Novel Vaccine against Pathological Pyroglutamate-Modified Amyloid Beta for Prevention of Alzheimer's Disease.

Post-translationally modified N-terminally truncated amyloid beta peptide with a cyclized form of glutamate at position 3 (pE3Aß) is a highly pathogenic molecule with increased neurotoxicity and propensity for aggregation. In the brains of Alzheimer's Disease (AD) cases, pE3Aß represents a major constituent of the amyloid plaque. The data show that pE3Aß formation is increased at early pre-symptomatic disease stages, while tau phosphorylation and aggregation mostly occur at later stages of the disease. This suggests that pE3Aß accumulation may be an early event in the disease pathogenesis and can be prophylactically targeted to prevent the onset of AD. The vaccine (AV-1986R/A) was generated by chemically conjugating the pE3Aß3-11 fragment to our universal immunogenic vaccine platform MultiTEP, then formulated in AdvaxCpG adjuvant. AV-1986R/A showed high immunogenicity and selectivity, with endpoint titers in the range of 105-106 against pE3Aß and 103-104 against the full-sized peptide in the 5XFAD AD mouse model. The vaccination showed efficient clearance of the pathology, including non-pyroglutamate-modified plaques, from the mice brains. AV-1986R/A is a novel promising candidate for the immunoprevention of AD. It is the first late preclinical candidate which selectively targets a pathology-specific form of amyloid with minimal immunoreactivity against the full-size peptide. Successful translation into clinic may offer a new avenue for the prevention of AD via vaccination of cognitively unimpaired individuals at risk of disease.

Immunogenicity of MultiTEP-Platform-Based Recombinant Protein Vaccine, PV-1950R, Targeting Three B-Cell Antigenic Determinants of Pathological α-Synuclein.

Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are characterized by the aberrant accumulation of intracytoplasmic misfolded and aggregated α-synuclein (α-Syn), resulting in neurodegeneration associated with inflammation. The propagation of α-Syn aggregates from cell to cell is implicated in the spreading of pathological α-Syn in the brain and disease progression. We and others demonstrated that antibodies generated after active and passive vaccinations could inhibit the propagation of pathological α-Syn in the extracellular space and prevent/inhibit disease/s in the relevant animal models. We recently tested the immunogenicity and efficacy of four DNA vaccines on the basis of the universal MultiTEP platform technology in the DLB/PD mouse model. The antibodies generated by these vaccines efficiently reduced/inhibited the accumulation of pathological α-Syn in the different brain regions and improved the motor deficit of immunized female mice. The most immunogenic and preclinically effective vaccine, PV-1950D, targeting three B-cell epitopes of pathological α-Syn simultaneously, has been selected for future IND-enabling studies. However, to ensure therapeutically potent concentrations of α-Syn antibodies in the periphery of the vaccinated elderly, we developed a recombinant protein-based MultiTEP vaccine, PV-1950R/A, and tested its immunogenicity in young and aged D-line mice. Antibody responses induced by immunizations with the PV-1950R/A vaccine and its homologous DNA counterpart, PV-1950D, in a mouse model of PD/DLB have been compared.

Characterization and preclinical evaluation of the cGMP grade DNA based vaccine, AV-1959D to enter the first-in-human clinical trials.

The DNA vaccine, AV-1959D, targeting N-terminal epitope of Aß peptide, has been proven immunogenic in mice, rabbits, and non-human primates, while its therapeutic efficacy has been shown in mouse models of Alzheimer's disease (AD). Here we report for the first time on IND-enabling biodistribution and safety/toxicology studies of cGMP-grade AV-1959D vaccine in the Tg2576 mouse model of AD. We also tested acute neuropathology safety profiles of AV-1959D in another AD disease model, Tg-SwDI mice with established vascular and parenchymal Aß pathology in a pre-clinical translational study. Biodistribution studies two days after the injection demonstrated high copy numbers of AV-1959D plasmid after single immunization of Tg2576 mice at the injection sites but not in the tissues of distant organs. Plasmids persisted at the injection sites of some mice 60 days after vaccination. In Tg2576 mice with established amyloid pathology, we did not observe short- or long-term toxicities after multiple immunizations with three doses of AV-1959D. Assessment of the repeated dose acute safety of AV-1959D in cerebral amyloid angiopathy (CAA) prone Tg-SwDI mice did not reveal any immunotherapy-induced vasogenic edema detected by magnetic resonance imaging (MRI) or increased microhemorrhages. Multiple immunizations of Tg-SwDI mice with AV-1959D did not induce T and B cell infiltration, glial activation, vascular deposition of Aß, or neuronal degeneration (necrosis and apoptosis) greater than that in the control group determined by immunohistochemistry of brain tissues. Taken together, the safety data from two different mouse models of AD substantiate a favorable safety profile of the cGMP grade AV-1959D vaccine supporting its progression to first-in-human clinical trials.

Comparison of Efficacy of Preventive and Therapeutic Vaccines Targeting the N Terminus of ß-Amyloid in an Animal Model of Alzheimer's Disease.

Previously, we reported that Alzheimer's disease (AD) epitope vaccines (EVs) composed of N-terminal ß-amyloid (Aß42) B cell epitope fused with universal foreign T helper (Th) epitope(s) were immunogenic, potent, and safe in different amyloid precursor protein (APP) transgenic mice with early AD-like pathology. However, developing an effective therapeutic vaccine is much more challenging, especially when a self-antigen such as Aß42 is a target. Here, we directly compare the efficacy of anti-Aß42 antibodies in Tg2576 mice with low or high levels of AD-like pathology at the start of immunizations: 6-6.5 months for preventive vaccinations and 16-19 months for therapeutic vaccinations. EV in a preventive setting induced high levels of anti-Aß antibodies, significantly reducing pathologic forms of Aß in the brains of Tg2576 mice. When used therapeutically for immunesenescent Tg2576 mice, EV induced low levels of antibodies not sufficient for clearing of AD-like pathology. Separately, we demonstrated that EV was also not effective in 11-11.5-month-old Tg2576 mice with moderate AD-like pathology. However, we augmented the titers of anti-Aß antibodies in transgenic (Tg) mice of the same age possessing the pre-existing memory Th cells and detected a significant decrease in diffuse and core plaques in cortical regions compared to control animals along with improved novel object recognition performance.

Immunogenicity, efficacy, safety, and mechanism of action of epitope vaccine (Lu AF20513) for Alzheimer's disease: prelude to a clinical trial.

The Alzheimer's disease (AD) process is understood to involve the accumulation of amyloid plaques and tau tangles in the brain. However, attempts at targeting the main culprits, neurotoxic Aß peptides, have thus far proven unsuccessful for improving cognitive function. Recent clinical trials with passively administrated anti-Aß antibodies failed to slow cognitive decline in mild to moderate AD patients, but suggest that an immunotherapeutic approach could be effective in patients with mild AD. Using an AD mouse model (Tg2576), we tested the immunogenicity (cellular and humoral immune responses) and efficacy (AD-like pathology) of clinical grade Lu AF20513 vaccine. We found that Lu AF20513 induces robust "non-self" T-cell responses and the production of anti-Aß antibodies that reduce AD-like pathology in the brains of Tg2576 mice without inducing microglial activation and enhancing astrocytosis or cerebral amyloid angiopathy. A single immunization with Lu AF20513 induced strong humoral immunity in mice with preexisting memory T-helper cells. In addition, Lu AF20513 induced strong humoral responses in guinea pigs and monkeys. These data support the translation of Lu AF20513 to the clinical setting with the aims of: (1) inducing therapeutically potent anti-Aß antibody responses in patients with mild AD, particularly if they have memory T-helper cells generated after immunizations with conventional tetanus toxoid vaccine, and (2) preventing pathological autoreactive T-cell responses.

Targeting TLR-4 with a novel pharmaceutical grade plant derived agonist, Immunomax®, as a therapeutic strategy for metastatic breast cancer.

BACKGROUND: Previously we demonstrated that the resection of primary 4T1 tumors only slightly prolongs mouse survival, but importantly, creates a "window of opportunity" with attenuated suppressor cell and increased activated T cell populations. This suggests that additional activation of the immune system by immunostimulatory agents during this period may enhance anti-tumor immunity and potentially eradicate micro-metastatic disease in this stringent model. We hypothesized that the immunostimulator Immunomax®, which is comprised of a plant-derived polysaccharide, is non-toxic in humans and stimulates immune defense during the infectious diseases treatment, may have also anti-tumor activity and be beneficial in the adjuvant setting when endogenous anti-tumor responses are present and during the "window of opportunity" in post-resection metastatic breast cancer model. Here we provide the initial report that Immunomax® demonstrates the capacity to eliminate micro-metastatic disease in the post-resection, 4T1 mouse model of breast cancer. METHODS: The efficacy of Immunomax® was evaluated by analyzing survival rate and the number of spontaneous clonogenic tumor cells in the lung homogenates of mice. The frequencies of activated NK, CD4(+) and CD8(+) cells as well as myeloid-derived suppressor cells and Treg cells were evaluated using flow cytometry. Highly purified mouse and human dendritic and NK cells were sorted and the effect of Immunomax® on activation status of these cells was assessed by flow cytometry. The property of Immunomax® as TLR-4 agonist was determined by NF-κB/SEAP reporter gene assay, WB, RT-PCR. RESULTS: Immunomax® injections significantly prolonged overall survival and cured 31% of mice. This immunostimulator activates DCs via the TLR-4, which in turn stimulates tumoricidal NK cells and in vitro, completely inhibits growth of 4T1 cells. Incubation of PBMC from healthy donors with Immunomax® activates NK cells via activation of plasmacytoid DC leading significantly higher efficacy in killing of human NK-target cells K562 compared with non-treated cells. CONCLUSION: This is the first demonstration that Immunomax® is a TLR-4 agonist and the first report of a documented role for this pharmaceutical grade immunostimulator in augmenting anti-tumor activity, suggesting that incorporation of Immunomax® into developing breast cancer therapeutic strategies may be beneficial and with less potential toxicity than checkpoint inhibitors.

The MultiTEP platform-based Alzheimer's disease epitope vaccine activates a broad repertoire of T helper cells in nonhuman primates.

BACKGROUND: As a prelude to clinical trials we have characterized B- and T-cell immune responses in macaques to AD vaccine candidates: AV-1955 and its slightly modified version, AV-1959 (with 3 additional promiscuous Th epitopes). METHODS: T- and B-cell epitope mapping was performed using the ELISPOT assay and competition ELISA, respectively. RESULTS: AV-1955 and AV-1959 did not stimulate potentially harmful autoreactive T cells, but instead activated a broad but individualized repertoire of Th cells specific to the MultiTEP platform in macaques. Although both vaccines induced robust anti-Aß antibody responses without producing antibodies specific to Th epitopes of MultiTEP platforms, analyses of cellular immune responses in macaques demonstrated that the addition of Th epitopes in the case of AV-1959 created a more potent, superior vaccine. CONCLUSION: AV-1959 is a promising vaccine candidate capable of producing therapeutically potent anti-amyloid antibody in a broader population of vaccinated subjects with high MHC class II gene polymorphisms.

Epitope-based DNA vaccine for Alzheimer's disease: translational study in macaques.

BACKGROUND: Clinical trials with passive and active Alzheimer's disease (AD) vaccines suggest that early interventions are needed for improvement of cognitive and/or functional performance in patients, providing impetus for the development of safe and immunologically potent active vaccines targeting amyloid ß (Aß). The AN-1792 trial has indicated that Aß-specific T cells may be unsafe for humans; therefore, other vaccines based on small Aß epitopes are undergoing preclinical and clinical testing. METHODS: Humoral and cellular immune responses elicited in response to a novel DNA epitope-based vaccine (AV-1955) delivered to rhesus macaques using the TriGrid electroporation device were evaluated. Functional activities of anti-Aß antibodies generated in response to vaccination were assessed in vitro. RESULTS: AV-1955 generates long-term, potent anti-Aß antibodies and cellular immune responses specific to foreign T-helper epitopes but not to self-Aß. CONCLUSIONS: This translational study demonstrates that a DNA-based epitope vaccine for AD could be appropriate for human clinical testing.

MultiTEP-Based Vaccines Targeting SARS-CoV-2 Spike Protein IgG Epitopes Elicit Robust Binding Antibody Titers with Limited Virus-Neutralizing Activity.

Within the last two decades, SARS-CoV-2 was the third zoonotic severe acute respiratory betacoronavirus (sarbecovirus) to infect humans, following SARS and MERS. The disruptions caused by the pandemic underscore the need for a universal vaccine against respiratory betacoronaviruses. Our group previously developed the universal platform for vaccine development, MultiTEP, which has been utilized in this study to generate a range of SARS-CoV-2 epitope vaccine candidates. We prepared and characterized 18 vaccines incorporating small peptide fragments from SARS-CoV-2 Spike protein fused with the MultiTEP sequence using overlapping PCR. Wild-type mice were immunized intramuscularly with the immunogen formulated in AdvaxCpG adjuvant. Serum antibodies were detected by ELISA, surrogate neutralization, and pseudovirus neutralization assays. Finally, the most promising vaccine candidate was administered to three non-human primates. All vaccines generated high titers of spike-binding IgG antibodies. However, only three vaccines generated antibodies that blocked RBD binding to the ACE2 receptor in a surrogate virus neutralization assay. However, none of the vaccines induced antibodies able to neutralize pseudotype viruses, including after the administration of the lead vaccine to NHPs. MultiTEP-based COVID-19 vaccines elicited robust, IgG-binding responses against the Spike protein in mice and non-human primates, but these antibodies were not neutralizing, underscoring the need to refine this approach further.

mRNA Vaccine for Alzheimer's Disease: Pilot Study.

The escalating global healthcare challenge posed by Alzheimer's Disease (AD) and compounded by the lack of effective treatments emphasizes the urgent need for innovative approaches to combat this devastating disease. Currently, passive and active immunotherapies remain the most promising strategy for AD. FDA-approved lecanemab significantly reduces Aß aggregates from the brains of early AD patients administered biweekly with this humanized monoclonal antibody. Although the clinical benefits noted in these trials have been modest, researchers have emphasized the importance of preventive immunotherapy. Importantly, data from immunotherapy studies have shown that antibody concentrations in the periphery of vaccinated people should be sufficient for targeting Aß in the CNS. To generate relatively high concentrations of antibodies in vaccinated people at risk of AD, we generated a universal vaccine platform, MultiTEP, and, based on it, developed a DNA vaccine, AV-1959D, targeting pathological Aß, completed IND enabling studies, and initiated a Phase I clinical trial with early AD volunteers. Our current pilot study combined our advanced MultiTEP technology with a novel mRNA approach to develop an mRNA vaccine encapsulated in lipid-based nanoparticles (LNPs), AV-1959LR. Here, we report our initial findings on the immunogenicity of 1959LR in mice and non-human primates, comparing it with the immunogenicity of its DNA counterpart, AV-1959D.

Immunogenicity of MultiTEP platform technology-based Tau vaccine in non-human primates.

Pathological forms of Tau protein are directly associated with neurodegeneration and correlate with Alzheimer's Disease (AD) symptoms, progression, and severity. Previously, using various mouse models of Tauopathies and AD, we have demonstrated the immunogenicity and efficacy of the MultiTEP-based adjuvanted vaccine targeting the phosphatase activating domain (PAD) of Tau, AV-1980R/A. Here, we analyzed its immunogenicity in non-human primates (NHP), the closest phylogenic relatives to humans with a similar immune system, to initiate the transition of this vaccine into clinical trials. We have demonstrated that AV-1980R/A is highly immunogenic in these NHPs, activating a broad but unique to each monkey repertoire of MultiTEP-specific T helper (Th) cells that, in turn, activate B cells specific to PAD. The resulting anti-PAD IgG antibodies recognize pathological Tau tangles and Tau-positive neuritis in AD case brain sections with no staining in control non-AD cases. These published data and efficacy results support the AV-1980R/A vaccine progression to first-in-human clinical trials.

Efficacy and immunogenicity of MultiTEP-based DNA vaccines targeting human α-synuclein: prelude for IND enabling studies.

Accumulation of misfolded proteins such as amyloid-ß (Aß), tau, and α-synuclein (α-Syn) in the brain leads to synaptic dysfunction, neuronal damage, and the onset of relevant neurodegenerative disorder/s. Dementia with Lewy bodies (DLB) and Parkinson's disease (PD) are characterized by the aberrant accumulation of α-Syn intracytoplasmic Lewy body inclusions and dystrophic Lewy neurites resulting in neurodegeneration associated with inflammation. Cell to cell propagation of α-Syn aggregates is implicated in the progression of PD/DLB, and high concentrations of anti-α-Syn antibodies could inhibit/reduce the spreading of this pathological molecule in the brain. To ensure sufficient therapeutic concentrations of anti-α-Syn antibodies in the periphery and CNS, we developed four α-Syn DNA vaccines based on the universal MultiTEP platform technology designed especially for the elderly with immunosenescence. Here, we are reporting on the efficacy and immunogenicity of these vaccines targeting three B-cell epitopes of hα-Syn aa85-99 (PV-1947D), aa109-126 (PV-1948D), aa126-140 (PV-1949D) separately or simultaneously (PV-1950D) in a mouse model of synucleinopathies mimicking PD/DLB. All vaccines induced high titers of antibodies specific to hα-Syn that significantly reduced PD/DLB-like pathology in hα-Syn D line mice. The most significant reduction of the total and protein kinase resistant hα-Syn, as well as neurodegeneration, were observed in various brain regions of mice vaccinated with PV-1949D and PV-1950D in a sex-dependent manner. Based on these preclinical data, we selected the PV-1950D vaccine for future IND enabling preclinical studies and clinical development.

A MultiTEP platform-based epitope vaccine targeting the phosphatase activating domain (PAD) of tau: therapeutic efficacy in PS19 mice.

Pathological tau correlates well with cognitive impairments in Alzheimer's disease (AD) patients and therefore represents a promising target for immunotherapy. Targeting an appropriate B cell epitope in pathological tau could in theory produce an effective reduction of pathology without disrupting the function of normal native tau. Recent data demonstrate that the N-terminal region of tau (aa 2-18), termed the "phosphatase activation domain (PAD)", is hidden within native Tau in a 'paperclip'-like conformation. Conversely, PAD is exposed in pathological tau and plays an essential role in the inhibition of fast axonal transport and tau polymerization. Thus, we hypothesized that anti-tau2-18 antibodies may safely and specifically reduce pathological tau and prevent further aggregation, which in turn would neutralize tau toxicity. Therefore, we evaluated the immunogenicity and therapeutic efficacy of our MultiTEP platform-based vaccine targeting tau2-18 formulated with AdvaxCpG adjuvant (AV-1980R/A) in PS19 tau transgenic mice. The AV-1980R/A induced extremely high antibody responses and the resulting sera recognized neurofibrillary tangles and plaque-associated dystrophic neurites in AD brain sections. In addition, under non-denaturing conditions AV-1980R/A sera preferentially recognized AD-associated tau. Importantly, vaccination also prevented age-related motor and cognitive deficits in PS19 mice and significantly reduced insoluble total and phosphorylated tau species. Taken together, these findings suggest that predominantly targeting misfolded tau with AV-1980R/A could represent an effective strategy for AD immunotherapy.

Testing a MultiTEP-based combination vaccine to reduce Aß and tau pathology in Tau22/5xFAD bigenic mice.

BACKGROUND: Alzheimer disease (AD) is characterized by the accumulation of beta-amyloid (Aß) plaques and neurofibrillary tangles composed of hyperphosphorylated tau, which together lead to neurodegeneration and cognitive decline. Current therapeutic approaches have primarily aimed to reduce pathological aggregates of either Aß or tau, yet phase 3 clinical trials of these approaches have thus far failed to delay disease progression in humans. Strong preclinical evidence indicates that these two abnormally aggregated proteins interact synergistically to drive downstream neurodegeneration. Therefore, combinatorial therapies that concurrently target both Aß and tau might be needed for effective disease modification. METHODS: A combinatorial vaccination approach was designed to concurrently target both Aß and tau pathologies. Tau22/5xFAD (T5x) bigenic mice that develop both pathological Aß and tau aggregates were injected intramuscularly with a mixture of two MultiTEP epitope vaccines: AV-1959R and AV-1980R, targeting Aß and tau, respectively, and formulated in AdvaxCpG, a potent polysaccharide adjuvant. Antibody responses of vaccinated animals were measured by ELISA, and neuropathological changes were determined in brain homogenates of vaccinated and control mice using ELISA and Meso Scale Discovery (MSD) multiplex assays. RESULTS: T5x mice immunized with a mixture of Aß- and tau-targeting vaccines generated high Aß- and tau-specific antibody titers that recognized senile plaques and neurofibrillary tangles/neuropil threads in human AD brain sections. Production of these antibodies in turn led to significant reductions in the levels of soluble and insoluble total tau, and hyperphosphorylated tau as well as insoluble Aß42, within the brains of bigenic T5x mice. CONCLUSIONS: AV-1959R and AV-1980R formulated with AdvaxCpG adjuvant are immunogenic and therapeutically potent vaccines that in combination can effectively reduce both of the hallmark pathologies of AD in bigenic mice. Taken together, these findings warrant further development of this vaccine technology for ultimate testing in human AD.

Alzheimer's disease Advax(CpG)- adjuvanted MultiTEP-based dual and single vaccines induce high-titer antibodies against various forms of tau and Aß pathological molecules.

Although ß-amyloid (Aß) may be the primary driver of Alzheimer's disease (AD) pathology, accumulation of pathological tau correlates with dementia in AD patients. Thus, the prevention/inhibition of AD may require vaccine/s targeting Aß and tau simultaneously or sequentially. Since high antibody titers are required for AD vaccine efficacy, we have decided to generate vaccines, targeting Aß (AV-1959R), Tau (AV-1980R) or Aß/tau (AV-1953R) B cell epitopes, based on immunogenic MultiTEP platform and evaluate the immunogenicity of these vaccines formulated with Advax(CpG), delta inulin, Alhydrogel(®), Montanide-ISA51, Montanide-ISA720, MPLA-SM pharmaceutical grade adjuvants. Formulation of AV-1959R in Advax(CpG) induced the highest cellular and humoral immune responses in mice. The dual-epitope vaccine, AV-1953R, or the combination of AV-1959R and AV-1980R vaccines formulated with Advax(CpG) induced robust antibody responses against various forms of both, Aß and tau pathological molecules. While anti-Aß antibody titers after AV-1953R immunization were similar to that in mice vaccinated with AV-1959R or AV-1959R/AV-1980R combination, anti-tau titers were significantly lower after AV-1953R injection when compared to the AV-1980R or AV-1959R/AV-1980R. In silico 3D-modeling provided insight into the differences in immunogenicity of these vaccine constructs. In sum, AV-1959R and AV-1980R formulated with Advax(CpG) adjuvant were identified as promising immunogenic vaccines for ongoing pre-clinical assessment and future human clinical trials.

A dual vaccine against influenza & Alzheimer's disease failed to enhance anti-ß-amyloid antibody responses in mice with pre-existing virus specific memory.

Novel dual vaccine, WSN-Aß(1-10), based on the recombinant influenza virus, expressing immunodominant B-cell epitope of ß-amyloid, simultaneously induced therapeutically potent anti-Aß and anti-influenza antibodies. In this study we showed that boosting of WSN-WT primed mice with WSN-Aß(1-10) enhances anti-viral, but fails to induce anti-Aß antibody responses. This inhibition is associated with expression of Aß(1-10) within the context of an inactivated influenza virus vaccine. These results demonstrate that the use of an inactivated influenza virus as a carrier for AD vaccine may not be applicable due to the possible inhibition of anti-Aß antibody response in individuals previously vaccinated or infected with influenza.

Primary 4T1 tumor resection provides critical "window of opportunity" for immunotherapy.

It is believed that primary tumor resection modulates host-tumor immune interaction, but this has not been characterized in a stringent breast cancer tumor model. This report, using the 4T1 murine mammary tumor model, characterizes for the first time the dynamic longitudinal changes in immunosuppressive and effector components of the immune system after resection of an established orthotopic primary tumor with a defined natural history of developing lung metastases. More specifically, we analyzed changes of absolute numbers and frequencies of MDSC, regulatory T cells (Treg), as well as activated CD4 and CD8 positive T cells in spleens and, in some studies, lungs of 4T1 tumor-bearing mice and mice after primary tumor resection. Importantly, using mathematical analyses we established that primary resection of an orthotopic tumor had created a "window of opportunity" with decreased tumor-associated immune suppression that existed for approximately 10 days. Although tumor resection did slightly prolong survival, it did not affect the ultimate development of metastatic disease since animals with resected tumors or intact primary tumors eventually died by day 47 and 43, respectively. This window of opportunity likely occurs in humans providing a rationale and parameters for integration and testing of immunotherapeutic strategies in this critical "window of opportunity" to combat the development of metastatic disease.

BTX AgilePulse(TM) system is an effective electroporation device for intramuscular and intradermal delivery of DNA vaccine.

DNA vaccines promote immune system activation in small animals and exhibit certain advantages when compared to conventional recombinant protein vaccines. However in clinical trials DNA vaccines are less effective in inducing potent immune responses due to the low delivery efficiency and expression of antigens. Currently, various delivery devices such as gene-guns, bioinjectors and electroporation systems are being used in order to increase the potency of DNA vaccines. However, the optimal delivery parameters are required and must be carefully set to obtain the highest levels of gene expression and strong immune responses in humans. The focus of this study was to optimize electroporation settings (voltage, pulse length, pulse intervals, and number of pulses), as well as the route of administration (intradermal vs. intramuscular) and dosage of the DNA epitope vaccine, AV-1959D, delivered by the BTX AgilePulse(TM) system. As a result, we have chosen the optimal settings for electroporation delivery using different routes of immunization with this vaccine, generating (i) robust antibody production to the B cell epitope (a small peptide, derived from ß-amyloid), and (ii) strong cellular immune responses to Th epitopes (a small synthetic peptide and eleven peptides from various pathogens) incorporated into DNA vaccine platform.

Immunostimulant patches containing Escherichia coli LT enhance immune responses to DNA- and recombinant protein-based Alzheimer's disease vaccines.

Immunotherapeutic approaches to treating Alzheimer's disease (AD) using vaccination strategies must overcome the obstacle of achieving adequate responses to vaccination in the elderly. Here we demonstrate for the first time that application of the Escherichia coli heat-labile enterotoxin adjuvant-laden immunostimulatory patches (LT-IS) dramatically enhances the onset and magnitude of immune responses to DNA- and protein-based vaccines for Alzheimer's disease following intradermal immunization via gene gun and conventional needles, respectively. Our studies suggest that the immune activation mediated by LT-IS offers improved potency for generating AD-specific vaccination responses that should be investigated as an adjuvant in the clinical arena.