# Human papillomavirus-associated head and neck squamous cell carcinoma cells lose viability during triggered myocyte lineage differentiation

## TL;DR

This study shows that human papillomavirus (HPV)-positive head and neck cancer cells can be triggered to differentiate into myocyte-like cells, which reduces their cancerous traits and viability.

## Contribution

The study is the first to report triggered myocyte-like differentiation in HPV+ head and neck squamous cell carcinoma cells.

## Key findings

- Differentiating HPV+ tumor cells showed reduced viability and loss of malignant characteristics.
- RNA-Seq and immunofluorescence confirmed myocyte-like differentiation with upregulated myofibril markers.
- HPV+ tumor tissue showed cells co-expressing myofibril markers, p16, and KRT17, supporting in vitro findings in human tissue.

## Abstract

Head and neck squamous cell carcinoma (HNSCC) is a highly malignant disease, and death rates have remained at approximately 50% for decades. New tumor-targeting strategies are desperately needed, and a previous report indicated the triggered differentiation of HPV-negative HNSCC cells to confer therapeutic benefits. Using patient-derived tumor cells, we created a similar HNSCC differentiation model of HPV+ tumor cells from two patients. We observed a loss of malignant characteristics in differentiating cell culture conditions, including irregularly enlarged cell morphology, cell cycle arrest with downregulation of Ki67, and reduced cell viability. RNA-Seq showed myocyte-like differentiation with upregulation of markers of myofibril assembly. Immunofluorescence staining of differentiated and undifferentiated primary HPV+ HNSCC cells confirmed an upregulation of these markers and the formation of parallel actin fibers reminiscent of myoblast-lineage cells. Moreover, immunofluorescence of HPV+ tumor tissue revealed areas of cells co-expressing the identified markers of myofibril assembly, HPV surrogate marker p16, and stress-associated basal keratinocyte marker KRT17, indicating that the observed myocyte-like in vitro differentiation occurs in human tissue. We are the first to report that carcinoma cells can undergo a triggered myocyte-like differentiation, and our study suggests that the targeted differentiation of HPV+ HNSCCs might be therapeutically valuable.