Abstract
Human cytomegalovirus (HCMV) uses two major ways for virus dissemination: infection by cell-free virus and direct cell-to-cell spread. Neutralizing antibodies can efficiently inhibit infection by cell-free virus but mostly fail to prevent cell-to-cell transmission. Here, we show that the ‘molecular tweezer’ CLR01, a broad-spectrum antiviral agent, is not only highly active against infection with cell-free virus but most remarkably inhibits antibody-resistant direct cell-to-cell spread of HCMV. The inhibition of cell-to-cell spread by CLR01 was not limited to HCMV but was also shown for the alphaherpesviruses herpes simplex viruses 1 and 2 (HSV-1, -2). CLR01 is a rapid acting small molecule that inhibits HCMV entry at the attachment and penetration steps. Electron microscopy of extracellular virus particles indicated damage of the viral envelope by CLR01, which likely impairs the infectivity of virus particles. The rapid inactivation of viral particles by CLR01, the viral envelope as the main target, and the inhibition of virus entry at different stages are presumably the key to inhibition of cell-free virus infection and cell-to-cell spread by CLR01. Importance: While cell-free spread enables the human cytomegalovirus (HCMV) and other herpesviruses to transmit between hosts, direct cell-to-cell spread is thought to be more relevant for in vivo dissemination within infected tissues. Cell-to-cell spread is resistant to neutralizing antibodies, thus contributing to the maintenance of virus infection and virus dissemination in the presence of an intact immune system. Therefore, it would be therapeutically interesting to target this mode of spread in order to treat severe HCMV infections and to prevent dissemination of virus within the infected host. The molecular tweezer CLR01 exhibits broad-spectrum antiviral activity against a number of enveloped viruses and efficiently blocks antibody-resistant cell-to-cell spread of HCMV, thus representing a novel class of small molecules with promising antiviral activity.
Highlights
We found that CLR01 is highly active in preventing the infection of cells with cell-free human cytomegalovirus (HCMV) and efficiently abrogates the cell-to-cell spread of HCMV and of other herpesviruses
CLR01 directly targeted virus particles and did not affect the susceptibility of cells to infection with HCMV when cells were pretreated with CLR01 prior to infection (Supplementary Figure S1A) [22]
To better understand the reduced infection by about 30–40% compared with the medium control and compared consequences of CLR01 binding to the HCMV envelope, we investigated which entry step with human immunoglobulins (IgG, HCMV negative), the latter used as a control for non-HCMV-specific effects of antibodies on virus penetration (Figure 3A,B)
Summary
Direct cell-to-cell spread appears to be highly relevant for herpesvirus dissemination within the infected host. Several possible mechanisms of cell-to-cell spread are proposed for various viruses, including direct transfer of mature enveloped particles between cells at tight junctions or synapses as well as the transfer of subviral particles via partial fusion of cell membranes or syncytia formation [1,11,12]. The cell-associated spread of HCMV, as well as other herpesviruses, likely represents an important mechanism to overcome immune responses. Effective antivirals that directly target the cell-associated spread would be of high therapeutic interest for the treatment of HCMV infections, since HCMV is a highly relevant opportunistic pathogen for individuals with a compromised or immature immune system, such as transplant recipients, patients with acquired immunodeficiency disease syndrome (AIDS), or connatally infected children
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