# Differential Effects of Snail-KO in Human Breast Epithelial Cells and Human Breast Epithelial × Human Breast Cancer Hybrids

## TL;DR

This study shows that Snail affects stemness in breast cancer hybrids but not EMT, with different effects in pure and hybrid cells.

## Contribution

The study reveals Snail's role in stemness rather than EMT in breast cancer hybrids, highlighting cell-type-specific functions.

## Key findings

- Snail-KO in pure cells induced EMT and increased colony formation.
- Snail-KO in hybrids reduced mammosphere formation but did not alter EMT markers.
- Hybrid Snail-KO cells showed increased invasive capacity.

## Abstract

Snail and Zeb1 have been suggested as markers for the hybrid/mixed epithelial (E)/mesenchymal (M) state of cancer cells. Such cancer cells co-express E- and M-specific transcripts and possess cancer stem cell properties. M13HS-2/-8 tumor hybrid clones derived from human M13SV1-EGFP-Neo breast epithelial cells and human HS578T-Hyg breast cancer cells exhibited co-expression of Snail and Zeb1. To explore the impact of Snail on stemness/epithelial-to-mesenchymal transition (EMT)-related properties in M13HS-2/-8 tumor hybrid clones, Snail was knocked out (KO) using CRISPR/Cas9. Mammosphere formation, colony formation, Western blot analyses, cell migration, and invasion assays were conducted for the characterization of Snail knockout cells. Interestingly, Snail-KO in M13SV1-EGFP-Neo cells resulted in the up-regulation of vimentin and N-cadherin, suggesting EMT induction, which was associated with a significantly enhanced colony formation capacity. In contrast, EMT marker pattern and colony formation capacities of M13HS-2/-8 Snail-KO tumor hybrid clones remained unchanged. Notably, the mammosphere formation capacities of M13HS-2/-8 Snail-KO tumor hybrid clones were significantly reduced. The migratory behavior of all Snail-KO cells was not altered compared with their wild-type counterparts. In contrast, M13HS-2 hybrids and their M13HS-2 Snail-KO variant exhibited a markedly enhanced invasive capacity. Therefore, Snail plays a role as a mediator of stemness properties rather than mediating EMT.