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Polymeric micellar nanoparticles for effective CRISPR/Cas9 genome editing in cancer
Yuzhen Li1, Chun Li1, Jiachang Yan1
1School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Shenzhen, 518107, China; School of Pharmaceutical Sciences (Shenzhen), Sun Yat-sen University, Shenzhen, 518107, China.
Abstract:
The clustered regularly interspaced short palindromic repeat (CRISPR)-associated protein 9 (Cas9) gene editing has attracted extensive attentions in various fields, however, its clinical application is hindered by the lack of effective and safe delivery system. Herein, we reported a cationic micelle nanoparticle composed of cholesterol-modified branched small molecular PEI (PEI-CHO) and biodegradable PEG-b-polycarbonate block copolymer (PEG-PC), denoted as PEG-PC/PEI-CHO/pCas9, for the CRISPR/Cas9 delivery to realize genomic editing in cancer. Specifically, PEI-CHO condensed pCas9 into nanocomplexes, which were further encapsulated into PEG-PC nanoparticles (PEG-PC/PEI-CHO/pCas9). PEG-PC/PEI-CHO/pCas9 had a PEG shell, protecting DNA from degradation by nucleases. Enhanced cellular uptake of PEG-PC/PEI-CHO/pCas9 nanoparticles was observed as compared to that mediated by Lipo2k/pCas9 nanoparticles, thus leading to significantly elevated transfection efficiency after escaping from endosomes via the proton sponge effect of PEI. In addition, the presence of PEG shell greatly improved biocompatibility, and significantly enhanced the in vivo tumor retention of pCas9 compared to PEI-CHO/pCas9. Notably, apparent downregulation of GFP expression could be achieved both in vitro and in vivo by using PEG-PC/PEI-CHO/pCas9-sgGFP nanoparticles. Furthermore, PEG-PC/PEI-CHO/pCas9-sgMcl1 induced effective apoptosis and tumor suppression in a HeLa tumor xenograft mouse model by downregulating Mcl1 expression. This work may provide an alternative paradigm for the efficient and safe genome editing in cancer.
Insights
A novel nanoparticle system delivers CRISPR-Cas9 gene editing tools for cancer therapy. This system enhances cellular uptake and tumor retention, leading to effective gene silencing and tumor suppression.
Area of Science:
- Biomedical Engineering
- Nanotechnology
- Gene Therapy
Background:
- CRISPR-Cas9 gene editing holds promise for cancer treatment but requires safe and effective delivery systems.
- Existing delivery methods face challenges in clinical application due to safety and efficacy concerns.
Purpose of the Study:
- To develop and evaluate a novel cationic micelle nanoparticle for CRISPR-Cas9 delivery in cancer.
- To assess the nanoparticle's efficiency in genomic editing, cellular uptake, biocompatibility, and in vivo tumor suppression.
Main Methods:
- Formulation of a nanoparticle system (PEG-PC/PEI-CHO/pCas9) using cholesterol-modified PEI and a biodegradable block copolymer.
- Evaluation of cellular uptake, endosomal escape, and transfection efficiency compared to a commercial liposome.
- Assessment of in vitro and in vivo gene silencing using GFP and Mcl1 targets.
- Testing of tumor suppression efficacy in a HeLa tumor xenograft mouse model.
Main Results:
- The developed PEG-PC/PEI-CHO/pCas9 nanoparticles demonstrated enhanced cellular uptake and transfection efficiency.
- The nanoparticle system showed improved biocompatibility and in vivo tumor retention.
- Effective downregulation of GFP and Mcl1 expression was achieved both in vitro and in vivo.
- Significant tumor suppression and apoptosis induction were observed in the xenograft model.
Conclusions:
- The PEG-PC/PEI-CHO/pCas9 nanoparticle system offers an efficient and safe platform for CRISPR-Cas9 delivery in cancer.
- This approach presents a promising alternative for advancing genome editing applications in oncology.
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