# Inhibiting the TGF-β1 Pathway Reduces the Aggressiveness of Intrahepatic CCA HuCCT1 CD90-Positive Cells

## TL;DR

Inhibiting the TGF-β1 pathway reduces the aggressive behavior of CD90-positive intrahepatic cholangiocarcinoma cells and improves response to chemotherapy.

## Contribution

The study identifies CD90 as a marker of aggressive cancer behavior and shows that combining TGF-β1 inhibition with chemotherapy enhances treatment effectiveness.

## Key findings

- HuCCT1/CD90+ cells are more proliferative and resistant to Gemcitabine compared to CD90− cells.
- Combining Gemcitabine with Galunisertib, a TGF-β1 inhibitor, synergistically reduces CD90+ cell proliferation.
- CD90+ cells express lower TGF-β1 levels but show increased resistance to chemotherapy.

## Abstract

Molecular mechanisms responsible for the poor prognosis in patients with intrahepatic cholangiocarcinoma (CCA) are still unknown, but stem cell marker Cluster Differentiation 90 (CD90) has been reported to be associated with a more aggressive cancer phenotype. In this scenario, the TGF-β1 signaling pathway likely has a role as master gene regulator. Aim of the study is to investigate the role of CD90 in iCCA aggressiveness. The molecular profile of HuCCT1/CD90+ and HuCCT1/CD90− cells was obtained through transcriptomic analysis (NGS). Bioinformatic data were confirmed in both cell lines by qRT-PCR and Western blot. Cells were treated with Gemcitabine in monotherapy or in combination with Galunisertib, a selective inhibitor of TGF-βRI, in 2D and 3D models. HuCCT1/CD90+ cells are more proliferative, less migratory, and resistant to Gemcitabine treatment. HuCCT1/CD90+ cells also express lower levels of TGF-β1 compared to /CD90− cell lines. Finally, HuCCT1/CD90+ cells are resistant to Gemcitabine, while the combination of Gemcitabine and Galunisertib displays a synergistic effect on HuCCT1/CD90+ cell proliferation. These results underline that CD90-induced Gemcitabine resistance can be overcome by adding a TGFβ1 inhibitor such as Galunisertib, thereby moving further toward a precision medicine approach in patients with iCCA.