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A New Toolkit for Evaluating Gene Functions using Conditional Cas9 Stabilization
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A New Tool for CRISPR-Cas13a-Based Cancer Gene Therapy

Jinliang Gao1, Tao Luo1, Na Lin1

  • 1State Key Laboratory of Bioelectronics, Southeast University, Nanjing 210096, P.R. China.

Molecular Therapy Oncolytics
|October 26, 2020
PubMed

Insights

Researchers developed a novel CRISPR-Cas13a tool (DCUg) for cancer gene therapy. This tool effectively targets and inhibits oncogene expression in cancer cells, leading to significant tumor growth reduction in vitro and in vivo.

Area of Science:

  • Molecular Biology
  • Gene Therapy
  • Oncology

Background:

  • The CRISPR-Cas13a system, known for virus detection, has underexplored potential in cancer treatment.
  • Targeting oncogenes offers a promising strategy for cancer therapy, but effective delivery and specificity remain challenges.

Purpose of the Study:

  • To engineer a novel Cas13a-based gene expression vector (DCUg) for targeted cancer gene therapy.
  • To evaluate the efficacy of DCUg in knocking down oncogene expression and inhibiting cancer cell growth.
  • To assess the therapeutic potential of DCUg delivered via adeno-associated virus (AAV) in vitro and in vivo.

Main Methods:

  • Construction of a new Cas13a expression vector (DCUg) utilizing specific promoters for controlled gene expression.
  • Assessment of DCUg's ability to knock down reporter and endogenous oncogene (TERT, EZH2, RelA) expression in cancer cells (HepG2) and normal cells (HL7702).
  • Delivery of DCUg via adeno-associated virus (AAV) for in vitro treatment of multiple cancer cell lines and in vivo treatment of tumor-bearing mice.

Main Results:

  • DCUg specifically and effectively reduced oncogene expression at both mRNA and protein levels in HepG2 cells, inducing apoptosis and inhibiting growth.
  • DCUg demonstrated specificity, with no significant impact on normal liver cells (HL7702).
  • AAV-packaged DCUg significantly inhibited the growth of various cancer cells in vitro and suppressed xenografted tumors in mice.

Conclusions:

  • The developed DCUg vector represents a novel and effective tool for CRISPR-Cas13a-based cancer gene therapy.
  • Targeting oncogenes with DCUg shows significant therapeutic potential for inhibiting cancer progression.
  • AAV-mediated delivery of DCUg offers a promising strategy for in vivo cancer treatment.
Keywords:
CRISPR-Cas13aNF-κBRNA interferencecancergene therapy

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