Population Pharmacokinetic Modeling of VL-2397, a Novel Systemic Antifungal Agent: Analysis of a Single- and Multiple-Ascending-Dose Study in Healthy Subjects.

VL-2397, a novel, systemic antifungal agent, has potent in vitro and in vivo fungicidal activity against Aspergillus species. Plasma concentrations from a phase 1 study were used to construct a population pharmacokinetic (PPK) model for VL-2397. Healthy subjects aged 18 to 55 years received single doses of VL-2397, ranging from 3 to 1,200 mg, multiple daily doses of 300, 600, or 1,200 mg for 7 days, or 300 mg three times/day for 7 days followed by 600 mg daily for 21 days. Plasma samples were collected throughout the dosing intervals. Sixty-six subjects provided 1,908 concentrations. Drug concentrations over time were increased less than dose proportionally for doses above 30 mg. Dose-normalized concentrations plotted over time did not overlap. A 3-compartment nonlinear saturable binding model fit the data well. Clearance increased with dose, and mean values ranged from 0.4 liters/h at 3 mg to 8.5 liters/h at 1,200 mg. Mean volume in the central compartment ranged from 4.8 to 6.9 liters across doses. In the first 24 h, once-daily dosing results in a rapid decrease in concentrations by hour 16 to approximately 1 mg/liter, regardless of dose, with slow clearance over time. Administration of 300 mg every 8 h achieved concentrations above 1 mg/liter over an entire 24-h period. There was a significant relationship between body surface area and clearance. The data suggest that VL-2397 has nonlinear saturable binding kinetics. Protein binding is the likely primary source of the nonlinearity. The PPK model can now be used to optimize dosing by bridging the kinetics to efficacious pharmacodynamic targets.

The Siderophore Transporter Sit1 Determines Susceptibility to the Antifungal VL-2397.

VL-2397 (previously termed ASP2397) is an antifungal, aluminum-chelating cyclic hexapeptide with a structure analogous to that of ferrichrome-type siderophores, whereby replacement of aluminum by iron was shown to decrease the antifungal activity of this compound. Here, we found that inactivation of an importer for ferrichrome-type siderophores, termed Sit1, renders Aspergillus fumigatus resistant to VL-2397. Moreover, expression of the endogenous sit1 gene under the control of a xylose-inducible promoter (to uncouple sit1 expression from iron repression) combined with C-terminal tagging with a fluorescent protein demonstrated localization of Sit1 in the plasma membrane and xylose-dependent VL-2397 susceptibility. This underlines that Sit1-mediated uptake is essential for VL-2397 susceptibility. Under xylose-induced sit1 expression, VL-2397 also retained antifungal activity after replacing aluminum with iron, which demonstrates that VL-2397 bears antifungal activity independent of cellular aluminum importation. Analysis of sit1 expression indicated that the reduced antifungal activity of the iron-chelated VL-2397 is caused by downregulation of sit1 expression by the imported iron. Furthermore, we demonstrate that defects in iron homeostatic mechanisms modulate the activity of VL-2397. In contrast to A. fumigatus and Candida glabrata, Saccharomyces cerevisiae displays intrinsic resistance to VL-2397 antifungal activity. However, expression of sit1 from A. fumigatus, or its homologue from C. glabrata, resulted in susceptibility to VL-2397, which suggests that the intrinsic resistance of S. cerevisiae is based on lack of uptake and that A. fumigatus, C. glabrata, and S. cerevisiae share an intracellular target for VL-2397.

First-in-Human Phase 1 Study To Assess Safety, Tolerability, and Pharmacokinetics of a Novel Antifungal Drug, VL-2397, in Healthy Adults.

VL-2397 is an antifungal drug with a novel mechanism of action, rapid fungicidal in vitro activity, and potent in vivo activity against Aspergillus fumigatus, including azole-resistant strains. VL2397-101, a phase 1 first-in-human, randomized, double-blind, placebo-controlled dose-escalation study, was conducted in healthy adults to determine the safety, tolerability, and pharmacokinetics (PK) of single and multiple ascending intravenous (i.v.) doses of VL-2397. All dosing cohorts were fully enrolled; all subjects completed the safety follow-up. A safety committee reviewed the safety data for each dosing cohort prior to recommending the initiation of each subsequent cohort. No serious adverse events (SAEs) occurred; the majority of treatment-emergent adverse events (TEAEs) were mild and self-limited. The most common drug-related TEAEs were infusion site reactions. No clinically concerning trends were noted in vital signs, electrocardiograms, physical examinations, or safety laboratory results. Following single infusions of VL-2397, the overall and maximum exposures rose less than proportionally with increasing doses from 3 mg to 1,200 mg as indicated by area under the concentration-time curve over 24 h (AUC24) and maximum concentration (Cmax). No signs of VL-2397 accumulation were observed following i.v. infusions of 300, 600, and 1,200 mg every 24 h (q24h) for 7 days. Renal elimination played a major role in total body clearance, with up to 47% of unmetabolized drug in urine 24 h after administration at single doses of >30 mg. Overall, VL-2397 dosing in the study appeared to be safe and well tolerated in the healthy subjects. The safety profile, consistent PK, and lack of drug accumulation support further development of VL-2397 in patients with invasive aspergillosis.

A randomised, placebo-controlled phase 3 study to evaluate the efficacy and safety of ASP0113, a DNA-based CMV vaccine, in seropositive allogeneic haematopoietic cell transplant recipients.

BACKGROUND: Cytomegalovirus (CMV) is a complication of allogeneic haematopoietic cell transplantation (allo-HCT). ASP0113, a DNA-based vaccine, contains two plasmids encoding human CMV glycoprotein B and phosphoprotein 65 (pp65). We assessed ASP0113 in CMV-seropositive allo-HCT recipients. METHODS: In this phase 3, randomised, placebo-controlled study, CMV-seropositive allo-HCT recipients were randomly assigned (1:1) via interactive response technology to receive five injections of 1 mL of 5 mg/mL ASP0113 or placebo. The pharmacist and designated staff were unblinded. Masked syringes maintained the blind for patients and study personnel. Efficacy and safety analyses included patients who received ≥1 dose of ASP0113/placebo. The primary efficacy endpoint was the proportion of allo-HCT recipients with composite all-cause mortality and adjudicated CMV end-organ disease (EOD) by 1 year post-transplant. ClinicalTrials.gov: NCT01877655 (not recruiting). FINDINGS: Patients were recruited between Sept 11, 2013 and Sept 21, 2016. Overall, 501 patients received ≥1 dose of ASP0113 (n = 246) or placebo (n = 255). The proportion of patients with composite all-cause mortality and adjudicated CMV EOD by 1 year post-transplant was 35.4% (n = 87) with ASP0113 and 30•2% (n = 77) with placebo (odds ratio 1.27; 95% confidence interval: 0.87 to 1.85; p = 0.205). Incidence of injection site-related treatment-emergent adverse events (TEAEs) was higher with ASP0113 than placebo. Overall incidence and severity of other TEAEs was similar between groups. T-cell response to pp65 increased over time and was greater with placebo than ASP0113 (p = 0.027). INTERPRETATION: ASP0113 did not reduce overall mortality or CMV EOD by 1 year post-transplant. Safety findings were similar between groups. FUNDING: Astellas Pharma Global Development, Inc .

Pharmacokinetics and Immunogenicity of ASP0113 in CMV-Seronegative Dialysis Patients and CMV-Seronegative and -Seropositive Healthy Subjects.

Cytomegalovirus (CMV) infection causes significant morbidity and mortality in immunocompromised transplant patients. ASP0113, a first-in-class DNA vaccine containing plasmids encoding CMV phosphoprotein 65 and glycoprotein B (gB), was evaluated in a phase 1b, subject-blinded study in CMV-seropositive (n = 13) and CMV-seronegative (n = 12) healthy and CMV-seronegative dialysis subjects (n = 12) randomized to ASP0113 or placebo. End points included pharmacokinetics, anti-gB antibody levels, phosphoprotein 65-specific T-cell responses measured by ex vivo enzyme-linked immune absorbent spot (ELISpot) assay and 10-day cultured ELISpot and Stat T-cell activation assays, and safety. ASP0113 concentrations peaked at 2-10 and 24-48 hours; the pharmacokinetics were similar across groups. No group demonstrated significant anti-gB antibody responses. T-cell responder rates in the cultured ELISpot assay were 8/12 (66.7%, 95%CI 35% to 90%) and 4/12 (33.3%, 95%CI 10% to 65%) in CMV-seronegative healthy subjects and dialysis patients, respectively, whereas ex vivo ELISpot assay response rates were 4/11 (36.4%, 95%CI 11% to 69%) and 0/12, respectively. Responses peaked at week 27, with lower magnitude observed in CMV-seronegative dialysis patients versus CMV-seronegative healthy subjects. No serious adverse events occurred; the most common adverse event in ASP0113-vaccinated patients was injection-site pain (64.9%). Some CMV-seronegative healthy subjects and dialysis patients had T-cell responses; no humoral responses were detected.

Evaluation of a plasmid DNA-based anthrax vaccine in rabbits, nonhuman primates and healthy adults.

VCL-AB01, a cationic lipid-formulated plasmid DNA (pDNA)-based vaccine that contains genes encoding genetically detoxified Bacillus anthracis protective antigen (PA) and lethal factor (LF), was assessed in a Phase 1, dose-escalating clinical trial in healthy adults for safety and immunogenicity, and in nonhuman primates for immunogenicity and efficacy against challenge with a lethal dose of B. anthracis spores. Healthy 18-45 year old subjects were randomly assigned to receive either the investigational vaccine containing 0.2 mg, 0.6 mg, or 2 mg of total pDNA per dose, or saline placebo, administered at 0, 1 and 2 months. The 0.2 mg and 0.6 mg dose levels were generally well tolerated; however, dose-limiting reactogenicity was observed among subjects given the first 2 mg dose and the remaining two injections in the 2 mg group were reduced to 0.6 mg. Dose-related increases in seroconversion frequencies were observed. Overall, 10%, 33.3% and 80% of subjects in the 0.2, 0.6 and 2 mg groups, respectively, developed antibodies to PA and/or LF as measured by ELISA; however, antibodies with toxin neutralizing activity (TNA) were detected in only one subject. In monkeys that received a 0.6 mg dose three times at 2 week intervals, low levels of antibodies were detected by ELISA but not by the TNA assay in all animals just prior to challenge. Despite the absence of TNA, 75% animals survived the lethal challenge. In summary, VCL-AB01 was generally well tolerated in humans at a dose that provided immunity in monkeys despite the lack of robust TNA titers in either species.

Cross-reactive neuraminidase antibodies afford partial protection against H5N1 in mice and are present in unexposed humans.

BACKGROUND: A pandemic H5N1 influenza outbreak would be facilitated by an absence of immunity to the avian-derived virus in the human population. Although this condition is likely in regard to hemagglutinin-mediated immunity, the neuraminidase (NA) of H5N1 viruses (avN1) and of endemic human H1N1 viruses (huN1) are classified in the same serotype. We hypothesized that an immune response to huN1 could mediate cross-protection against H5N1 influenza virus infection. METHODS AND FINDINGS: Mice were immunized against the NA of a contemporary human H1N1 strain by DNA vaccination. They were challenged with recombinant A/Puerto Rico/8/34 (PR8) viruses bearing huN1 (PR8-huN1) or avN1 (PR8-avN1) or with H5N1 virus A/Vietnam/1203/04. Additional naïve mice were injected with sera from vaccinated mice prior to H5N1 challenge. Also, serum specimens from humans were analyzed for reactivity with avN1. Immunization elicited a serum IgG response to huN1 and robust protection against the homologous challenge virus. Immunized mice were partially protected from lethal challenge with H5N1 virus or recombinant PR8-avN1. Sera transferred from immunized mice to naïve animals conferred similar protection against H5N1 mortality. Analysis of human sera showed that antibodies able to inhibit the sialidase activity of avN1 exist in some individuals. CONCLUSIONS: These data reveal that humoral immunity elicited by huN1 can partially protect against H5N1 infection in a mammalian host. Our results suggest that a portion of the human population could have some degree of resistance to H5N1 influenza, with the possibility that this could be induced or enhanced through immunization with seasonal influenza vaccines.

Novel microneutralization assay for HCMV using automated data collection and analysis.

In addition to being sensitive and specific, an assay for the assessment of neutralizing antibody activity from clinical trial samples must be amenable to automation for use in high-volume screening. To that effect, we developed a 96-well microplate assay for the measurement of HCMV-neutralizing activity in human sera using the HCMV-permissive human cell line HEL-299 and the laboratory strain of HCMV AD169. The degree to which neutralizing antibodies diminish HCMV infection of cells in the assay is determined by quantifying the nuclei of infected cells based on expression of the 72 kDa IE1 viral protein. Nuclear IE1 is visualized using a highly sensitive immunoperoxidase staining and the stained nuclei are counted using an automated ELISPOT analyzer. The use of Half Area 96-well microplates, with wells in which the surface area of the well bottom is half the area of a standard 96-well microplate plate, improves signal detection compared with standard microplates and economizes on the usage of indicator cells, virus, and reagents. The staining process was also streamlined by using a microplate washer and data analysis was simplified and accelerated by employing a software program that automatically plots neutralization curves and determines NT(50) values using 4-PL curve fitting. The optimized assay is not only fast and convenient, but also specific, sensitive, precise and reproducible and thus has the characteristics necessary for use in measuring HCMV-neutralizing activity in the sera of vaccine trial subjects such as the recipients of Vical's HCMV pDNA vaccine candidates.

Establishment of an HLA-A*0201 human papillomavirus type 16 tumor model to determine the efficacy of vaccination strategies in HLA-A*0201 transgenic mice.

With the increasing generation of new cancer vaccine strategies, there is also an increasing demand for preclinical models that can carefully predict the efficacy of these vaccines in humans. However, the only tumor models available to study vaccines against human papillomavirus (HPV) 16 have been developed in C57BL/6 mice. To test the HLA-restricted capabilities of vaccination strategies, it is important to establish a tumor model in HLA-A*0201 transgenic mice. By transfecting heart lung fibroblasts from HLA-A*0201 mice with HPV16 E6 and E7 oncogenes and H-Ras V12, we have generated a transgenic cell line that is tumorigenic in HLA-A*0201 mice. The dominant H-2D(b) HPV16 E7 epitope was removed from the E7 construct to ensure that all antitumor responses were mediated through the HLA-A*0201-restricted epitopes. We used this tumor model to test the efficacy of two genetic vaccines: a plasmid DNA multi-epitope vaccine encoding human epitopes of HPV16, and a Venezuelan equine encephalitis (VEE) virus-based vector to deliver HPV16 E6 and E7 RNA. We show that both our multi-epitope DNA- and VEE-based vaccines protect 100% of HLA-A*0201 transgenic mice from tumor challenge and elicit a specific T-cell response against multiple HLA-A*0201-restricted HPV16 epitopes. Furthermore, both vaccines significantly decreased tumor burden when tested therapeutically. In conclusion, this is the first tumor model that allows for the assessment of the potential of a vaccine to induce HPV-directed, HLA-A*0201-restricted, antitumor responses in mice. These results pave the way for the clinical evaluation of these vaccines.

A cytomegalovirus DNA vaccine induces antibodies that block viral entry into fibroblasts and epithelial cells.

A vaccine to prevent congenital cytomegalovirus (CMV) infections is a national priority. Investigational vaccines have targeted the viral glycoprotein B (gB) as an inducer of neutralizing antibodies and phosphoprotein 65 (pp65) as an inducer of cytotoxic T cells. Antibodies to gB neutralize CMV entry into all cell types but their potency is low compared to antibodies that block epithelial cell entry through targeting the pentameric complex (gH/gL/UL128/UL130/UL131). Hence, more potent overall neutralizing responses may result from a vaccine that combines gB with pentameric complex-derived antigens. To assess the ability of pentameric complex subunits to generate epithelial entry neutralizing antibodies, DNA vaccines encoding UL128, UL130, and/or UL131 were formulated with Vaxfectin(®), an adjuvant that enhances antibody responses to DNA vaccines. Mice were immunized with individual DNA vaccines or with pair-wise or trivalent combinations. Only the UL130 vaccine induced epithelial entry neutralizing antibodies and no synergy was observed from bi- or trivalent combinations. In rabbits the UL130 vaccine again induced epithelial entry neutralizing antibodies while UL128 or UL131 vaccines did not. To evaluate compatibility of the UL130 vaccine with DNA vaccines encoding gB or pp65, mono-, bi-, or trivalent combinations were evaluated. Fibroblast and epithelial entry neutralizing titers did not differ between rabbits immunized with gB alone vs. gB/UL130, gB/pp65, or gB/UL130/pp65 combinations, indicating a lack of antagonism from coadministration of DNA vaccines. Importantly, gB-induced epithelial entry neutralizing titers were substantially higher than activities induced by UL130, and both fibroblast and epithelial entry neutralizing titers induced by gB alone as well as gB/pp65 or gB/UL130/pp65 combinations were comparable to those observed in sera from humans with naturally-acquired CMV infections. These findings support further development of Vaxfectin(®)-formulated gB-expressing DNA vaccine for prevention of congenital CMV infections.

Questionnaire for biotechs: Vical incorporated.

This feature contains forward-looking statements subject to risks and uncertainties that could cause actual results to differ materially from those projected. Forward-looking statements include statements about the company's focus, collaborative partners, product candidates, and developmental status. Risks and uncertainties include whether any product candidates will be shown to be safe and efficacious in clinical trials, the timing of clinical trials, whether Vical or its collaborative partners will seek or gain approval to market any product candidates, the dependence of the company on its collaborative partners, and additional risks set forth in the company's filings with the Securities and Exchange Commission. These forward-looking statements represent the company's judgment as of the date of this release. The company disclaims, however, any intent or obligation to update these forward-looking statements.

Clinical Development of a Cytomegalovirus DNA Vaccine: From Product Concept to Pivotal Phase 3 Trial.

2013 marks a milestone year for plasmid DNA vaccine development as a first-in-class cytomegalovirus (CMV) DNA vaccine enters pivotal phase 3 testing. This vaccine consists of two plasmids expressing CMV antigens glycoprotein B (gB) and phosphoprotein 65 (pp65) formulated with a CRL1005 poloxamer and benzalkonium chloride (BAK) delivery system designed to enhance plasmid expression. The vaccine's planned initial indication under investigation is for prevention of CMV reactivation in CMV-seropositive (CMV⁺) recipients of an allogeneic hematopoietic stem cell transplant (HCT). A randomized, double-blind placebo-controlled phase 2 proof-of-concept study provided initial evidence of the safety of this product in CMV⁺ HCT recipients who underwent immune ablation conditioning regimens. This study revealed a significant reduction in viral load endpoints and increased frequencies of pp65-specific interferon-γ-producing T cells in vaccine recipients compared to placebo recipients. The results of this endpoint-defining trial provided the basis for defining the primary and secondary endpoints of a global phase 3 trial in HCT recipients. A case study is presented here describing the development history of this vaccine from product concept to initiation of the phase 3 trial.

Preclinical evaluation of Vaxfectin-adjuvanted Vero cell-derived seasonal split and pandemic whole virus influenza vaccines.

Increasing the potency and supply of seasonal and pandemic influenza vaccines remains an important unmet medical need which may be effectively accomplished with adjuvanted egg- or cell culture-derived vaccines. Vaxfectin, a cationic lipid-based adjuvant with a favorable safety profile in phase 1 plasmid DNA vaccines trials, was tested in combination with seasonal split, trivalent and pandemic whole virus, monovalent influenza vaccines produced in Vero cell cultures. Comparison of hemagglutination inhibition (HI) antibody titers in Vaxfectin-adjuvanted to nonadjuvanted vaccinated mice and guinea pigs revealed 3- to 20-fold increases in antibody titers against each of the trivalent influenza virus vaccine strains and 2- to 8-fold increases in antibody titers against the monovalent H5N1 influenza virus vaccine strain. With the vaccine doses tested, comparable antibody responses were induced with formulations that were freshly prepared or refrigerated at conventional 2-8°C storage conditions for up to 6 mo. Comparison of T-cell frequencies measured by interferon-gamma ELISPOT assay between groups revealed increases of between 2- to 10-fold for each of the adjuvanted trivalent strains and up to 22-fold higher with monovalent H5N1 strain. Both trivalent and monovalent vaccines were easy to formulate with Vaxfectin by simple mixing. These preclinical data support further testing of Vaxfectin-adjuvanted Vero cell culture vaccines toward clinical studies designed to assess safety and immunogenicity of these vaccines in humans.

Preclinical evaluation of the immunogenicity and safety of plasmid DNA-based prophylactic vaccines for human cytomegalovirus.

Human cytomegalovirus (CMV) establishes a lifelong persistent infection characterized by periods of latency and sporadic viral replication and is a major infectious cause of birth defects following congenital infection. Currently, no licensed vaccine is available that would prevent CMV infection. In an effort to develop a prophylactic CMV vaccine, the effects of different formulations, immunization routes and delivery devices on the immunogenicity of plasmid DNA (pDNA)-based vaccines were evaluated in rabbits and mice. Compared with PBS- and poloxamer-based formulations, significantly higher antibody responses were obtained with pDNA formulated with Vaxfectin (®) , a cationic lipid-based adjuvant. With low vaccine doses, the intradermal (ID) route resulted in higher antibody responses than obtained when the same dose was administered intramuscularly (IM). Since the IM route allowed injection of larger volumes and higher doses than could be administered at a single ID site, better antibody responses were obtained using the IM route. The needle-free injection system Biojector (®) 2000 and electroporation devices enhanced antibody responses only marginally compared with responses obtained with Vaxfectin (®) -formulated pDNA injected IM with a needle. A single-vial Vaxfectin (®) formulation was developed in a dosage form ready for use after thawing at room temperature. Finally, in a GLP-compliant repeat-dose toxicology study conducted in rabbits, single-vial Vaxfectin (®) -formulated vaccines, containing pDNA and Vaxfectin (®) up to 4.5 mg and 2 mg/injection, respectively, showed a favorable safety profile and were judged as well-tolerated. The results support further development of a Vaxfectin (®) -formulated pDNA vaccine to target congenital CMV infection.

A novel therapeutic cytomegalovirus DNA vaccine in allogeneic haemopoietic stem-cell transplantation: a randomised, double-blind, placebo-controlled, phase 2 trial.

BACKGROUND: Cytomegalovirus reactivation occurs within 6 months in 60-70% of cytomegalovirus-seropositive patients after allogeneic haemopoietic stem-cell transplantation (HSCT), mainly due to immunosuppression associated with the procedure. Pre-emptive antiviral therapy reduces incidence of cytomegalovirus disease but can be toxic. To reduce the potential for disease and subsequent need for such antiviral drugs, we aimed to assess safety and efficacy of a cytomegalovirus therapeutic DNA vaccine compared with placebo. METHODS: In this exploratory double-blind, placebo-controlled, parallel group, phase 2 trial, up to 80 donor-recipient pairs and 80 unpaired recipients undergoing allogeneic HSCT were planned for enrolment at 16 transplant centres in the USA. Eligible recipients were cytomegalovirus-seropositive, 18-65 years old, without high-risk primary disease, T-cell depletion, previous vaccination for cytomegalovirus, or autoimmune diseases. We randomly allocated participants in both parallel groups in a 1:1 ratio to receive a cytomegalovirus therapeutic DNA vaccine (TransVax; Vical, San Diego, CA, USA) or placebo before conditioning and at 1, 3, and 6 months after transplantation. The vaccine contains plasmids encoding cytomegalovirus glycoprotein B and phosphoprotein 65 formulated with poloxamer CRL1005 and benzalkonium chloride. Randomisation was done by sequential allocation based on Pocock and Simon's method, and stratified by site, donor-recipient HLA matching status, and donor's cytomegalovirus serostatus. The primary outcome was the occurrence rate of clinically significant viraemia resulting in initiation of cytomegalovirus-specific antiviral therapy in the per-protocol assessable population. We assessed rates of adverse events in all participants who received at least one dose of vaccine or placebo. This study is registered with ClinicalTrials.gov, number NCT00285259. FINDINGS: We randomly allocated 108 participants (94 HSCT recipients and 14 paired donors) between June 29, 2006, and Dec 11, 2009. Enrolment of the paired arm was halted in February 2008 for logistical reasons. Safety was assessed in all participants; the efficacy population was restricted to 74 unpaired recipients. Groups were balanced for demographic and clinical variables. 19 (48%) of 40 vaccine recipients required cytomegalovirus-specific antiviral therapy, compared with 21 (62%) of 34 controls (p=0·145). However, during follow-up vaccine significantly reduced the occurrence and recurrence of cytomegalovirus viraemia and improved the time-to-event for viraemia episodes compared with placebo. The vaccine was well-tolerated; only one participant discontinued after an allergic reaction. Incidence of common adverse events after HSCT (eg, graft-versus-host disease or secondary infections) did not differ between groups. INTERPRETATION: We show proof of concept for an immunotherapeutic cytomegalovirus vaccine (TransVax) for clinically significant viraemia in the HSCT setting. The reported safety and efficacy outcomes support further development in a phase 3 trial, notwithstanding a lack of significant reduction in the use of cytomegalovirus-specific antiviral therapy compared with placebo in this phase 2 trial. FUNDING: Vical and US National Institute of Allergy and Infectious Diseases.

Phase 1 clinical trials of the safety and immunogenicity of adjuvanted plasmid DNA vaccines encoding influenza A virus H5 hemagglutinin.

BACKGROUND: Development of vaccines against highly pathogenic avian influenza virus H5N1 subtypes posing a pandemic threat remains a priority. Limitations in manufacturing capacity and production time of conventional inactivated vaccines highlight the need for additional approaches. METHODS: We conducted two double-blind, placebo-controlled phase 1 studies involving a total of 103 healthy adults who received two intramuscular injections of Vaxfectin-adjuvanted plasmid DNA vaccine or placebo 21 days apart. Vaccine cohorts received either a monovalent vaccine containing an A/Vietnam/1203/04 H5 hemagglutinin-encoding plasmid or a trivalent vaccine with plasmids encoding H5, NP, and M2 proteins in doses from 0.1 to 1mg of DNA/injection. RESULTS: All doses were well tolerated without vaccine-related serious adverse events or discontinuations. In the monovalent cohorts, hemagglutination inhibition (HI) titers of > or =40 and 4-fold rises from baseline were achieved in 47-67% of subjects and H5-specific T-cell responses in 75-100%. Trivalent cohorts had lower HI response rates (< or = 20%), but 72% of subjects achieved T-cell and/or antibody responses to one or more antigens. CONCLUSIONS: Vaxfectin-adjuvanted monovalent H5 DNA vaccines were well tolerated and induced HI response rates and titers in the reported range of inactivated protein-based H5 vaccines, suggesting that adjuvanted DNA vaccines with rapid vaccine production could be useful for pandemic control.

Vaxfectin, a cationic lipid-based adjuvant for protein-based influenza vaccines.

Mice were immunized either with unadjuvanted seasonal trivalent influenza vaccine (TIV) or TIV formulated with Vaxfectin, a cationic lipid-based adjuvant. Increasing doses of Vaxfectin resulted in increased hemagglutination-inhibition or anti-TIV ELISA antibody titers, with up to a 200-fold increase obtained with 900 microg of Vaxfectin. A >or=10-fold dose-sparing effect was demonstrated with Vaxfectin formulations. Vaxfectin preferentially increased IgG2 titers compared to IgG1 titers, resulting in a balanced IgG isotype distribution. Lower doses of Vaxfectin (30 microg) did not enhance antibody responses, but increased the number of IFN-gamma secreting T-cells by up to 18-fold. The data demonstrate that Vaxfectin enhances Th1 responses with protein-based seasonal influenza vaccine, and suggest that cellular or humoral immune responses may be preferentially induced by modifying the Vaxfectin:antigen ratio in the vaccine formulation.

Plasmid DNA-based vaccines protect mice and ferrets against lethal challenge with A/Vietnam/1203/04 (H5N1) influenza virus.

Plasmid DNA (pDNA) vaccines represent an alternative to conventional inactivated influenza vaccines that are likely to experience supply constraints during a pandemic. Several Vaxfectin-formulated pDNA vaccines were tested in mice and ferrets for efficacy against a lethal challenge with the highly pathogenic A/Vietnam/1203/04 (H5N1) influenza virus strain; the vaccines encoded influenza A virus hemagglutinin (HA), and/or nucleoprotein (NP), and M2 protein. Complete protection from death and disease was achieved in mice and ferrets with 2 doses of a Vaxfectin-formulated vaccine containing H5 HA, NP, and M2 plasmids and in ferrets with only 1 dose. A Vaxfectin-formulated vaccine containing NP and M2 pDNA provided significant protection against death in mice and provided some benefit in ferrets (i.e., 17% survival, delayed time to illness and death, and significant reduction in viral load compared with that in negative control animals). These experiments support the clinical testing of pDNA vaccine candidates that may ultimately increase global vaccine supply options during pandemics.

Safety and immunogenicity of a bivalent cytomegalovirus DNA vaccine in healthy adult subjects.

BACKGROUND: VCL-CB01, a candidate cytomegalovirus (CMV) DNA vaccine that contains plasmids encoding CMV phosphoprotein 65 (pp65) and glycoprotein B (gB) to induce cellular and humoral immune responses and that is formulated with poloxamer CRL1005 and benzalkonium chloride to enhance immune responses, was evaluated in a phase 1 clinical trial. METHODS: VCL-CB01 was evaluated in 44 healthy adult subjects (22 CMV seronegative and 22 CMV seropositive) 18-43 years old. Thirty-two subjects received 1- or 5-mg doses of vaccine on a 0-, 2-, and 8-week schedule, and 12 subjects received 5-mg doses of vaccine on a 0-, 3-, 7-, and 28-day schedule. RESULTS: Overall, the vaccine was well tolerated, with no serious adverse events. Local reactions included mild to moderate injection site pain and tenderness, induration, and erythema. Systemic reactions included mild to moderate malaise and myalgia. All reactions resolved without sequelae. Through week 16 of the study, immunogenicity, as measured by enzyme-linked immunosorbant assay and/or ex vivo interferon (IFN)-gamma enzyme-linked immunospot assay, was documented in 45.5% of CMV-seronegative subjects and in 25.0% of CMV-seropositive subjects who received the full vaccine series, and 68.1% of CMV-seronegative subjects had memory IFN-gamma T cell responses at week 32. CONCLUSION: The safety and immunogenicity data from this trial support further evaluation of VCL-CB01.