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Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells

Gene therapy research has advanced rapidly to clinical trials, but it is greatly hampered by the unstable nucleic acids particularly short interference RNA (siRNA) packaged inside carriers, and the lack of specificity towards targeted sites in the body. Hence, development of a stable carrier with sp...

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Main Author: Akwiditya, Made Angga
Format: Thesis
Language:English
Published: 2021
Subjects:
Gene therapy
Hepatitis B virus
Online Access:http://psasir.upm.edu.my/id/eprint/104615/1/FBSB%202022%2016%20IR.pdf
http://psasir.upm.edu.my/id/eprint/104615/
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spelling my.upm.eprints.1046152023-10-31T01:41:04Z http://psasir.upm.edu.my/id/eprint/104615/ Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells Akwiditya, Made Angga Gene therapy research has advanced rapidly to clinical trials, but it is greatly hampered by the unstable nucleic acids particularly short interference RNA (siRNA) packaged inside carriers, and the lack of specificity towards targeted sites in the body. Hence, development of a stable carrier with specific targeted delivery is urgently needed. This study aimed to address gene therapy limitations by encapsidating a plasmid synthesizing short hairpin RNA (shRNA) that targets the anti-apoptotic Bcl-2 gene (namely PshRNA) using truncated hepatitis B virus core antigen (tHBcAg) virus-like particle (VLP). A siRNA sequence targeting the anti-apoptotic Bcl-2 was synthesized and cloned into pSilencer 2.0-U6 vector, and encapsidated inside tHBcAg VLP. The VLP encapsidating PsiRNA was conjugated with folic acid (FA) to produce FA-tHBcAg-PsiRNA VLP. Scanning transmission electron microscopy revealed that FA-tHBcAg-PsiRNA VLP has icosahedral structure similar to that of the unmodified tHBcAg VLP. Delivery of FA-tHBcAg-PsiRNA VLP into HeLa cells overexpressing folate receptor (FR) significantly downregulated the expression of anti-apoptotic Bcl-2 at 48- and 72-hours post-transfection. MTT assay demonstrated that the cells’ viability was significantly reduced from 89.46% at 24 h to 64.52% and 60.63%, respectively, at 48- and 72-hours post-transfection. As a conclusion, tHBcAg VLP can be used as a carrier for a receptor-mediated targeted delivery of a therapeutic plasmid encoding shRNA for gene silencing in cancer cells. 2021-10 Thesis NonPeerReviewed text en http://psasir.upm.edu.my/id/eprint/104615/1/FBSB%202022%2016%20IR.pdf Akwiditya, Made Angga (2021) Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells. Masters thesis, Universiti Putra Malaysia. Gene therapy Hepatitis B virus
institution Universiti Putra Malaysia
building UPM Library
collection Institutional Repository
continent Asia
country Malaysia
content_provider Universiti Putra Malaysia
content_source UPM Institutional Repository
url_provider http://psasir.upm.edu.my/
language English
topic Gene therapy
Hepatitis B virus
spellingShingle Gene therapy
Hepatitis B virus
Akwiditya, Made Angga
Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells
description Gene therapy research has advanced rapidly to clinical trials, but it is greatly hampered by the unstable nucleic acids particularly short interference RNA (siRNA) packaged inside carriers, and the lack of specificity towards targeted sites in the body. Hence, development of a stable carrier with specific targeted delivery is urgently needed. This study aimed to address gene therapy limitations by encapsidating a plasmid synthesizing short hairpin RNA (shRNA) that targets the anti-apoptotic Bcl-2 gene (namely PshRNA) using truncated hepatitis B virus core antigen (tHBcAg) virus-like particle (VLP). A siRNA sequence targeting the anti-apoptotic Bcl-2 was synthesized and cloned into pSilencer 2.0-U6 vector, and encapsidated inside tHBcAg VLP. The VLP encapsidating PsiRNA was conjugated with folic acid (FA) to produce FA-tHBcAg-PsiRNA VLP. Scanning transmission electron microscopy revealed that FA-tHBcAg-PsiRNA VLP has icosahedral structure similar to that of the unmodified tHBcAg VLP. Delivery of FA-tHBcAg-PsiRNA VLP into HeLa cells overexpressing folate receptor (FR) significantly downregulated the expression of anti-apoptotic Bcl-2 at 48- and 72-hours post-transfection. MTT assay demonstrated that the cells’ viability was significantly reduced from 89.46% at 24 h to 64.52% and 60.63%, respectively, at 48- and 72-hours post-transfection. As a conclusion, tHBcAg VLP can be used as a carrier for a receptor-mediated targeted delivery of a therapeutic plasmid encoding shRNA for gene silencing in cancer cells.
format Thesis
author Akwiditya, Made Angga
author_facet Akwiditya, Made Angga
author_sort Akwiditya, Made Angga
title Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells
title_short Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells
title_full Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells
title_fullStr Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells
title_full_unstemmed Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells
title_sort targeted delivery of short hairpin rna expressing plasmid using hepatitis b virus-like particle for bcl-2 gene silencing in cervical cancer cells
publishDate 2021
url http://psasir.upm.edu.my/id/eprint/104615/1/FBSB%202022%2016%20IR.pdf
http://psasir.upm.edu.my/id/eprint/104615/
_version_ 1781706768374562816
score 13.211869

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