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

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
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spelling my-upm-ir.1046152023-10-31T01:41:04Z Targeted delivery of short hairpin RNA expressing plasmid using hepatitis B virus-like particle for bcl-2 gene silencing in cervical cancer cells 2021-10 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. Gene therapy Hepatitis B virus 2021-10 Thesis http://psasir.upm.edu.my/id/eprint/104615/ http://psasir.upm.edu.my/id/eprint/104615/1/FBSB%202022%2016%20IR.pdf text en public masters Universiti Putra Malaysia Gene therapy Hepatitis B virus Tan, Wen Siang
institution Universiti Putra Malaysia
collection PSAS Institutional Repository
language English
advisor Tan, Wen Siang
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
qualification_level Master's degree
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
granting_institution Universiti Putra Malaysia
publishDate 2021
url http://psasir.upm.edu.my/id/eprint/104615/1/FBSB%202022%2016%20IR.pdf
_version_ 1783725827113877504

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