CRISPR-Cas13a Targeting the FGFR3-TACC3 Fusion Gene Inhibits Proliferation of Bladder Cancer Cells in vitro and in vivo.

Abstract

Introduction: The FGFR3-TACC3 fusion gene exists in a variety of malignant tumors, including bladder cancer. In our ongoing research on the CRISPR-Cas13a gene-editing system, we reported the use of CRISPR-Cas13a gene-editing system to knockout FGFR3-TACC3 and inhibit the proliferation of bladder tumor cells.

Purpose:  This study aimed to use the CRISPR-Cas13a gene-editing system to target the FGFR3-TACC3 fusion gene in bladder cancer cells, which has the potential to be a new and effective treatment for bladder cancer.

Materials and methods: The efficacy of the CRISPR-Cas13a gene-editing system was analysed by qRT-PCR. The inhibitory effects of Cas13a-mediated knockdown of the FGFR3-TACC3 fusion gene on the proliferation of RT4 and RT112 cell lines were assessed utilizing CCK-8, EdU, and organoid formation assays. Subsequently, the comparative tumorigenic capability of RT4 cells with FGFR3-TACC3 knockdown achieved by Cas13a was examined in a nude mouse model.

Results: At the cellular level, the comparative analysis of FGFR3-TACC3 knockdown efficacy between CRISPR-Cas13a and shRNA revealed a more pronounced reduction with the former. This knockdown effectively curtailed cellular proliferation, with CRISPR-Cas13a-mediated knockdown exhibiting a superior inhibitory effect over shRNA-mediated knockdown. In organoid cultures derived from RT4 cells, a similar trend was observed, with Cas13a-mediated knockdown of FGFR3-TACC3 leading to a more substantial suppression of proliferation compared to shRNA-mediated knockdown. In vivo tumor models corroborated these findings, demonstrating a significantly diminished tumor volume in the Cas13a-treated cohort relative to both the control and shRNA-treated groups.

Conclusion: The CRISPR-Cas13a gene-editing system has been demonstrated to significantly suppress tumor proliferation both in vitro and in vivo, thereby presenting itself as a promising candidate for a novel and efficacious therapeutic intervention in bladder cancer treatment.