Production Strategies for Pentamer-Positive Subviral Dense Bodies as a Safe Human Cytomegalovirus Vaccine.

Gogesch Gogesch,Plachter Plachter,Büscher Büscher,Penner Penner,Grode Grode,Aydin Aydin,Krauter Krauter

doi:10.3390/vaccines7030104

Abstract

Infections with the human cytomegalovirus (HCMV) are associated with severe clinical manifestations in children following prenatal transmission and after viral reactivation in immunosuppressed individuals. The development of an HCMV vaccine has long been requested but there is still no licensed product available. Subviral dense bodies (DB) are immunogenic in pre-clinical models and are thus a promising HCMV vaccine candidate. Recently, we established a virus based on the laboratory strain Towne that synthesizes large numbers of DB containing the pentameric protein complex gH/gL/UL128-131 (Towne-UL130repΔGFP). The work presented here focuses on providing strategies for the production of a safe vaccine based on that strain. A GMP-compliant protocol for DB production was established. Furthermore, the DB producer strain Towne-UL130rep was attenuated by deleting the UL25 open reading frame. Additional genetic modifications aim to abrogate its capacity to replicate in vivo by conditionally expressing pUL51 using the Shield-1/FKBP destabilization system. We further show that the terminase inhibitor letermovir can be used to reduce infectious virus contamination of a DB vaccine by more than two orders of magnitude. Taken together, strategies are provided here that allow for the production of a safe and immunogenic DB vaccine for clinical testing.

Highlights

The human cytomegalovirus (HCMV) is well-recognized as a clinically important pathogen.Transmission of the virus during pregnancy and the resulting congenital HCMV infection are frequently associated with severe sequelae [1,2,3]
Separating dense bodies (DB) from virions is achieved by glycerol-tartrate gradient centrifugation of culture supernatants [37], DB fractions are still potentially contaminated by infectious virus
HCMV DB Material Can be Produced in a Large-Scale GMP-Compliant Manner

Summary

Introduction

The human cytomegalovirus (HCMV) is well-recognized as a clinically important pathogen.Transmission of the virus during pregnancy and the resulting congenital HCMV infection (cCMV) are frequently associated with severe sequelae [1,2,3]. The establishment of a vaccine for the prevention of HCMV-related complications in these settings is highly desirable [8]. DB contain major HCMV antigens and have been shown to be highly immunogenic without the addition of adjuvant [27,28,29,32,33,34,39]. DB contain major viral envelope proteins in their pre-fusion conformation, providing important antigens for the induction of a nabs response. One obstacle in the course of generating DB for clinical studies relates to the fact that the synthesis of these particles requires viral infection of culture cells. This work focused on reducing the viral load in culture supernatants used for DB purification and on the establishment of an attenuated HCMV seed strain that would be replication incompetent in vivo

Methods

Results

Discussion

Conclusion