Design of trans-splicing RNATechnology #15306n
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Gene therapy is considered to be a highly effective treatment for chronic diseases with minimum side effects. The global cancer gene therapy market was valued at $289 million in 2016, and is estimated to reach $2,082 million by 2023, at a compound annual growth rate of 32.4% from 2017 to 2023.
An advanced tsRNA design towards a Herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV) suicide gene therapy approach. Tested for hepatocellular carcinoma and cervical cancer.
Figure 1: Design of tsRNA for 5′ and 3′ exon replacement (ER) and structure of an alpha fetoprotein (AFP) mini-gene.
1. Hepatocellular carcinoma (HCC): alpha-fetoprotein (AFP), hepatocellular carcinoma-associated gene 2/ Yin Yang 1-associated protein 1 (HCCA2/YY1AP1), cluster of differentiation 24 (CD24) and Vascular endothelial growth factor (VEGF)
2. Cervical cancer: HPV-16
In vitro anti-tumour efficacy:
1. Hepatocellular carcinoma:
a. Investigation of death of HepG2 cells triggered by 5’ or 3’ exon replacement (ER) toward overexpressed and endogenous AFP pre-mRNA using AlamarBlue cell viability assay:
i. up to 80% at 100 μM or 60% to 70% at 10 μM GCV
ii. No cytotoxicity was triggered by the AFP mini-gene or the trans-splicing constructs in the absence of GCV.
b. Investigation of death of HepG2 cells triggered by 5’ or 3’ ER toward overexpressed and endogenous AFP pre-mRNA and another HCC biomarker (HCCA2/YY1AP1, CD24, or VEGF as secondary targets) using AlamarBlue cell viability assay:
i. At 10 μM GCV, all dual-targeting constructs triggered significantly higher levels of cell death compared with the construct targeting AFP only.
ii. At 100 μM GCV, single- and dual-targeting constructs showed comparable effects
2. Cervical cancer:
a. Investigation of death of SiHa and C3 cells triggered by 5’ or 3’ ER toward overexpressed and endogenous HPV-16 pre-mRNA using AlamarBlue cell viability assay:
i. In SiHa cells, all tsRNAs triggered cell death to the same extent as the positive control
ii. In C3 cells, 3’ ER was more efficient than 5’ ER.
STAGE OF DEVELOPMENT
• Gene therapy
• High efficacy
• Minimum side effects