Production of Cytomegalovirus Dense Bodies by Scalable Bioprocess Methods Maintains Immunogenicity and Improves Neutralizing Antibody Titers

Schneider-Ohrum, Kirsten; Cayatte, Corinne; Liu, Yi; Wang, Zhaoti; Irrinki, Alivelu; Cataniag, Floro; Nguyen, Nga; Lambert, Stacie; Liu, Hui; Aslam, Shahin; Duke, Gregory; McCarthy, Michael; McCormick, Louise

doi:10.1128/jvi.00463-16

Cited by 12 publications

(26 citation statements)

References 57 publications

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“…The antibody response resulting from vaccination with these DBs was also shown to prevent infection of both fibroblasts and epithelial cells by the clinical CMV isolate VR1814 in cell culture (112). Methods to scale up the processes of manufacture and purification have been previously described (115). The Serum Institute of India has licensed the technology for production of a DB vaccine, designated CAP CMV001, from Vakzine Projekt Management GmbH.…”

Section: Vaccines In Preclinical Developmentmentioning

confidence: 99%

Progress toward Development of a Vaccine against Congenital Cytomegalovirus Infection

Schleiss

Permar

Plotkin

2017

Clin Vaccine Immunol

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A vaccine against congenital human cytomegalovirus (CMV) infection is a major public health priority. Congenital CMV causes substantial long-term morbidity, particularly sensorineural hearing loss (SNHL), in newborns, and the public health impact of this infection on maternal and child health is underrecognized. Although progress toward development of a vaccine has been limited by an incomplete understanding of the correlates of protective immunity for the fetus, knowledge about some of the key components of the maternal immune response necessary for preventing transplacental transmission is accumulating. Moreover, although there have been concerns raised about observations indicating that maternal seropositivity does not fully prevent recurrent maternal CMV infections during pregnancy, it is becoming increasing clear that preconception immunity does confer some measure of protection against both CMV transmission and CMV disease (if transmission occurs) in the newborn infant. Although the immunity to CMV conferred by both infection and vaccination is imperfect, there are encouraging data emerging from clinical trials demonstrating the immunogenicity and potential efficacy of candidate CMV vaccines. In the face of the knowledge that between 20,000 and 30,000 infants are born with congenital CMV in the United States every year, there is an urgent and compelling need to accelerate the pace of vaccine trials. In this minireview, we summarize the status of CMV vaccines in clinical trials and provide a perspective on what would be required for a CMV immunization program to become incorporated into clinical practice.

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Section: Vaccines In Preclinical Developmentmentioning

confidence: 99%

Progress toward Development of a Vaccine against Congenital Cytomegalovirus Infection

Schleiss

Permar

Plotkin

2017

Clin Vaccine Immunol

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“…We and others have shown that DBs are highly immunogenic, bearing the exceptional capacity to induce both humoral and cellular immune responses without requiring an adjuvant (25,28,29,31,33,34). DBs induce maturation and activation of immature dendritic cells in addition, likely explaining their impact on the priming of immune responses (30).…”

Section: Discussionmentioning

confidence: 99%

“…This is likely related to the maturation and activation of dendritic cells (DCs) following incubation of these cells with DBs (30). DB application also induces high levels of NT-abs against HCMV infection (25,28,29,31). Since viral envelope proteins are inserted in their functional conformation into the envelopes of DBs, these particles likely induce humoral responses comparable to those induced by viral infection.…”

mentioning

confidence: 99%

“…All previous experiments using purified DBs as immunogens have been conducted with particles from laboratory HCMV strains which are devoid of the PC (25,(27)(28)(29)(31)(32)(33)(34). Here, we attempted to analyze DBs containing the PC (PC-positive) alongside PC-negative DBs in order to address the contribution of PC-specific antibodies to DB-induced neutralizing antibody responses.…”

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confidence: 99%

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Dense Bodies of a gH/gL/UL128/UL130/UL131 Pentamer-Repaired Towne Strain of Human Cytomegalovirus Induce an Enhanced Neutralizing Antibody Response

Lehmann

Falk²,

Büscher

et al. 2019

J Virol

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The development of a vaccine against human cytomegalovirus infection (HCMV) is a high-priority medical goal. The viral pentameric protein complex consisting of glycoprotein H (gH)/gL/UL128-131A (PC) is considered to be an important vaccine component. Its relevance to the induction of a protective antibody response is, however, still a matter of debate. We addressed this issue by using subviral dense bodies (DBs) of HCMV. DBs are exceptionally immunogenic. Laboratory HCMV strain DBs harbor important neutralizing antibody targets, like the glycoproteins B, H, L, M, and N, but they are devoid of the PC. To be able to directly compare the impact of the PC on the levels of neutralizing antibody (NT-abs) responses, a PC-positive variant of the HCMV laboratory strain Towne was established by bacterial artificial chromosome (BAC) mutagenesis (Towne-UL130rep). This strain synthesized PC-positive DBs upon infection of fibroblasts. These DBs were used in side-by-side immunizations with PC-negative Towne DBs. Mouse and rabbit sera were tested to address the impact of the PC on DB immunogenicity. The neutralizing antibody response to PC-positive DBs was superior to that of PC-negative DBs, as tested on fibroblasts, epithelial cells, and endothelial cells and for both animal species used. The experiments revealed the potential of the PC to enhance the antibody response against HCMV. Of particular interest was the finding that PC-positive DBs induced an antibody response that blocked the infection of fibroblasts by a PC-positive viral strain more efficiently than sera following immunizations with PC-negative particles. IMPORTANCE Infections with the human cytomegalovirus (HCMV) may cause severe and even life-threatening disease manifestations in newborns and immunosuppressed individuals. Several strategies for the development of a vaccine against this virus are currently pursued. A critical question in this respect refers to the antigenic composition of a successful vaccine. Using a subviral particle vaccine candidate, we show here that one protein complex of HCMV, termed the pentameric complex (PC), enhances the neutralizing antibody response against viral infection of different cell types. We further show for the first time that this not only relates to the infection of epithelial or endothelial cells; the presence of the PC in the particles also enhanced the neutralizing antibody response against the infection of fibroblasts by HCMV. Together, these findings argue in favor of including the PC in strategies for HCMV vaccine development.

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“…However, an impact of pUL25 loss either on the morphogenesis of DBs within the cell or on the yield of DBs that could be purified from infected cell culture supernatants was not seen (not shown). DBs are a promising vaccine candidate (31)(32)(33)(34)(35). Production of DBs depends on their purification from infected cell cultures.…”

Section: Discussionmentioning

confidence: 99%

The Abundant Tegument Protein pUL25 of Human Cytomegalovirus Prevents Proteasomal Degradation of pUL26 and Supports Its Suppression of ISGylation

Zimmermann

Büscher

Krauter

et al. 2018

J Virol

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The tegument of human cytomegalovirus (HCMV) virions contains proteins that interfere with both the intrinsic and the innate immunity. One protein with a thus far unknown function is pUL25. The deletion of pUL25 in a viral mutant (Towne-ΔUL25) had no impact on the release of virions and subviral dense bodies or on virion morphogenesis. Proteomic analyses showed few alterations in the overall protein composition of extracellular particles. A surprising result, however, was the almost complete absence of pUL26 in virions and dense bodies of Towne-ΔUL25 and a reduction of the large isoform pUL26-p27 in mutant virus-infected cells. pUL26 had been shown to inhibit protein conjugation with the interferon-stimulated gene 15 protein (ISG15), thereby supporting HCMV replication. To test for a functional relationship between pUL25 and pUL26, we addressed the steady-state levels of pUL26 and found them to be reduced in Towne-ΔUL25-infected cells. Coimmunoprecipitation experiments proved an interaction between pUL25 and pUL26. Surprisingly, the overall protein ISGylation was enhanced in Towne-ΔUL25-infected cells, thus mimicking the phenotype of a pUL26-deleted HCMV mutant. The functional relevance of this was confirmed by showing that the replication of Towne-ΔUL25 was more sensitive to beta interferon. The increase of protein ISGylation was also seen in cells infected with a mutant lacking the tegument protein pp65. Upon retesting, we found that pUL26 degradation was also increased when pp65 was unavailable. Our experiments show that both pUL25 and pp65 regulate pUL26 degradation and the pUL26-dependent reduction of ISGylation and add pUL25 as another HCMV tegument protein that interferes with the intrinsic immunity of the host cell.IMPORTANCE Human cytomegalovirus (HCMV) expresses a number of tegument proteins that interfere with the intrinsic and the innate defense mechanisms of the cell. Initial induction of the interferon-stimulated gene 15 protein (ISG15) and conjugation of proteins with ISG15 (ISGylation) by HCMV infection are subsequently attenuated by the expression of the viral IE1, pUL50, and pUL26 proteins. This study adds pUL25 as another factor that contributes to suppression of ISGylation. The tegument protein interacts with pUL26 and prevents its degradation by the proteasome. By doing this, it supports its restrictive influence on ISGylation. In addition, a lack of pUL25 enhances the levels of free ISG15, indicating that the tegument protein may interfere with the interferon response on levels other than interacting with pUL26. Knowledge obtained in this study widens our understanding of HCMV immune evasion and may also provide a new avenue for the use of pUL25-negative strains for vaccine production.

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Production of Cytomegalovirus Dense Bodies by Scalable Bioprocess Methods Maintains Immunogenicity and Improves Neutralizing Antibody Titers

Cited by 12 publications

References 57 publications

Progress toward Development of a Vaccine against Congenital Cytomegalovirus Infection

Progress toward Development of a Vaccine against Congenital Cytomegalovirus Infection

Dense Bodies of a gH/gL/UL128/UL130/UL131 Pentamer-Repaired Towne Strain of Human Cytomegalovirus Induce an Enhanced Neutralizing Antibody Response

The Abundant Tegument Protein pUL25 of Human Cytomegalovirus Prevents Proteasomal Degradation of pUL26 and Supports Its Suppression of ISGylation

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