# Niclosamide and Palbociclib Act Synergistically to Reduce Cholangiocarcinoma Cell Viability In Vitro and Inhibit Tumour Growth in a Mouse Model

## TL;DR

Niclosamide and Palbociclib together reduce the viability of cholangiocarcinoma cells and inhibit tumor growth in mice, offering a promising new treatment strategy.

## Contribution

The study identifies a synergistic drug combination of Niclosamide and Palbociclib for treating cholangiocarcinoma.

## Key findings

- Niclosamide reduces the viability of cholangiocarcinoma cells more than normal bile duct cells.
- The combination of Niclosamide and Palbociclib is more effective than either drug alone in reducing tumor growth in mice.
- Niclosamide reduces PRH protein expression via a proteasome-dependent mechanism, but PRH knockout does not affect Niclosamide sensitivity.

## Abstract

Cancer of the bile duct, or cholangiocarcinoma, is increasing in incidence and mortality. Despite the emergence of new targeted therapies that benefit a minority of patients whose tumours carry specific mutations, the outcomes for most patients are very poor, and new treatment strategies are urgently required. Here we show that in preclinical research cholangiocarcinoma cells are more sensitive to Niclosamide, a drug that is commonly used to treat parasitic infections, than normal bile duct cells. Importantly, the combination of Niclosamide with Palbociclib, a drug that blocks cell division, is significantly more effective than either drug alone. Further studies will be required to determine whether this drug combination is effective in the clinic.

Background: Despite the emergence of new treatment modalities, including targeted therapies that are of benefit to patients whose tumours carry specific mutations, the prognosis for most patients with cholangiocarcinoma remains poor. Novel therapeutic approaches that can benefit the majority of patients whose tumour cells do not carry targetable mutations are urgently needed. Results: To identify mutation-agnostic treatment approaches, we screened a library of well-tolerated off-patent drugs against cholangiocarcinoma cells and normal biliary epithelial cells. The screen identified Niclosamide as a drug that reduces the viability of multiple cholangiocarcinoma cell lines but has a lesser effect on normal primary biliary epithelial cells. Moreover, Niclosamide treatment reduces the growth of cholangiocarcinoma tumour cells as tumour spheroids in vitro and reduces the growth of cholangiocarcinoma cells as tumours in a xenograft mouse model of this disease. Through a proteasome-dependent mechanism, Niclosamide treatment reduces the expression of the Proline-Rich Homeodomain (PRH) protein, a transcription factor which acts as an oncoprotein in cholangiocarcinoma cells. However, PRH knockout does not alter the sensitivity of cholangiocarcinoma cells to Niclosamide, indicating that this drug is not dependent on PRH to reduce cell viability. Interestingly, the CDK4/6 kinase inhibitor Palbociclib selectively reduces the viability of cholangiocarcinoma cell lines compared to normal biliary epithelial cells and, importantly, Palbociclib synergises with Niclosamide to reduce cholangiocarcinoma cell viability in vitro as well as to reduce tumour growth in a mouse xenograft model. Conclusion: These preclinical results suggest that the combination of Niclosamide and an inhibitor of CDK4/6 is worthy of clinical evaluation as a potential treatment for this disease.