A Respiratory Syncytial Virus Vaccine Based on the Small Hydrophobic Protein Ectodomain Presented With a Novel Lipid-Based Formulation Is Highly Immunogenic and Safe in Adults: A First-in-Humans Study.

Background: Respiratory syncytial virus infection can cause lower respiratory tract infection in older adults comparable to influenza, but no vaccines are available. Methods: This was a randomized, observer-blinded, first-in-humans study of a novel synthetic RSV antigen based on the ectodomain of the small hydrophobic glycoprotein (SHe) of RSV subgroup A, formulated with either the lipid and oil-based vaccine platform DepoVax (DPX-RSV[A]) or alum (RSV[A]-Alum), in healthy, 50-64-year-old individuals. Two dose levels (10 or 25 µg) of SHe with each formulation were compared to placebo. A booster dose was administered on day 56. Results: There was no indication that the vaccine was unsafe. Mild pain, drowsiness, and muscles aches were the most common solicited adverse events (AEs), and the frequencies of the AEs did not increase after dose 2. Robust anti-SHe-specific immune responses were demonstrated in the DPX-RSV(A) 10-µg and 25-µg groups (geometric mean titer, approximately 10-fold and 100-fold greater than that of placebo at days 56 and 236, respectively), and responses were sustained in the DPX-RSV(A) 25-µg group at day 421. Responses to the RSV(A)-Alum vaccines were very low. Conclusions: A novel antigen from the SH protein of RSV, formulated in a lipid and oil-based vaccine platform, was highly immunogenic, with sustained antigen-specific antibody responses, and had an acceptable safety profile.

Maveropepimut-S, a DPX-Based Immune-Educating Therapy, Shows Promising and Durable Clinical Benefit in Patients with Recurrent Ovarian Cancer, a Phase II Trial.

PURPOSE: Patients with platinum-resistant ovarian cancer respond poorly to existing therapies. Hence there is a need for more effective treatments. PATIENTS AND METHODS: The DeCidE1 trial is a multicenter, randomized, open-label, single-arm phase II study to evaluate the safety and effectiveness of maveropepimut-S with cyclophosphamide in patients with recurrent ovarian cancer. Median follow-up for evaluable subjects was 4.4 months. Data were collected from March 2019 to June 2021. Subjects received two injections of 0.25 mL maveropepimut-S 3 weeks apart, followed by one 0.1-mL doses, every 8 weeks up to progression. Oral cyclophosphamide, 50 mg twice daily, was administered in repeating weekly on and off cycles. RESULTS: Twenty-two patients were enrolled. Median age was 58 years (38-78 years). Among the evaluable population, the objective response rate (ORR) was 21% [90% confidence interval (CI), 7.5%-41.9%], with a disease control rate (DCR) of 63% (90% CI, 41.8%-81.3%), including 4 (21%) patients with partial responses, 8 (42%) stable disease, and 7 (37%) progressive disease. The ORRs were consistent across subgroups based on platinum sensitivity, and DCR was higher in the platinum-resistant subpopulation. Four SD patients maintained clinical benefit up to 25 months. Most treatment-related adverse events (TRAE) were grade 1 and 2 (87% of unique events). Most common AEs were injection site reactions. Eight subjects reported grade 3 and no grade 4 AEs. Survivin-specific T-cell responses were observed in treated patients with clinical benefit. CONCLUSIONS: Maveropepimut-S with intermittent low-dose cyclophosphamide is well-tolerated, with clinical benefit for patients with recurrent ovarian cancer. Observed responses are irrespective of the platinum status.

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.

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.

Type III hypersensitivity reactions to a B cell epitope antigen are abrogated using a depot forming vaccine platform.

Peptide antigens are combined with an adjuvant in order to increase immunogenicity in vivo. The immunogenicity and safety of a RSV vaccine formulated in a novel oil-based platform, DepoVax™ (DPX), was compared to an alum formulation. A peptide B cell epitope derived from RSV small hydrophobic ectodomain (SHe) served as the antigen. Both vaccines induced SHe-specific antibodies after immunization of mice. A single dose of the DPX-based formulation resulted in anti-SHe titres for up to 20 weeks. Boosting with Alum-SHe, but not with DPX-SHe, led to unexpected clinical signs such as decreased activity, cyanosis and drop in body temperature in mice but not in rabbits. The severity of adverse reactions correlated with magnitude of SHe-specific IgG immune responses and decreased complement component 3 plasma levels, indicating a type III hypersensitivity reaction. By RP-HPLC analysis, we found that only 8-20% of the antigen was found to be adsorbed to alum in vitro, indicating that this antigen is likely released systemically upon injection in vivo. Clinical signs were not observed in rabbits, indicating the response correlates with peptide dose relative to size of animal. These results suggest that peptide antigens targeted to produce B cell mediated response may result in increased incidence of type III hypersensitivity reactions when delivered in non-depot forming vaccines. The DPX formulation induced strong antibody titres to the antigen without causing adverse events, likely due to the strength of the depot in vivo, and demonstrates the potential safety and immunogenicity of this platform for B cell peptide antigens.

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.

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.

A C-terminal glycine suppresses production of pleurocidin as a fusion peptide in Escherichia coli.

The winter flounder (Pseudopleuronectes americanus) antimicrobial peptide pleurocidin was produced in Escherichia coli using a synthetic gene constructed by PCR. The gene expresses pleurocidin from pET21a fused to the C-terminus of an insoluble carrier peptide. Once expressed, the fusion peptide formed inclusion bodies in the cytoplasm that were collected, solubilized in guanidine-HCl, and chemically cleaved using hydroxylamine at a unique asparaginyl-glycyl dipeptide. This released recombinant pleurocidin (r-pleurocidin), which was purified using ultrafiltration followed by reverse phase chromatography. The r-pleurocidin peptide resolved as a single band (2.7 kDa) when analyzed by Tris-Tricine buffered SDS-PAGE, and its amino acid sequence was confirmed using tandem mass spectrometry. Extending the pleurocidin sequence with a C-terminal glycine (r-pleurocidin-G) suppressed production of the fusion peptide 15-fold. When pleurocidin was extended further to include aspartate (r-pleurocidin-GD), the same effect was observed, and when pleurocidin was extended with aspartate alone, no effect was observed. Expression of fusion peptide containing either r-pleurocidin-G or r-pleurocidin-GD with low concentrations of inductant caused E. coli to enter stationary phase prematurely, but did not affect overall growth rates. A partial production recovery of r-pleurocidin-G was achieved by inducing expression in stationary phase cells. We observed r-pleurocidin-G to have enhanced antimicrobial activity compared with r-pleurocidin, and we propose that this activity interferes with E. coli metabolism during expression. This antimicrobial effect is probably facilitated by residual solubility of the fusion peptide and by a C-terminal cap structure, which stabilizes the r-pleurocidin-G alpha-helix that is thought to be important for activity.