Single dose of DPX-rPA, an enhanced-delivery anthrax vaccine formulation, protects against a lethal Bacillus anthracis spore inhalation challenge.

Anthrax is a serious biological threat caused by pulmonary exposure to aerosolized spores of Bacillus anthracis. Biothrax® (anthrax vaccine adsorbed (AVA)) is the only Food and Drug Administration-licensed vaccine and requires five administrations over 12 months with annual boosting to maintain pre-exposure prophylaxis. Here we report the evaluation of a single intramuscular injection of recombinant B. anthracis-protective antigen (rPA) formulated in the DPX delivery platform. Immune responses were compared to an alum-based formulation in mice and rabbits. Serological analysis of anti-rPA immunoglobulin G and toxin neutralization activity demonstrated higher responses induced by DPX-rPA when compared to rPA in alum. DPX-rPA was compared to AVA in rabbits and non-human primates (NHPs). In both species, DPX-rPA generated responses after a single immunization, whereas AVA required two immunizations. In rabbits, single injection of DPX-rPA or two injections of AVA conferred 100% protection from anthrax challenge. In NHPs, single-dose DPX-rPA was 100% protective against challenge, whereas one animal in the two-dose AVA group and all saline administered animals succumbed to infection. DPX-rPA was minimally reactogenic in all species tested. These data indicate that DPX-rPA may offer improvement over AVA by reducing the doses needed for protective immune responses and is a promising candidate as a new-generation anthrax vaccine.

Unique depot formed by an oil based vaccine facilitates active antigen uptake and provides effective tumour control.

BACKGROUND: Oil emulsions are commonly used as vaccine delivery platforms to facilitate slow release of antigen by forming a depot at the injection site. Antigen is trapped in the aqueous phase and as the emulsion degrades in vivo the antigen is passively released. DepoVax™ is a unique oil based delivery system that directly suspends the vaccine components in the oil diluent that forces immune cells to actively take up components from the formulation in the absence of passive release. The aim of this study was to use magnetic resonance imaging (MRI) with additional biological markers to evaluate and understand differences in clearance between several different delivery systems used in peptide-based cancer vaccines. METHODS: C57BL/6 mice were implanted with a cervical cancer model and vaccinated 5 days post-implant with either DepoVax (DPX), a water-in-oil emulsion (w/o), a squalene oil-in-water emulsion (squal o/w) or a saponin/liposome emulsion (sap/lip) containing iron oxide-labeled targeted antigen. MRI was then used to monitor antigen clearance, the site of injection, tumour and inguinal lymph node volumes and other gross anatomical changes. HLA-A2 transgenic mice were also vaccinated to evaluate immune responses of human directed peptides. RESULTS: We demonstrated differences in antigen clearance between DPX and w/o both in regard to how quickly the antigen was cleared and the pattern in which it was cleared. We also found differences in lymph node responses between DPX and both squal o/w and sap/lip. CONCLUSIONS: These studies underline the unique mechanism of action of this clinical stage vaccine delivery system.

Survivin-targeted immunotherapy drives robust polyfunctional T cell generation and differentiation in advanced ovarian cancer patients.

DepoVax™ is an innovative and strongly immunogenic vaccine platform. Survivin is highly expressed in many tumor types and has reported prognostic value. To generate tumor-specific immune response, a novel cancer vaccine was formulated in DepoVax platform (DPX-Survivac) using survivin HLA class I peptides. Safety and immune potency of DPX-Survivac was tested in combination with immune-modulator metronomic cyclophosphamide in ovarian cancer patients. All the patients receiving the therapy produced antigen-specific immune responses; higher dose vaccine and cyclophosphamide treatment generating significantly higher magnitude responses. Strong T cell responses were associated with differentiation of naïve T cells into central/effector memory (CM/EM) and late differentiated (LD) polyfunctional antigen-specific CD4+ and CD8+ T cells. This approach enabled rapid de novo activation/expansion of vaccine antigen-specific CD8+ T cells and provided a strong rationale for further testing to determine clinical benefits associated with this immune activation. These data represent vaccine-induced T cell activation in a clinical setting to a self-tumor antigen previously described only in animal models.

First-in-man application of a novel therapeutic cancer vaccine formulation with the capacity to induce multi-functional T cell responses in ovarian, breast and prostate cancer patients.

BACKGROUND: DepoVax is a novel non-emulsion depot-forming vaccine platform with the capacity to significantly enhance the immunogenicity of peptide cancer antigens. Naturally processed HLA-A2 restricted peptides presented by breast, ovarian and prostate cancer cells were used as antigens to create a therapeutic cancer vaccine, DPX-0907. METHODS: A phase I clinical study was designed to examine the safety and immune activating potential of DPX-0907 in advanced stage breast, ovarian and prostate cancer patients. A total of 23 late stage cancer patients were recruited and were divided into two dose/volume cohorts in a three immunization protocol. RESULTS: DPX-0907 was shown to be safe with injection site reactions being the most commonly reported adverse event. All breast cancer patients (3/3), most of ovarian (5/6) and one third of prostate (3/9) cancer patients exhibited detectable immune responses, resulting in a 61% immunological response rate. Immune responses were generally observed in patients with better disease control after their last prior treatment. Antigen-specific responses were detected in 73% of immune responders (44% of evaluable patients) after the first vaccination. In 83% of immune responders (50% of evaluable patients), peptide-specific T cell responses were detected at ≥2 time points post vaccination with 64% of the responders (39% of evaluable patients) showing evidence of immune persistence. Immune monitoring also demonstrated the generation of antigen-specific T cell memory with the ability to secrete multiple Type 1 cytokines. CONCLUSIONS: The novel DepoVax formulation promotes multifunctional effector memory responses to peptide-based tumor associated antigens. The data supports the capacity of DPX-0907 to elicit Type-1 biased immune responses, warranting further clinical development of the vaccine. This study underscores the importance of applying vaccines in clinical settings in which patients are more likely to be immune competent. TRIAL REGISTRATION: ClinicalTrials.gov NCT01095848.

Development and validation of an HPLC/UV assay for separation and quantification of peptide antigens from a liposomal vaccine delivery platform.

The development and validation of an HPLC method for the quantification of eight peptide antigens from the therapeutic cancer vaccine DPX-0907 is described. The antigens were formulated in DepoVax™, a patented liposomal vaccine delivery platform used in a phase 1 study for breast, ovarian, and prostate cancers. A gradient reversed-phase method with UV detection was optimized for separating and quantifying the peptide mixture. Several extraction methods investigated to extract the peptides from the lipids led to poor recovery of one or more of the peptides. A simple, reproducible, and high-recovery extraction procedure for the simultaneous quantification of hydrophilic and hydrophobic peptides was discovered using a liquid-liquid extraction with water-saturated n-butanol and sodium bicarbonate (0.1 M). The method was found to be specific, linear, accurate, precise, and reliable within the range of 50-150% of the nominal concentration for DPX-0907. The validated method was successfully applied to the assay of peptide content in pre-clinical and clinical batches of DPX-0907.

Efficacy of a single dose hepatitis B depot vaccine.

A single dose formulation of a novel hepatitis B vaccine, consisting of an adjuvant emulsion of liposomes in oil was produced at a manufacturing scale and delivered to rabbits. This single dose vaccine generated a significantly higher antibody response than two doses of an alum-adjuvanted control vaccine in the short term, and was as effective as three doses of the control vaccine in the long term.

A novel breast/ovarian cancer peptide vaccine platform that promotes specific type-1 but not Treg/Tr1-type responses.

In light of lack of efficacy associated with current cancer vaccines, we aimed to develop a novel vaccine platform called DepoVax as a therapeutic vaccine for breast/ovarian cancer. This study was designed to examine the efficacy of this novel platform over conventional emulsion vaccine using human class I MHC transgenic mice. We have developed a water-free depot vaccine formulation (DPX-0907) with high immune activating potential. Naturally processed peptides bound to HLA-A2 molecules isolated from independent breast and ovarian tumor cell lines, but not normal cells, were isolated and used as antigens in DPX-0907 along with a proprietary adjuvant and a T helper peptide epitope. Efficacy of vaccine was tested in immunized HLA-A*0201/H2Dd transgenic mice by measuring the frequency of IFN-gamma secreting cells in the draining lymph nodes, and regulatory T-cell frequencies in the spleen. Compared with a water-in-oil emulsion vaccine, DPX-0907 enhanced IFN-gamma+CD8+ T cells in vaccine site-draining lymph nodes, as seen by immunofluorescence staining and increased the frequency of IFN-gamma+ lymph node cells as seen by enzyme-linked immunosorbent spot assay. Notably, while conventional vaccine formulations elicited elevated levels of splenic Foxp3+CD4+ and IL10-secreting T cells, this was not the case for DPX-0907-based vaccines, with treated animals exhibiting normal levels of regulatory T cells. These data support the unique capabilities of a vaccine formulation containing novel tumor peptides and DPX-0907 to elicit type-1 dominated, specific immunity that may represent a potent clinical therapeutic modality for patients with breast or ovarian carcinoma.