The development of a staphylococcus aureus four antigen vaccine for use prior to elective orthopedic surgery.

Staphylococcus aureus (S. aureus) is a challenging bacterial pathogen which can cause a range of diseases, from mild skin infections, to more serious and invasive disease including deep or organ space surgical site infections, life-threatening bacteremia, and sepsis. S. aureus rapidly develops resistance to antibiotic treatments. Despite current infection control measures, the burden of disease remains high. The most advanced vaccine in clinical development is a 4 antigen S. aureus vaccine (SA4Ag) candidate that is being evaluated in a phase 2b/3 efficacy study in patients undergoing elective spinal fusion surgery (STaphylococcus aureus suRgical Inpatient Vaccine Efficacy [STRIVE]). SA4Ag has been shown in early phase clinical trials to be generally safe and well tolerated, and to induce high levels of bactericidal antibodies in healthy adults. In this review we discuss the design of SA4Ag, as well as the proposed clinical development plan supporting licensure of SA4Ag for the prevention of invasive disease caused by S. aureus in elective orthopedic surgical populations. We also explore the rationale for the generalizability of the results of the STRIVE efficacy study (patients undergoing elective open posterior multilevel instrumented spinal fusion surgery) to a broad elective orthopedic surgery population due to the common pathophysiology of invasive S. aureus disease and commonalties of patient and procedural risk factors for developing postoperative S. aureus surgical site infections.

Meningococcal carriage in Dutch adolescents and young adults; a cross-sectional and longitudinal cohort study.

OBJECTIVES: Current information on rates and dynamics of meningococcal carriage is essential for public health policy. This study aimed to determine meningococcal carriage prevalence, its risk factors and duration in the Netherlands, where meningococcal C vaccine coverage is >90%. Several methods to identify serogroups of meningococcal carriage isolates among adolescent and young adults were compared. METHODS: Oropharyngeal swabs were collected from 1715 participants 13-23 years of age in 2013-2014; 300 were prospectively followed over 8 months. Cultured isolates were characterized by Ouchterlony, real-time (rt-) PCR or whole-genome sequencing (WGS). Direct swabs were assessed by rt-PCR. Questionnaires on environmental factors and behaviour were also obtained. RESULTS: A meningococcal isolate was identified in 270/1715 (16%) participants by culture. Of MenB isolates identified by whole genome sequencing, 37/72 (51%) were correctly serogrouped by Ouchterlony, 46/51 (90%) by rt-PCR of cultured isolates, and 39/51 (76%) by rt-PCR directly on swabs. A sharp increase in carriage was observed before the age of 15 years. The age-related association disappeared after correction for smoking, level of education, frequent attendance to crowded social venues, kissing in the previous week and alcohol consumption. Three participants carried the same strain identified at three consecutive visits in an 8-month period. In these isolates, progressively acquired mutations were observed. CONCLUSIONS: Whole genome sequencing of culture isolates was the most sensitive method for serogroup identification. Based upon results of this study and risk of meningococcal disease, an adolescent meningococcal vaccination might include children before the age of 15 years to confer individual protection and potentially to establish herd protection.

Vaccine development to prevent Staphylococcus aureus surgical-site infections.

BACKGROUND: Staphylococcus aureus surgical-site infections (SSIs) are a major cause of poor health outcomes, including mortality, across surgical specialties. Despite current advances as a result of preventive interventions, the disease burden of S. aureus SSI remains high, and increasing antibiotic resistance continues to be a concern. Prophylactic S. aureus vaccines may represent an opportunity to prevent SSI. METHODS: A review of SSI pathophysiology was undertaken in the context of evaluating new approaches to developing a prophylactic vaccine to prevent S. aureus SSI. RESULTS: A prophylactic vaccine ideally would provide protective immunity at the time of the surgical incision to prevent initiation and progression of infection. Although the pathogenicity of S. aureus is attributed to many virulence factors, previous attempts to develop S. aureus vaccines targeted only a single virulence mechanism. The field has now moved towards multiple-antigen vaccine strategies, and promising results have been observed in early-phase clinical studies that supported the recent initiation of an efficacy trial to prevent SSI. CONCLUSION: There is an unmet medical need for novel S. aureus SSI prevention measures. Advances in understanding of S. aureus SSI pathophysiology could lead to the development of effective and safe prophylactic multiple-antigen vaccines to prevent S. aureus SSI.

Predicting pneumococcal community-acquired pneumonia in the emergency department: evaluation of clinical parameters.

The aim of this study was to quantify the value of clinical predictors available in the emergency department (ED) in predicting Streptococcus pneumoniae as the cause of community-acquired pneumonia (CAP). A prospective, observational, cohort study of patients with CAP presenting in the ED was performed. Pneumococcal aetiology of CAP was based on either bacteraemia, or S. pneumoniae being cultured from sputum, or urinary immunochromatographic assay positivity, or positivity of a novel serotype-specific urinary antigen detection test. Multivariate logistic regression was used to identify independent predictors and various cut-off values of probability scores were used to evaluate the usefulness of the model. Three hundred and twenty-eight (31.0%) of 1057 patients with CAP had pneumococcal CAP. Nine independent predictors for pneumococcal pneumonia were identified, but the clinical utility of this prediction model was disappointing, because of low positive predictive values or a small yield. Clinical criteria have insufficient diagnostic capacity to predict pneumococcal CAP. Rapid antigen detection tests are needed to diagnose S. pneumoniae at the time of hospital admission.

Use of a rapid test of pneumococcal colonization density to diagnose pneumococcal pneumonia.

BACKGROUND: There is major need for a more sensitive assay for the diagnosis of pneumococcal community-acquired pneumonia (CAP). We hypothesized that pneumococcal nasopharyngeal (NP) proliferation may lead to microaspiration followed by pneumonia. We therefore tested a quantitative lytA real-time polymerase chain reaction (rtPCR) on NP swab samples from patients with pneumonia and controls. METHODS: In the absence of a sensitive reference standard, a composite diagnostic standard for pneumococcal pneumonia was considered positive in South African human immunodeficiency virus (HIV)-infected adults hospitalized with radiographically confirmed CAP, if blood culture, induced good-quality sputum culture, Gram stain, or urinary Binax demonstrated pneumococci. Results of quantitative lytA rtPCR in NP swab samples were compared with quantitative colony counts in patients with CAP and 300 HIV-infected asymptomatic controls. RESULTS: Pneumococci were the leading pathogen identified in 76 of 280 patients with CAP (27.1%) using the composite diagnostic standard. NP colonization density measured by lytA rtPCR correlated with quantitative cultures (r = 0.67; P < .001). The mean lytA rtPCR copy number in patients with pneumococcal pneumonia was 6.0 log(10) copies/mL, compared with patients with CAP outside the composite standard (2.7 log(10) copies/mL; P < .001) and asymptomatic controls (0.8 log(10) copies/mL; P < .001). A lytA rtPCR density ≥8000 copies/mL had a sensitivity of 82.2% and a specificity of 92.0% for distinguishing pneumococcal CAP from asymptomatic colonization. The proportion of CAP cases attributable to pneumococcus increased from 27.1% to 52.5% using that cutoff. CONCLUSIONS: A rapid molecular assay of NP pneumococcal density performed on an easily available specimen may significantly increase pneumococcal pneumonia diagnoses in adults.

Relationship between IgG titers and opsonocytophagocytic activity of anti-pneumococcal antibodies after immunization with the 7-valent conjugate vaccine in allogeneic stem cell transplant.

The European Group for Blood and Marrow Transplantation has recently run a prospective, randomized trial comparing an early (3 months) vs a late (9 months) immunization program after allo-SCT with three doses of the conjugate 7-valent pneumococcal vaccine. This trial has shown that the response rate assessed 1 month after the third dose of conjugate vaccine was not inferior after an early vaccination vs a late vaccination. Part of the responder cohort of these patients (n=28) were chosen to assess the functionality of the anti-pneumococcal antibodies through an opsonophagocytic assay, 1 month after the third dose of conjugate vaccine. We have assessed the relationship between IgG titers measured by the pneumococcal ELISA and functional titers measured by opsonophagocytic assay of anti-pneumococcal antibodies. We found a significant correlation between titers for both assays, and conclude that the response to PCV7 after SCT induces functional antibodies.

A novel human papillomavirus type 6 neutralizing domain comprising two discrete regions of the major capsid protein L1.

We have mapped the binding sites on human papillomavirus (HPV) type 6 for three HPV 6-specific neutralizing monoclonal antibodies (mAbs). The critical binding residues were first identified by making HPV 11-like amino acid substitutions in the HPV 6 major capsid protein L1 and assaying the resulting virus-like particles (VLPs) for reactivity with the mAbs. To confirm the relevance of these residues for mAb binding, we demonstrated that HPV 6 type-specificity could be transferred to HPV 11 VLPs by making the appropriate HPV 6-like amino acid substitutions in the HPV 11 L1. Two binding regions were found. For one mAb, all critical residues are centered at residue 53, while for the other two mAbs, type-specific binding also requires a second site located more than 100 residues distal to the first. Both binding sites coincide with regions of L1 where the sequences of the closely related HPV 6 and 11 diverge. These regions are where the L1 sequences are the least well conserved among all HPV types and they have been implicated in type-specific binding for other HPV types. This suggests that clusters of diverged residues, surrounded by conserved L1 sequences, are presented on the surface of assembled particles and are responsible for eliciting critical humoral immune responses to the virus.

Neutralization of human papillomavirus type 11 (HPV-11) by serum from women vaccinated with yeast-derived HPV-11 L1 virus-like particles: correlation with competitive radioimmunoassay titer.

Neutralization of human papillomavirus type 11 (HPV-11) has been demonstrated using serum and cervical secretions from primates vaccinated with virus-like particles (VLPs). Theoretically, neutralizing antibodies could protect women from HPV infection. The immunogenicity of a yeast-derived HPV-11 L1 VLP vaccine was tested in women. Serum specimens were evaluated for HPV-11 titer by competitive radioimmunoassay (cRIA) and for neutralization by use of the athymic mouse xenograft system. Analysis of serum from 104 subjects showed a dose response in HPV-11 cRIA titers and neutralization. Overall, 68 (82.9%) of 82 postimmunization serum specimens from VLP recipients were 100% neutralizing when used in the assay at a 1:50 dilution. Of 69 serum specimens, 63 (91.3%) with cRIA titers >200 milliMerck units per milliliter were neutralizing. Immunization with HPV VLPs elicits a vigorous serum immune response in a high percentage of women. The HPV-11 cRIA titer appears to be a surrogate marker for neutralization.

Antibody, cytokine and cytotoxic T lymphocyte responses in chimpanzees immunized with human papillomavirus virus-like particles.

We evaluated antibody, cytokine (IFN-gamma, IL-5, TNF-alpha), and cytotoxic T lymphocyte (CTL) responses in chimpanzees immunized with monovalent or quadrivalent (HPV-6, -11, -16, -18) L1 virus-like particle (VLP) vaccines administered i.m. on aluminum hydroxyphosphate (alum) at weeks 0, 8 and 24. Maximum serum antibody titers to type-specific, neutralizing, conformational epitopes on HPV-11 or -16 L1 VLPs were detected by radioimmunoassay (RIA) four weeks after the second and third immunizations. HPV-11 and -16 neutralizing antibodies were also detected at similar time points with an Human papillomaviruses (HPV) neutralization assay using pseudovirions. Depending on the VLP type used for immunization, HPV type-specific cytokine responses were most frequently seen four weeks after the second or third immunizations and between weeks 44-52. Transient HPV-16 L1-specific CTL activity was observed only between weeks 16-24 in 3 of 22 (13.6%) chimpanzees immunized with HPV-16 L1 VLPs. These findings provide evidence that immunization with multivalent L1 VLPs on alum can evoke both neutralizing antibodies and Th1 and Th2 cytokine responses to several HPV types; however, induction of CTLs is infrequent.

A simple and efficient method for the monitoring of antigen-specific T cell responses using peptide pool arrays in a modified ELISpot assay.

In this study, we describe a simple and efficient method for both the monitoring of antigen-specific CD4 and CD8 T cell responses as well as the identification of novel CD4 and CD8 T cell epitopes using a modified ELISpot assay and pools of 20mer peptides. We have demonstrated that pools containing as many as 64 20mer peptides may be used to screen for CD4 and CD8 T cell responses to HPV16 L1, E1, and E7 in mice. Using arrays of pools of overlapping 20mer peptides, we have identified novel CD4 and CD8 epitopes in both HPV16L1 and HPV16E1 which are presented in Balb/c mice. We have further shown that the use of 20mer peptides is equivalent to using minimal 9mer epitopes for the stimulation of CD8 T cell responses in our assay. While our experiments are conducted in mice, the use of peptide pool arrays allows for the identification of epitope-specific responses using far fewer cells than is required for testing a panel of overlapping peptides individually, making this strategy particularly useful in clinical settings where immune cells may be limiting.

Hybrid papillomavirus L1 molecules assemble into virus-like particles that reconstitute conformational epitopes and induce neutralizing antibodies to distinct HPV types.

Human papillomavirus (HPV) hybrid virus-like particles (VLPs) were prepared using complementary regions of the major capsid L1 proteins of HPV-11 and -16. These hybrid L1 proteins were tested for assembly into VLPs, for presentation and mapping of conformational neutralizing epitopes, and as immunogens in rabbits and mice. Two small noncontiguous hypervariable regions of HPV-16 L1, when replaced into the HPV-11 L1 backbone, produced an assembly-positive hybrid L1 which was recognized by the type-specific, conformationally dependent HPV-16 neutralizing monoclonal antibody (N-MAb) H16.V5. Several new N-MAbs that were generated following immunization of mice with wild-type HPV-16 L1 VLPs also recognized this reconstructed VLP, demonstrating that these two hypervariable regions collectively constituted an immunodominant epitope. When a set of hybrid VLPs was tested as immunogens in rabbits, antibodies to both HPV-11 and -16 wild-type L1 VLPs were obtained. One of the hybrid VLPs containing hypervariable FG and HI loops of HPV-16 L1 replaced into an HPV-11 L1 background provoked neutralizing activity against both HPV-11 and HPV-16. In addition, conformationally dependent and type-specific MAbs to both HPV-11 and HPV-16 L1 VLP were obtained from mice immunized with hybrid L1 VLPs. These data indicated that hybrid L1 proteins can be constructed that retain VLP-assembly properties, retain type-specific conformational neutralizing epitopes, can map noncontiguous regions of L1 which constitute type-specific conformational neutralizing epitopes recognized by N-MAbs, and trigger polyclonal antibodies which can neutralize antigenically unrelated HPV types.

Neutralization of human papillomavirus (HPV) pseudovirions: a novel and efficient approach to detect and characterize HPV neutralizing antibodies.

The development of vaccines against human papillomaviruses (HPVs) has long been hampered by the inability to grow HPVs in tissue culture and the lack of an efficient neutralization assay. To date, less than 10% of more than 100 different HPV types can be grown in athymic and "SCID" mouse xenograft systems or raft culture systems. Recently, the in vitro generation of HPV pseudovirions and their use in neutralization assays were demonstrated. The major shortcomings of the current approaches to HPV neutralization are the lack of HPV virions for most types for the xenograft methods and the time-consuming and inefficient generation of infective pseudovirions for the latter methods, which precludes their use in large-scale HPV clinical trials or epidemiological studies. We describe here a novel and efficient approach to generating pseudovirions in which HPV virus-like particles (VLPs) are coupled to the beta-lactamase gene as a reporter. We show that it is not necessary to encapsidate the reporter gene constructs into the pseudovirions. Using sera from human volunteers immunized with HPV-11 VLPs expressed in yeast, we demonstrate that our novel neutralization assay compares favorably with the athymic mouse neutralization assay. Furthermore, our assay was used to define neutralizing monoclonal antibodies to HPV-6, which were previously unknown.

HPV11 mutant virus-like particles elicit immune responses that neutralize virus and delineate a novel neutralizing domain.

Characterization of the regions of human papillomaviruses (HPVs) that elicit neutralizing immune responses supports studies on viral infectivity and provides insight for the development and evaluation of prophylactic vaccines. HPV11 is a major etiologic agent of genital warts and a likely vaccine candidate. A conformationally dependent epitope for the binding of three neutralizing monoclonal antibodies (mAbs) has been mapped to residues G(131)T(132) of the L1 major capsid protein. The mAbs bind L1 only when it is assembled into virions or into virus-like particles (VLPs) that mimic the capsid structure. We were interested in identifying other domains of L1 that elicit neutralizing responses. To this end, we have generated a panel of mAbs against VLPs derived from HPV11 L1 harboring a G131S substitution. The new mAbs are unlike the neutralizing mAbs previously mapped to residues G(131)T(132) in that they bind both prototype and HPV11:G131S mutant VLPs. Some of the new mAbs neutralized virus in vitro. We have mapped epitopes for three of these new mAbs, as well as a neutralizing mAb generated against HPV11 virions, by measuring binding to HPV6 VLPs substituted with HPV11-like amino acids. Two regions are critical: one defined by HPV11 L1 residues 263-290 and the other by residues 346-349. mAbs H11.H3 and H11.G131S.G3 bind HPV6 VLPs with substitutions derived from the 346-349 region; in addition, H11.G131S.G3 binds HPV6 VLPs with substitutions derived only from the 263-290 region. Although H11.H3 does not bind HPV6 VLPs with substitutions derived from the 263-290 region, binding to HPV6 VLPs is enhanced when both sets of substitutions are present. mAbs H11.G131S.I1 and H11.G131S.K5 bind HPV6 VLPs with the 263-290 substitutions, but show little binding to HPV6 VLPs with the 346-349 substitutions. However, binding to HPV6 VLPs is enhanced when substitutions at both regions are present. The 346-349 region has not previously been described as eliciting a neutralizing response for any HPV type. In addition, the work demonstrates a complex binding site contributed by two distinct regions of L1.

Purification of virus-like particles of recombinant human papillomavirus type 11 major capsid protein L1 from Saccharomyces cerevisiae.

Recombinant major capsid protein, L1 (M(r) = 55,000), of human papillomavirus type 11 was expressed intracellularly at high levels in a galactose-inducible Saccharomyces cerevisiae expression system by an HPV6/11 hybrid gene. The capsid protein self-assembled into virus-like particles (VLPs) and accounted for 15% of the total soluble protein. A purification process was developed that consisted of two main steps: microfiltration and cation-exchange chromatography. The purified VLPs were 98% homogeneous, and the overall purification yield was 10%. The final product was characterized by several analytical methods and was highly immunogenic in mice.

The L1 major capsid protein of human papillomavirus type 11 recombinant virus-like particles interacts with heparin and cell-surface glycosaminoglycans on human keratinocytes.

The L1 major capsid protein of human papillomavirus (HPV) type 11, a 55-kDa polypeptide, forms particulate structures resembling native virus with an average particle diameter of 50-60 nm when expressed in the yeast Saccharomyces cerevisiae. We show in this report that these virus-like particles (VLPs) interact with heparin and with cell-surface glycosaminoglycans (GAGs) resembling heparin on keratinocytes and Chinese hamster ovary cells. The binding of VLPs to heparin is shown to exhibit an affinity comparable to that of other identified heparin-binding proteins. Immobilized heparin chromatography and surface plasmon resonance were used to show that this interaction can be specifically inhibited by free heparin and dextran sulfate and that the effectiveness of the inhibitor is related to its molecular weight and charge density. Sequence comparison of nine human L1 types revealed a conserved region of the carboxyl terminus containing clustered basic amino acids that bear resemblance to proposed heparin-binding motifs in unrelated proteins. Specific enzymatic cleavage of this region eliminated binding to both immobilized heparin and human keratinocyte (HaCaT) cells. Removal of heparan sulfate GAGs on keratinocytes by treatment with heparinase or heparitinase resulted in an 80-90% reduction of VLP binding, whereas treatment of cells with laminin, a substrate for alpha6 integrin receptors, provided minimal inhibition. Cells treated with chlorate or substituted beta-D-xylosides, resulting in undersulfation or secretion of GAG chains, also showed a reduced affinity for VLPs. Similarly, binding of VLPs to a Chinese hamster ovary cell mutant deficient in GAG synthesis was shown to be only 10% that observed for wild type cells. This report establishes for the first time that the carboxyl-terminal portion of HPV L1 interacts with heparin, and that this region appears to be crucial for interaction with the cell surface.

Human papillomavirus type 11 neutralization in the athymic mouse xenograft system: correlation with virus-like particle IgG concentration.

Neutralization of virus is likely to be necessary for development of an effective prophylactic vaccine against genital human papillomavirus (HPV) infection. Two New Zealand white rabbits were immunized with purified HPV type 11 (HPV 11) virions in Freund's adjuvant. An enzyme linked immunoassay (ELISA) was used to determine the quantity of IgG which recognized the HPV 11 major capsid protein (L1 protein) virus-like particles (VLPs) in the two anti-HPV 11 sera (serum A and serum B). The concentration of HPV 11 L1 VLP-specific IgG in the A and B sera were determined to be 37 and 90 micrograms per ml, respectively. The A and B sera were used in neutralization experiments in the athymic mouse xenograft system with known quantities of purified HPV 11 virions. The concentration of HPV 11 L1 VLP-specific IgG required to neutralize HPV 11 was determined for each antiserum. This concentration of IgG was approximately 700 to 900 ng per ml. This study demonstrates a positive correlation between the level of HPV 11 L1 VLP-specific IgG in animals immunized with HPV 11 virions and neutralization of HPV 11 in the athymic mouse model. Further studies are needed 1) to determine if sera or genital secretions from other species are neutralizing in the athymic mouse xenograft system, and 2) to determine if the VLP ELISA can be used as a reliable substitute for more cumbersome neutralization assays.

Human papillomavirus type 11 (HPV-11) neutralizing antibodies in the serum and genital mucosal secretions of African green monkeys immunized with HPV-11 virus-like particles expressed in yeast.

It has been shown previously that immunization of animals with recombinant virus-like particles (VLPs) consisting of the viral capsid proteins L1 or L1 plus L2 protected animals against experimental viral challenge. However, none of these experimental models addresses the issue of whether systemic immunization with VLPs elicits a neutralizing antibody response in the genital mucosa. Such a response may be necessary to protect the uterine cervix against infection with genital human papillomavirus (HPV) types. African green monkeys systemically immunized with HPV-11 VLPs expressed in Saccharomyces cerevisiae and formulated on aluminum adjuvant elicited high-titered HPV-11 VLP-specific serum antibody responses. Sera from these immunized monkeys neutralized HPV-11 in the athymic mouse xenograft system. Significant levels of HPV-11-neutralizing antibodies also were observed in cervicovaginal secretions. These findings suggest that protection against HPV infection of the uterine cervix may be possible through systemic immunization with HPV VLPs.

Expression of the major capsid protein of human papillomavirus type 11 in Saccharomyces cerevisae.

The major capsid protein L1 of papillomaviruses expressed recombinantly or in infected cells has the intrinsic ability to form virus-like particles (VLPs) which display conformational epitopes necessary to elicit high-titered, virus-neutralizing antibodies. We have shown previously that the L1 gene of human papillomavirus type 6a (HPV6) can be expressed efficiently in Saccharomyces cerevisae (Sc) as VLPs. However, when we attempted to express the L1 gene cloned from the closely related HPV11 in yeast, few VLPs were observed in crude lysates. The lower expression level of HPV11 L1 protein was found to result from a truncation of the HPV11 L1 mRNA. Since sequence requirements for transcriptional termination in yeast are still unclear, the HPV6 L1 gene was used as the basis for the complete synthetic reconstruction of the entire 1506-bp HPV11 L1 gene. Expression of this HPV6/11 hybrid L1 gene in yeast resulted in predominantly full-length L1 mRNA and a > 7-fold increased level of production of HPV11 VLPs compared to that expressed by the wild-type HPV11 L1 gene. The VLPs were shown to display the conformational epitopes important to elicit virus-neutralizing antibodies.

Sequence conservation within the major capsid protein of human papillomavirus (HPV) type 18 and formation of HPV-18 virus-like particles in Saccharomyces cerevisiae.

The major capsid protein L1 of human papillomaviruses (HPVs) has been identified as a promising candidate antigen for a prophylactic HPV vaccine. Since amino acid sequence heterogeneity has been demonstrated for the L1 genes within individual HPV types, nucleotide sequences of L1 were determined from six HPV-18 clinical isolates and the cervical carcinoma cell line SW756 and compared to the published HPV-18 prototype sequence. The sequences were almost identical between the clinical isolates and SW756 but differed markedly from the published prototype sequence. Resequencing the prototype HPV-18 revealed that these differences were due to sequencing artifacts of the prototype HPV-18 sequence archived in GenBank. Thus, the HPV-18 L1 genes seem to display a very high level of sequence conservation. The HPV-18 L1 gene derived from SW756 was expressed in Saccharomyces cerevisiae and self-assembly of the L1 protein into virus-like particles was demonstrated.

Protection against papillomavirus with a polynucleotide vaccine.

Genital infections with human papillomavirus (HPV) are increasingly recognized as a significant source of human disease; HPV is now implicated in up to 90% of cervical carcinomas. Neutralizing antibodies against papillomaviruses recognize conformational epitopes formed when viral capsid proteins assemble into virions or virus-like particles. Immunization with plasmid DNA encoding the major viral capsid protein L1 was studied as a means of inducing neutralizing antibodies and protection against virus challenge. In a cottontail rabbit papillomavirus (CRPV) model, immunization with plasmid DNA encoding L1 elicited conformationally specific neutralizing antibodies and provided immunity against papilloma formation upon challenge with CRPV. Immunization with DNA encoding the capsid protein may provide a means of protecting humans against HPV and would simplify the production of multivalent vaccines by combining plasmids that encode the viral capsid proteins of different strains. This may be of importance given the multiplicity of HPV types capable of causing disease.