# Unraveling the impact of AXIN1 mutations on HCC development: Insights from CRISPR/Cas9 repaired AXIN1-mutant liver cancer cell lines

## TL;DR

This study examines how repairing AXIN1 mutations in liver cancer cells affects cancer-related signaling and growth.

## Contribution

The study uses CRISPR/Cas9 to repair AXIN1 mutations and investigates their impact on β-catenin signaling and cancer cell behavior.

## Key findings

- AXIN1 repair reduced β-catenin signaling and cell viability in HCC cell lines.
- Wnt3A exposure partially restored signaling but not growth, suggesting other mechanisms are involved.
- RNA-sequencing showed no consistent gene/pathway changes beyond β-catenin targets.

## Abstract

Hepatocellular carcinoma (HCC) is a highly aggressive liver cancer with significant morbidity and mortality rates. AXIN1 is one of the top-mutated genes in HCC, but the mechanism by which AXIN1 mutations contribute to HCC development remains unclear.

In this study, we utilized CRISPR/Cas9 genome editing to repair AXIN1-truncated mutations in five HCC cell lines.

For each cell line we successfully obtained 2–4 correctly repaired clones, which all show reduced β-catenin signaling accompanied with reduced cell viability and colony formation. Although exposure of repaired clones to Wnt3A-conditioned medium restored β-catenin signaling, it did not or only partially recover their growth characteristics, indicating the involvement of additional mechanisms. Through RNA-sequencing analysis, we explored the gene expression patterns associated with repaired AXIN1 clones. Except for some highly-responsive β-catenin target genes, no consistent alteration in gene/pathway expression was observed. This observation also applies to the Notch and YAP/TAZ-Hippo signaling pathways, which have been associated with AXIN1-mutant HCCs previously. The AXIN1-repaired clones also cannot confirm a recent observation that AXIN1 is directly linked to YAP/TAZ protein stability and signaling.

Our study provides insights into the effects of repairing AXIN1 mutations on β-catenin signaling, cell viability, and colony formation in HCC cell lines. However, further investigations are necessary to understand the complex mechanisms underlying HCC development associated with AXIN1 mutations.