteau ₂ , Michael J Mitchell _{1,

3,

4,

5,

6,

7,

8}

Affiliation

mRNA-based gene editing platforms have tremendous promise in the treatment of genetic diseases. However, for this potential to be realized in vivo, these nucleic acid cargos must be delivered safely and effectively to cells of interest. Ionizable lipid nanoparticles (LNPs), the most clinically advanced non-viral RNA delivery system, have been well-studied for the delivery of mRNA but have not been systematically optimized for the delivery of mRNA-based CRISPR-Cas9 platforms. In this study, we investigated the effect of microfluidic and lipid excipient parameters on LNP gene editing efficacy. Through in vitro screening in liver cells, we discovered distinct trends in delivery based on phospholipid, cholesterol, and lipid-PEG structure in LNP formulations. Combination of top-performing lipid excipients produced an LNP formulation that resulted in 3-fold greater gene editing in vitro and facilitated 3-fold greater reduction of a therapeutically-relevant protein in vivo relative to the unoptimized LNP formulation. Thus, systematic optimization of LNP formulation parameters revealed a novel LNP formulation that has strong potential for delivery of gene editors to the liver to treat metabolic disease.

中文翻译：

优化的微流体配方和有机赋形剂用于改进脂质纳米颗粒介导的基因组编辑

基于 mRNA 的基因编辑平台在遗传疾病的治疗中具有巨大的前景。然而，为了在体内实现这种潜力，这些核酸货物必须安全有效地递送至感兴趣的细胞。可电离脂质纳米粒子 (LNP) 是临床上最先进的非病毒 RNA 递送系统，已针对 mRNA 的递送进行了深入研究，但尚未针对基于 mRNA 的 CRISPR-Cas9 平台的递送进行系统优化。在这项研究中，我们研究了微流体和脂质赋形剂参数对 LNP 基因编辑功效的影响。通过肝细胞的体外筛选，我们发现了 LNP 制剂中基于磷脂、胆固醇和脂质-PEG 结构的独特递送趋势。与未优化的 LNP 制剂相比，结合性能最佳的脂质赋形剂产生的 LNP 制剂可实现 3 倍的体外基因编辑，并促进体内治疗相关蛋白的减少 3 倍。因此，LNP 制剂参数的系统优化揭示了一种新型 LNP 制剂，该制剂具有将基因编辑器递送至肝脏以治疗代谢疾病的强大潜力。

更新日期：2024-07-22

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