An Attenuated Strain of Human Cytomegalovirus for the Establishment of a Subviral Particle Vaccine

Abstract

Human cytomegalovirus (HCMV) infection is associated with severe disease conditions either following congenital transmission of the virus or viral reactivation in immunosuppressed individuals. Consequently, the establishment of a protective vaccine is of high medical need. Several candidates have been tested in preclinical and clinical studies, yet no vaccine has been licensed. Subviral dense bodies (DB) are a promising vaccine candidate. We have recently provided a GMP-compliant protocol for the production of DB, based on a genetically modified version of the HCMV laboratory strain Towne, expressing the pentameric complex of envelope protein gH-gL-pUL128-131 (Towne-UL130rep). In this work, we genetically attenuated Towne-UL130rep by abrogating the expression of the tegument protein pUL25 and by fusing the destabilizing domain ddFKBP to the N-terminus of the IE1- and IE2-proteins of HCMV. The resulting strain, termed TR-VAC, produced high amounts of DB under IE1/IE2 repressive conditions and concomitant supplementation of the viral terminase inhibitor letermovir to the producer cell culture. TR-VAC DB retained the capacity to induce neutralizing antibodies. A complex pattern of host protein induction was observed by mass spectrometry following exposure of primary human monocytes with TR-VAC DB. Human monocyte-derived dendritic cells (DC) moderately increased the expression of activation markers and MHC molecules upon stimulation with TR-VAC DB. In a co-culture with autologous T cells, the TR-VAC DB-stimulated DC induced a robust HCMV-specific T cell-activation and –proliferation. Exposure of donor-derived monocytic cells to DB led to the activation of a rapid innate immune response. This comprehensive data set thus shows that TR-VAC is an optimal attenuated seed virus strain for the production of a DB vaccine to be tested in clinical studies.

Epidemiological characteristics of Cytomegalovirus infection in children before and after the COVID-19 pandemic in Henan, China, 2016–2022

“… 7 Unfortunately, there is no effective vaccine to prevent the infection of CMV. 8 Therefore, CMV is a major and important pathogen in children all over the world, which is a serious threat to children’s health and brings a heavy burden to patients, families, and society. It is urgent to take precise prevention strategies for controlling the infection of CMV by understanding the situation of children CMV infection in different countries or regions.…”

Bacterial-Artificial-Chromosome-Based Genome Editing Methods and the Applications in Herpesvirus Research

“…The galK gene is a dual-selection cassette widely used in genome editing of herpesviruses [ 82 , 83 , 84 ]. The galK gene encodes galactokinase, which allows galK -deficient E. coli to grow on the medium with galactose as the sole carbon source during positive selection.…”

Subviral Dense Bodies of Human Cytomegalovirus Enhance Interferon-Beta Responses in Infected Cells and Impair Progeny Production

“…Quantitative real-time polymerase chain reaction (qPCR) using the TaqMan ® technology and quantification of viral progeny by limiting the dilution and IE1 protein staining were performed as described before [ 26 ].…”

“…For the pretreatment of cells with DB, 100 µL of UV-irradiated DBs was diluted in 2900 µL of 5% MEM and applied to cells for 2 h. Afterwards, the cells were washed twice with PBS and then infected. Quantitative real-time polymerase chain reaction (qPCR) using the TaqMan ® technology and quantification of viral progeny by limiting the dilution and IE1 protein staining were performed as described before [26].…”