Dense Bodies of Human Cytomegalovirus Induce both Humoral and Cellular Immune Responses in the Absence of Viral Gene Expression

Abstract

Infection of fibroblast cell cultures with human cytomegalovirus (HCMV) leads to the production of significant amounts of defective enveloped particles, termed dense bodies (DB). These noninfectious structures contain major antigenic determinants which are responsible for induction of both the humoral and the cellular immune response against HCMV. We tested the hypothesis that, by virtue of their unique antigenic and structural properties, DB could induce a significant immune response in the absence of infectious virus. Mice were immunized with gradient-purified DB, which were either left untreated or subjected to sequential rounds of sonication and freeze-thawing to prevent cellular entry. Titers of neutralizing antibodies induced by DB were in a range comparable to levels present in convalescent human sera. The virus-neutralizing antibody response was surprisingly durable, with neutralizing antibodies detected 12 months following primary immunization. The HCMV-specific major histocompatibility complex class I-restricted cytolytic T-cell (CTL) response was assayed using mice transgenic for the human HLA-A2 molecule. Immunization with DB led to high levels of HCMV-specific CTL in the absence of de novo viral protein synthesis. Maximal total cytolytic activity in mice immunized with DB was nearly as efficient as the cytolytic activity induced by a standard immunization with murine cytomegalovirus. Furthermore, DB induced a typical T-helper 1 (Th1)-dominated immune response in mice, as determined by cytokine and immunoglobulin G isotype analysis. Induction of humoral and cellular immune responses was achieved without the concomitant use of adjuvant. We thus propose that DB can serve as a basis for the future development of a recombinant nonreplicating vaccine against HCMV. Finally, such particles could be engineered for efficient delivery of antigens from other pathogens to the immune system.

Protein delivery by subviral particles of human cytomegalovirus

“…[4][5][6] We have previously shown that DB elicit strong humoral and cellular immune responses in mice. 5 Thus, by virtue of their biophysical properties, DB can be considered a potentially rewarding system for the development of both vaccines and therapeutics.…”

“…In a mouse model, DB have been shown to be highly immunogenic. 5 In contrast to viruslike particles from small animal viruses, used for vaccine purposes, DB are relatively large structures. Consequently, they have a high packaging capacity of protein molecules per particle.…”

“…CTLL were generated by immunizing HLA-A2/CD8 double-transgenic mice (a kind gift of L. Sherman, The Scripps Institute, LaJolla, USA) with DB resuspended in incomplete Freunds adjuvant, followed by spleenectonomy 10 days later, similar to procedures described previously. 5,10,11 Splenic cells were restimulated weekly with irradiated, pp65 495-503 -loaded feeder cells and TCGF according to established protocols. 10,11 IFN-g-based ELISPOT analyses were carried out as detailed by others.…”

Exogenous introduction of an immunodominant peptide from the non-structural IE1 protein of human cytomegalovirus into the MHC class I presentation pathway by recombinant dense bodies

“…Previous studies provided evidence that peptides derived from exogenously delivered, particle-associated HCMV proteins are presented by MHC class I molecules without requiring de novo protein synthesis (Besold et al, 2007;Manley et al, 2004;McLaughlin-Taylor et al, 1994;Pepperl et al, 2000;Reddehase et al, 1984;Riddell et al, 1991). It was previously unknown whether peptides not usually included in viral particles would also be introduced into MHC class I presentation.…”

“…Although several vaccine approaches have been developed, none have yet entered routine clinical practice (Pepperl-Klindworth & Plachter, 2006;Plotkin, 2004;Schleiss & Heineman, 2005;Zhong & Khanna, 2007). We have demonstrated that HCMV dense bodies (DB) provide a promising basis for vaccine development (Pepperl et al, 2000;Pepperl-Klindworth et al, 2002). DB are enveloped, subviral particles devoid of capsids and viral DNA.…”