Thomas C. Heineman

A randomized, controlled study in adults of the immunogenicity of a novel hepatitis B vaccine containing MF59 adjuvant

The safety and immunogenicity of a novel hepatitis B virus (HBV) vaccine containing recombinant PreS2 and S antigens combined with MF59 adjuvant (HBV/MF59) was evaluated in healthy adults (N=230) who were randomized to receive 2 or 3 immunizations of either the study vaccine or a licensed control vaccine (Recombivax HB). After a single immunization, 105 of 118 (89%) recipients of HBV/MF59 achieved protective serum levels of anti-HBs antibody (> 10 mIU/ml), compared with 13 of 110 (12%) recipients of licensed vaccine (P < 0.001). The geometric mean titer (GMT) after 2 doses of HBV/MF59 given 2 months apart (13,422 mIU/ml) was more than 5-fold higher than that following 3 doses of licensed vaccine given over 6 months (2,346 mIU/ml; P < 0.001). The GMT following 3 injections of HBV/MF59 (249,917 mIU/ml) was 100-fold higher than licensed vaccine (P < 0.001). Anti-PreS2 antibodies were elicited in over 90% of the subset of HBV/MF59 recipients tested. Both vaccines were well tolerated; transient, mild-to-moderate local inflammation was the major postinjection reaction.

A Phase 1 Study of 4 Live, Recombinant Human Cytomegalovirus Towne/Toledo Chimeric Vaccines

BACKGROUNDnHuman cytomegalovirus (HCMV) infection acquired in utero often results in severe consequences, including mental retardation and deafness. Although not evaluated for this indication, live attenuated HCMV vaccines based on the Towne strain are well-tolerated and have demonstrated moderate efficacy in other clinical settings.nnnMETHODSnTo produce live HCMV vaccine candidates that retain the excellent safety profile of the Towne strain but are more immunogenic, the genomes of the Towne strain and the unattenuated HCMV Toledo strain were recombined to yield 4 independent chimeric vaccine candidates. These vaccine candidates were evaluated in 20 HCMV-seropositive persons, in a phase 1, double-blinded, placebo-controlled trial. Participants received a single dose of vaccine or placebo, and the safety and tolerability of the vaccine candidates were evaluated.nnnRESULTSnThere was no difference in systemic symptoms between the vaccine and placebo recipients. As a group, vaccine recipients experienced more injection-site reactions than did placebo recipients; however, these were generally minor and short-lived. Vaccine virus could not be detected in blood, urine, or saliva samples obtained from any vaccine recipient.nnnCONCLUSIONSnThe Towne/Toledo chimeric vaccine candidates were well tolerated and did not cause systemic infection. Additional human trials are warranted to further evaluate the potential of these vaccine candidates as live virus vaccines.

Progress toward an elusive goal: current status of cytomegalovirus vaccines

Although infection with human cytomegalovirus (CMV) is ubiquitous and generally asymptomatic in most individuals, certain patient populations are at high risk for CMV-associated disease. These include HIV-infected individuals with AIDS, transplant patients, and newborn infants with congenital CMV infection. Immunity to CMV infection, both in the transplant setting and among women of childbearing age, plays a vital role in the control of CMV-induced injury and disease. Although immunity induced by CMV infection is not completely protective against reinfection, there is nevertheless a sound basis on which to believe that vaccination could help control CMV disease in high-risk patient populations. Evidence from several animal models of CMV infection indicates that a variety of vaccine strategies are capable of inducing immune responses sufficient to protect against CMV-associated illness following viral challenge. Vaccination has also proven effective in improving pregnancy outcomes following CMV challenge of pregnant guinea pigs, providing a ‘proof-of-principle’ relevant to human clinical trials of CMV vaccines. Although there are no licensed vaccines currently available for human CMV, progress toward this goal has been made, as evidenced by ongoing clinical trial testing of a number of immunization strategies. CMV vaccines currently in various stages of preclinical and clinical testing include: protein subunit vaccines; DNA vaccines; vectored vaccines using viral vectors, such as attenuated pox- and alphaviruses; peptide vaccines; and live attenuated vaccines. This review summarizes some of the obstacles that must be overcome in development of a CMV vaccine, and provides an overview of the current state of preclinical and clinical trial evaluation of vaccines for this important public health problem.

Varicella-Zoster Virus Infection Triggers Formation of an Interleukin-1β (IL-1β)-processing Inflammasome Complex

Innate cellular immunity is the immediate host response against pathogens, and activation of innate immunity also modulates the induction of adaptive immunity. The nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) are a family of intracellular receptors that recognize conserved patterns associated with intracellular pathogens, but information about their role in the host defense against DNA viruses is limited. Here we report that varicella-zoster virus (VZV), an alphaherpesvirus that is the causative agent of varicella and herpes zoster, induces formation of the NLRP3 inflammasome and the associated processing of the proinflammatory cytokine IL-1β by activated caspase-1 in infected cells. NLRP3 inflammasome formation was induced in VZV-infected human THP-1 cells, which are a transformed monocyte cell line, primary lung fibroblasts, and melanoma cells. Absent in melanoma gene-2 (AIM2) is an interferon-inducible protein that can form an alternative inflammasome complex with caspase-1 in virus-infected cells. Experiments in VZV-infected melanoma cells showed that NLRP3 protein recruits the adaptor protein ASC and caspase-1 to form an NLRP3 inflammasome complex independent of AIM2 protein and in the absence of free radical reactive oxygen species release. NLRP3 was also expressed extensively in infected skin xenografts in the severe combined immunodeficiency mouse model of VZV pathogenesis in vivo. We conclude that NLRP3 inflammasome formation is an innate cellular response to infection with this common pathogenic human herpesvirus.

Role of the Varicella-Zoster Virus gB Cytoplasmic Domain in gB Transport and Viral Egress

ABSTRACT To study the function of the varicella-zoster virus (VZV) gB cytoplasmic domain during viral infection, we produced a VZV recombinant virus that expresses a truncated form of gB lacking the C-terminal 36 amino acids of its cytoplasmic domain (VZV gB-36). VZV gB-36 replicates in noncomplementing cells and grows at a rate similar to that of native VZV. However, cells infected with VZVgB-36 form extensive syncytia compared to the relatively small syncytia formed during native VZV infection. In addition, electron microscopy shows that very little virus is present on the surfaces of cells infected with VZV gB-36, while cells infected with native VZV exhibit abundant virions on the cell surface. The C-terminal 36 amino acids of the gB cytoplasmic domain have been shown in transfection-based experiments to contain both an endoplasmic reticulum-to-Golgi transport signal (the C-terminal 17 amino acids) and a consensus YXXφ (where Y is tyrosine, X is any amino acid, and φ is any bulky hydrophobic amino acid) signal sequence (YSRV) that mediates the internalization of gB from the plasma membrane. As predicted based on these data, gB-36 expressed during the infection of cultured cells is transported inefficiently to the Golgi. Despite lacking the YSRV signal sequence, gB-36 is internalized from the plasma membrane; however, in contrast to native gB, it fails to localize to the Golgi. Therefore, the C-terminal 36 amino acids of the VZV gB cytoplasmic domain are required for normal viral egress and for both the pre- and post-Golgi transport of gB.

Cytoplasmic Domain Signal Sequences That Mediate Transport of Varicella-Zoster Virus gB from the Endoplasmic Reticulum to the Golgi

ABSTRACT Normal herpesvirus assembly and egress depend on the correct intracellular localization of viral glycoproteins. While several post-Golgi transport motifs have been characterized within the cytoplasmic domains of various viral glycoproteins, few specific endoplasmic reticulum (ER)-to-Golgi transport signals have been described. We report the identification of two regions within the 125-amino-acid cytoplasmic domain of Varicella-Zoster virus gB that are required for its ER-to-Golgi transport. Native gB or gB containing deletions and specific point mutations in its cytoplasmic domain was expressed in mammalian cells. ER-to-Golgi transport of gB was assessed by indirect immunofluorescence and by the acquisition of Golgi-dependent posttranslational modifications. These studies revealed that the ER-to-Golgi transport of gB requires a nine-amino-acid region (YMTLVSAAE) within its cytoplasmic domain. Mutations of individual amino acids within this region markedly impaired the transport of gB from the ER to the Golgi, indicating that this domain functions by a sequence-dependent mechanism. Deletion of the C-terminal 17 amino acids of the gB cytoplasmic domain was also shown to impair the transport of gB from the ER to the Golgi. However, internal mutations within this region did not disrupt the transport of gB, indicating that its function during gB transport is not sequence dependent. Native gB is also transported to the nuclear membrane of transfected cells. gB lacking as many as 67 amino acids from the C terminus of its cytoplasmic domain continued to be transported to the nuclear membrane at apparently normal levels, indicating that the cytoplasmic domain of gB is not required for nuclear membrane localization.