A humanized mouse model of HPV-associated pathology driven by E7 expression.

Águeda Buitrago-Pérez,Marta Dueñas,José L Rodríguez-Peralto,Jesús M Paramio,Marcela Del Rio,Baki Akgül,Almudena Santos,Ramón García-Escudero,Fernando Larcher,Blanca Duarte,Juan Luis Santiago,Belén Lloveras,Almudena Holguín,Mariam Hachimi,Alan Storey,Catalina Ribas

doi:10.1371/journal.pone.0041743

Abstract

Human papillomavirus (HPV) is the causative agent of human cervical cancer and has been associated with oropharyngeal squamous cell carcinoma development. Although prophylactic vaccines have been developed, there is a need to develop new targeted therapies for individuals affected with malignant infected lesions in these locations, which must be tested in appropriate models. Cutaneous beta HPV types appear to be involved in skin carcinogenesis. Virus oncogenicity is partly achieved by inactivation of retinoblastoma protein family members by the viral E7 gene. Here we show that the E7 protein of cutaneous beta HPV5 binds pRb and promotes its degradation. In addition, we described an in vivo model of HPV-associated disease in which artificial human skin prepared using primary keratinocytes engineered to express the E7 protein is engrafted onto nude mice. Expression of E7 in the transplants was stably maintained for up to 6 months, inducing the appearance of lesions that, in the case of HPV16 E7, histologically resembled human anogenital lesions caused by oncogenic HPVs. Moreover, it was confirmed through biomarker expression analysis via immunodetection and/or quantitative PCR from mRNA and miRNA that the 16E7-modified engrafted skin shares molecular features with human HPV-associated pretumoral and tumoral lesions. Finally, our findings indicate a decrease of the in vitro capacity of HPV5 E7 to reduce pRb levels in vivo, possibly explaining the phenotypical differences when compared with 16E7-grafts. Our model seems to be a valuable platform for basic research into HPV oncogenesis and preclinical testing of HPV-associated antitumor therapies.

Highlights

Human papillomaviruses (HPVs) are small DNA viruses that show strict tropism for squamous epithelium, such as cutaneous epidermis or genital and oral mucosas
Cutaneous HPV5 E7 Protein Reduces pRb Protein Levels The transforming activities of the E7 oncogenes of HR-HPVs are mainly attributable to their ability to inhibit pocket proteins by reducing their protein levels
Using purified GST-E7-Flag proteins (Fig. S2A and B), we showed that E7 from HPV5 is able to interact with pRb in vitro using lysates obtained from the human skin HaCaT cell line (Fig. S2C) or purified His-pRb (Fig. S2D), as has been described [22,57,58]

Summary

Introduction

Human papillomaviruses (HPVs) are small DNA viruses that show strict tropism for squamous epithelium, such as cutaneous epidermis or genital and oral mucosas. Certain HPV types are related with human cancer of the skin, anogenital region, and oropharynx [1,2,3,4]. The link between cervical cancer (CC), the second most frequent cancer in women worldwide, and HPVs was first described in 1983 by Harald zur Hausen [5]. Today there is overwhelming evidence that certain high-risk (HR) types of mucosal HPVs cause CC. HPV types 16 and 18 are responsible for around 70% of CC worldwide [6]. In recent decades there has been a notable increase in the number of cases detected of HPV-positive cancers of the tonsils and base of the tongue and of oropharyngeal squamous cell carcinoma (OSCC)

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Conclusion