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Efficiency of Chitosan-Coated PLGA Nanocarriers for Cellular Delivery of siRNA and CRISPR/Cas9 Complex

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Abstract

Purpose

Most studies based on siRNA or CRISPR/Cas9 systems rely on viral vectors for their transfection. However, these viral vectors are immunogenic, which limits their application in gene therapy. Thus, identification of novel vectors with higher safety and improved targeting is desirable. Hence, we have explored the potential of biodegradable, chitosan coated PLGA nanocarriers for intracellular delivery of CRISPR/Cas9, and siRNA. In this investigation, we have compared the efficiency of chitosan-coated PLGA NPs (CS-PLGA NPs) with that of the PLGA NPs for the delivery of CRISPR and siRNA.

Methods

Presented here is the preparation and evaluation of specifically surface-modified CS-PLGA NPs and PLGA NPs on their efficacy for binding and delivery of siRNA and CRISPR/Cas9 complex.

Results

Cy3 siRNA loaded on PLGA NPs showed an internalization of 4.6% and mean fluorescent intensity (MFI) of 13.76%, while that of CS PLGA resulted in 89% internalization and MFI of 67.95%. The in vitro GFP silencing assessed by anti-GFP siRNA and NPs resulted in 10–15% silencing by PLGA NPs and 50–55% silencing by CS-PLGA NPs. The GFP silencing by CRISPR-Cas9 plasmid pX459 with CS PLGA was 80–83%, while that of PLGA was 11% and of commercial Lipofectamine agent was 13%.

Conclusions

The biodegradable CS-PLGA NPs exhibited successful loading and high binding efficiency for siRNA as well as CRISPRCas9 and resulted in effective silencing. Our studies report that the CS-PLGA NPs can be a novel suitable candidate for the effective delivery of siRNA and CRISPR/Cas9 complex.

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Acknowledgements

The author is very thankful to Mr. Lalit Borade from TIFR, Mumbai, for giving the facility of TEM imaging. Mr. Anjan Ghosh has helped in acquiring the FACS data from BD FACS Melody. We also like to acknowledge Ms. Namrata, Ms. Soumya, and Mr. Lalit from the laboratory of Dr. Debjyoti, IGIB, New Delhi, for providing CRISPR/Cas9, plasmid, and HEK 293T GFP-positive cell sample.

Funding

The authors are thankful to Department of Science Technology (DST) Nanomission (SR/NM/NS1145/2012) and Indian Council of Medical Research (ICMR) (No. 3/1/3/JRF-2015/HRD-LS/29/31232/85) Government of India for the research grant and fellowship.

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Authors and Affiliations

  1. Department of Chemical Engineering, Institute of Chemical Technology, NP Marg, Matunga E, Mumbai, 400019, India

    Ashu Srivastav, Kritika Gupta & Ratnesh Jain

  2. Department of Pharmaceutical Sciences and Technology, Institute of Chemical Technology, NP Marg, Matunga E, Mumbai, 400019, India

    Ashu Srivastav, Kritika Gupta & Prajakta Dandekar

  3. RNA lab, CSIR-Institute of Genomics & Integrative Biology, Mathura Road, Delhi, 110025, India

    Debojyoti Chakraborty

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  1. Ashu Srivastav
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Correspondence to Prajakta Dandekar or Ratnesh Jain.

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Srivastav, A., Gupta, K., Chakraborty, D. et al. Efficiency of Chitosan-Coated PLGA Nanocarriers for Cellular Delivery of siRNA and CRISPR/Cas9 Complex. J Pharm Innov 17, 180–193 (2022). https://doi.org/10.1007/s12247-020-09496-4

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