Direct cellular reprogramming enables development of viral T antigen-driven Merkel cell carcinoma in mice.

Monique Verhaegen ,Ping Chen,Arul M Chinnaiyan,Paul W Harms,Thomas L Saunders,Li-Jyun Syu,Chia-Jen Liu,Andrzej A Dlugosz,John S Runge,Sunny Y Wong,Aaron M Udager,Kevin Hu,Marina Grachtchouk,Isaac Brownell,Michael C Kelly,Stephan Weidinger,Matthew Patrick ,Marcin Cieślik ,Julia J Van Goor ,L.c Tsoi ,H Zabawa ,Jóhann E Guðjónsson ,Jacob R Arche ,D Wilbert

doi:10.1172/jci152069

Abstract

Merkel cell carcinoma (MCC) is an aggressive neuroendocrine skin cancer that frequently carries an integrated Merkel cell polyomavirus (MCPyV) genome and expresses viral transforming antigens (TAgs). MCC tumor cells also express signature genes detected in skin-resident, postmitotic Merkel cells, including atonal bHLH transcription factor 1 (ATOH1), which is required for Merkel cell development from epidermal progenitors. We now report the use of in vivo cellular reprogramming, using ATOH1, to drive MCC development from murine epidermis. We generated mice that conditionally expressed MCPyV TAgs and ATOH1 in epidermal cells, yielding microscopic collections of proliferating MCC-like cells arising from hair follicles. Immunostaining of these nascent tumors revealed p53 accumulation and apoptosis, and targeted deletion of transformation related protein 53 (Trp53) led to development of gross skin tumors with classic MCC histology and marker expression. Global transcriptome analysis confirmed the close similarity of mouse and human MCCs, and hierarchical clustering showed conserved upregulation of signature genes. Our data establish that expression of MCPyV TAgs in ATOH1-reprogrammed epidermal cells and their neuroendocrine progeny initiates hair follicle–derived MCC tumorigenesis in adult mice. Moreover, progression to full-blown MCC in this model requires loss of p53, mimicking the functional inhibition of p53 reported in human MCPyV-positive MCCs.

Highlights

Merkel cell carcinoma (MCC) is an aggressive neuroendocrine skin tumor with a poor prognosis [1], up to 50% of patients with advanced disease respond to immunotherapy [2]
We generated and validated transgenic mouse strains with doxycycline-inducible co-expression of Merkel cell polyomavirus (MCPyV) sTAg and tLTAg carrying IRES-driven RFP and GFP reporters, respectively (Fig. 1A and Supplementary Fig. 1)
Histological analysis of sections from SLA mice collected two weeks after transgene induction revealed spatially restricted, atypical-appearing cellular aggregates near the normally quiescent hair follicle stem cell compartment called the bulge [29, 30] (Fig. 1). The cells in these aggregates contained scant cytoplasm, condensed chromosomes, and pyknotic nuclei (Fig. 1B), and they expressed ATOH1, the Merkel cell/MCC markers Keratin 8 (KRT8) and SOX2, proliferation marker Ki67, apoptosis marker cleaved caspase 3 (CC3), MCPyV tLTAg, and p53 (Fig. 1C), none of which were detectable at appreciable levels in control hair follicles (Supplementary Fig. 3)

Summary

Introduction

MCC is an aggressive neuroendocrine skin tumor with a poor prognosis [1], up to 50% of patients with advanced disease respond to immunotherapy [2]. Given the failure of conventional approaches to generate a mouse model of MCC, we set out to ascertain whether ATOH1 could be utilized as a novel tool to reprogram TAg-expressing epidermal cells into the Merkel cell lineage in adult mice, enabling the development of murine tumors resembling human MCC.

Results

Conclusion