Dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN) is a cellular receptor for delta inulin adjuvant.

Delta inulin, or Advax, is a polysaccharide vaccine adjuvant that significantly enhances vaccine-mediated immune responses against multiple pathogens and was recently licensed for use in the coronavirus disease 2019 (COVID-19) vaccine SpikoGen. Although Advax has proven effective as an immune adjuvant, its specific binding targets have not been characterized. In this report, we identify a cellular receptor for Advax recognition. In vitro uptake of Advax particles by macrophage cell lines was substantially greater than that of latex beads of comparable size, suggesting an active uptake mechanism by phagocytic cells. Using a lectin array, Advax particles were recognized by lectins specific for various carbohydrate structures including mannosyl, N-acetylgalactosamine and galactose moieties. Expression in nonphagocytic cells of dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN), a C-type lectin receptor, resulted in enhanced uptake of fluorescent Advax particles compared with mock-transfected cells. Advax uptake was reduced with the addition of ethylenediaminetetraacetic acid and mannan to cells, which are known inhibitors of DC-SIGN function. Finally, a specific blockade of DC-SIGN using a neutralizing antibody abrogated Advax uptake in DC-SIGN-expressing cells. Together, these results identify DC-SIGN as a putative receptor for Advax. Given the known immunomodulatory role of DC-SIGN, the findings described here have implications for the use of Advax adjuvants in humans and inform future mechanistic studies.

Randomized controlled trial demonstrating the benefits of delta inulin adjuvanted immunotherapy in patients with bee venom allergy.

BACKGROUND: Allergic reactions to Hymenoptera insect stings remain a major global clinical problem. Although effective, parenteral desensitization regimens require use of costly venom extracts and require frequent visits over extended periods of time. OBJECTIVE: Adjuvants are commonly used to enhance the efficacy of infectious disease vaccines, and this study asked whether Advax (Vaxine Pty Ltd, Adelaide, Australia), a novel noninflammatory polysaccharide adjuvant, might provide similar benefits for allergy desensitization. METHODS: A randomized, controlled phase 1/2 trial was undertaken in 27 adults with a history of rapid-onset systemic allergic reactions to honeybee stings and positive specific IgE levels to evaluate the safety and efficacy of honeybee venom immunotherapy (HBVIT) combined with Advax adjuvant. Venom immunotherapy (VIT) was administered monthly for 30 months after achievement of maintenance doses. RESULTS: Advax-adjuvanted HBVIT was well tolerated. Around week 14 of VIT, specific IgG4 responses peaked in both groups but increased earlier, peaked higher, and were better maintained through the end of the study in the Advax-adjuvanted arm. Several different patterns of serologic response to VIT were seen; some subjects had a dominant IgG4 response, some had a combined IgG4 and IgG1 response, and some had an exclusively IgG1 response. In some subjects specific IgE levels increased during the induction phase and then decreased, whereas in others specific IgE levels progressively decreased from the start of VIT. CONCLUSION: Advax adjuvant favorably enhanced the immunogenicity of HBVIT, with an early and prolonged switch to specific IgG4 production. The ability of Advax adjuvant to enhance VIT efficacy warrants further study.

Replacement of glycaemic carbohydrates by inulin-type fructans from chicory (oligofructose, inulin) reduces the postprandial blood glucose and insulin response to foods: report of two double-blind, randomized, controlled trials.

PURPOSE: Inulin-type fructans are recognized as prebiotic dietary fibres and classified as non-digestible carbohydrates that do not contribute to glycaemia. The aim of the present studies was to investigate the glycaemic response (GR) and insulinaemic response (IR) to foods in which sucrose was partially replaced by inulin or oligofructose from chicory. METHODS: In a double-blind, randomized, controlled cross-over design, 40-42 healthy adults consumed a yogurt drink containing oligofructose or fruit jelly containing inulin and the respective full-sugar variants. Capillary blood glucose and insulin were measured in fasted participants and at 15, 30, 45, 60, 90, and 120 min after starting to drink/eat. For each test food, the incremental area under the curve (iAUC) for glucose and insulin was calculated and the GR and IR determined. RESULTS: Consumption of a yogurt drink with oligofructose which was 20% reduced in sugars significantly lowered the glycaemic response compared to the full-sugar reference (iAUC120min 31.9 and 37.3 mmol/L/min, respectively; p < 0.05). A fruit jelly made with inulin and containing 30% less sugars than the full-sugar variant likewise resulted in a significantly reduced blood glucose response (iAUC120min 53.7 and 63.7 mmol/L/min, respectively; p < 0.05). In both studies, the postprandial insulin response was lowered in parallel (p < 0.05). The reduction of postprandial glycaemia was positively correlated to the proportion of sugars replaced by inulin-type fructans (p < 0.001). CONCLUSIONS: In conclusion, the studies confirmed that substitution of glycaemic sugars by inulin or oligofructose from chicory may be an effective strategy to reduce the postprandial blood glucose response to foods.

Optimizing Water Exchange Rates and Rotational Mobility for High-Relaxivity of a Novel Gd-DO3A Derivative Complex Conjugated to Inulin as Macromolecular Contrast Agents for MRI.

Thanks to the understanding of the relationships between the residence lifetime τM of the coordinated water molecules to macrocyclic Gd-complexes and the rotational mobility τR of these structures, and according to the theory for paramagnetic relaxation, it is now possible to design macromolecular contrast agents with enhanced relaxivities by optimizing these two parameters through ligand structural modification. We succeeded in accelerating the water exchange rate by inducing steric compression around the water binding site, and by removing the amide function from the DOTA-AA ligand [1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid mono(p-aminoanilide)] (L) previously designed. This new ligand 10[2(1-oxo-1-p-propylthioureidophenylpropyl]-1,4,7,10-tetraazacyclodecane-1,4,7-tetraacetic acid (L1 ) was then covalently conjugated to API [O-(aminopropyl)inulin] to get the complex API-(GdL1 )x with intent to slow down the rotational correlation time (τR ) of the macromolecular complex. The evaluation of the longitudinal relaxivity at different magnetic fields and the study of the 17 O-NMR at variable temperature of the low-molecular-weight compound (GdL1 ) showed a slight decrease of the τM value (τM310 = 331 ns vs. τM310 = 450 ns for the GdL complex). Consequently to the increase of the size of the API-(GdL1 )x complex, the rotational correlation time becomes about 360 times longer compared to the monomeric GdL1 complex (τR  = 33,700 ps), which results in an enhanced proton relaxivity.

Rethinking CKD Evaluation: Should We Be Quantifying Basal or Stimulated GFR to Maximize Precision and Sensitivity?

Chronic kidney disease (CKD) is an increasing clinical problem. Although clinical risk factors and biomarkers for the development and progression of CKD have been identified, there is no commercial surveillance technology to definitively diagnose and quantify the severity and progressive loss of glomerular filtration rate (GFR) in CKD. This has limited the study of potential therapies to late stages of CKD when FDA-registerable events are more likely. Because patient outcomes, including the rate of CKD progression, correlate with disease severity and effective therapy may require early intervention, being able to diagnose and stratify patients by their level of decreased kidney function early on is key for translational progress. In addition, renal reserve, defined as the increase in GFR following stimulation, may improve the quantification of GFR based solely on basal levels. Various groups are developing and characterizing optical measurement techniques using new minimally invasive or noninvasive approaches for quantifying basal and stimulated kidney function. This development has the potential to allow widespread individualization of therapy at an earlier disease stage. Therefore, the purposes of this review are to suggest why quantifying stimulated GFR, by activating renal reserve, may be advantageous in patients and to review fluorescent technologies to deliver patient-specific GFR.

Severe acute respiratory syndrome-associated coronavirus vaccines formulated with delta inulin adjuvants provide enhanced protection while ameliorating lung eosinophilic immunopathology.

UNLABELLED: Although the severe acute respiratory syndrome-associated coronavirus (SARS-CoV) epidemic was controlled by nonvaccine measures, coronaviruses remain a major threat to human health. The design of optimal coronavirus vaccines therefore remains a priority. Such vaccines present major challenges: coronavirus immunity often wanes rapidly, individuals needing to be protected include the elderly, and vaccines may exacerbate rather than prevent coronavirus lung immunopathology. To address these issues, we compared in a murine model a range of recombinant spike protein or inactivated whole-virus vaccine candidates alone or adjuvanted with either alum, CpG, or Advax, a new delta inulin-based polysaccharide adjuvant. While all vaccines protected against lethal infection, addition of adjuvant significantly increased serum neutralizing-antibody titers and reduced lung virus titers on day 3 postchallenge. Whereas unadjuvanted or alum-formulated vaccines were associated with significantly increased lung eosinophilic immunopathology on day 6 postchallenge, this was not seen in mice immunized with vaccines formulated with delta inulin adjuvant. Protection against eosinophilic immunopathology by vaccines containing delta inulin adjuvants correlated better with enhanced T-cell gamma interferon (IFN-γ) recall responses rather than reduced interleukin-4 (IL-4) responses, suggesting that immunopathology predominantly reflects an inadequate vaccine-induced Th1 response. This study highlights the critical importance for development of effective and safe coronavirus vaccines of selection of adjuvants based on the ability to induce durable IFN-γ responses. IMPORTANCE: Coronaviruses such as SARS-CoV and Middle East respiratory syndrome-associated coronavirus (MERS-CoV) cause high case fatality rates and remain major human public health threats, creating a need for effective vaccines. While coronavirus antigens that induce protective neutralizing antibodies have been identified, coronavirus vaccines present a unique problem in that immunized individuals when infected by virus can develop lung eosinophilic pathology, a problem that is further exacerbated by the formulation of SARS-CoV vaccines with alum adjuvants. This study shows that formulation of SARS-CoV spike protein or inactivated whole-virus vaccines with novel delta inulin-based polysaccharide adjuvants enhances neutralizing-antibody titers and protection against clinical disease but at the same time also protects against development of lung eosinophilic immunopathology. It also shows that immunity achieved with delta inulin adjuvants is long-lived, thereby overcoming the natural tendency for rapidly waning coronavirus immunity. Thus, delta inulin adjuvants may offer a unique ability to develop safer and more effective coronavirus vaccines.

A new method to measure intestinal secretion using fluorescein isothiocyanate-inulin in small bowel of rats.

BACKGROUND: Small intestine ischemia can be seen in various conditions such as intestinal transplantation. To further understand the pathologic disruption in ischemia-reperfusion injury, we have developed a method to measure fluid changes in the intestinal lumen of rats. METHODS: Two 10-cm rat intestine segments were procured, connected to the terminal apertures of a perfusion device, and continuously infused with 3 mL of HEPES solution (control solution) containing 50 µM of fluorescein isothiocyanate (FITC)-inulin. The perfusion device consists of concentric chambers that contain the perfused bowel segments, which are maintained at 37°C via H2O bath. The individual chamber has four apertures as follows: two fill and/or drain the surrounding HEPES solution on the blood side of the tissue. The others provide flow of HEPES solution containing FITC-inulin through the lumens. The experimental intestine was infused with the same solution with 100 µM of Forskolin. A pump continuously circulated solutions at 6 mL/min. Samples were collected at 15-min intervals until 150 min and were measured by the nanoflourospectrometer. RESULTS: A mean of 6-µM decrease in the FITC-inulin concentration in the Forskolin-treated experimental intestine was observed in comparison with that in the control intestine. The FITC-inulin count dilution in the experimental intestine is a result of an increase of fluid secretion produced by the effect of Forskolin, with P values <0.0001. CONCLUSIONS: We demonstrate that it is possible to measure luminal fluid changes over time using our new modified perfusion system along with FITC-inulin to allow real-time determinations of fluid and/or electrolyte movement along the small intestine.

JE-ADVAX vaccine protection against Japanese encephalitis virus mediated by memory B cells in the absence of CD8(+) T cells and pre-exposure neutralizing antibody.

JE-ADVAX is a new, delta inulin-adjuvanted, Japanese encephalitis (JE) candidate vaccine with a strong safety profile and potent immunogenicity that confers efficient immune protection not only against JE virus but also against related neurotropic flaviviruses such as West Nile virus. In this study, we investigated the immunological mechanism of protection by JE-ADVAX vaccine using knockout mice deficient in B cells or CD8(+) T cells and poor persistence of neutralizing antibody or by adoptive transfer of immune splenocyte subpopulations. We show that memory B cells induced by JE-ADVAX provide long-lived protection against JE even in the absence of detectable pre-exposure serum neutralizing antibodies and without the requirement of CD8(+) T cells. Upon virus encounter, these vaccine-induced memory B cells were rapidly triggered to produce neutralizing antibodies that then protected immunized mice from morbidity and mortality. The findings suggest that the extent of the B-cell memory compartment might be a better immunological correlate for clinical efficacy of JE vaccines than the currently recommended measure of serum neutralizing antibody. This may explain the paradox where JE protection is observed in some subjects even in the absence of detectable serum neutralizing antibody. Our investigation also established the suitability of a novel flavivirus challenge model (ß(2)-microglobulin-knockout mice) for studies of the role of B-cell memory responses in vaccine protection.

An inactivated cell culture Japanese encephalitis vaccine (JE-ADVAX) formulated with delta inulin adjuvant provides robust heterologous protection against West Nile encephalitis via cross-protective memory B cells and neutralizing antibody.

West Nile virus (WNV), currently the cause of a serious U.S. epidemic, is a mosquito-borne flavivirus and member of the Japanese encephalitis (JE) serocomplex. There is currently no approved human WNV vaccine, and treatment options remain limited, resulting in significant mortality and morbidity from human infection. Given the availability of approved human JE vaccines, this study asked whether the JE-ADVAX vaccine, which contains an inactivated cell culture JE virus antigen formulated with Advax delta inulin adjuvant, could provide heterologous protection against WNV infection in wild-type and ß2-microglobulin-deficient (ß2m(-/-)) murine models. Mice immunized twice or even once with JE-ADVAX were protected against lethal WNV challenge even when mice had low or absent serum cross-neutralizing WNV titers prior to challenge. Similarly, ß2m(-/-) mice immunized with JE-ADVAX were protected against lethal WNV challenge in the absence of CD8(+) T cells and prechallenge WNV antibody titers. Protection against WNV could be adoptively transferred to naive mice by memory B cells from JE-ADVAX-immunized animals. Hence, in addition to increasing serum cross-neutralizing antibody titers, JE-ADVAX induced a memory B-cell population able to provide heterologous protection against WNV challenge. Heterologous protection was reduced when JE vaccine antigen was administered alone without Advax, confirming the importance of the adjuvant to induction of cross-protective immunity. In the absence of an approved human WNV vaccine, JE-ADVAX could provide an alternative approach for control of a major human WNV epidemic.

Measurement of glomerular filtration rate during flight in a migratory bird using a single bolus injection of FITC-inulin.

During migration, passerine birds typically complete a series of multi-hour flights, each followed by a period of stopover. During flight, rates of respiratory water loss are high, yet these birds show no signs of dehydration after flights. During stopover, birds become hyperphagic to replenish fat reserves, often consuming food with high water content, such as fruit. Thus migratory birds seem to face an osmoregulatory challenge; they must reduce water losses during flight but retain the ability to excrete large quantities of water while maintaining osmotic balance at stopover. Our goal was to measure glomerular filtration rate (GFR) and fractional water reabsorption (FWR) of a migratory bird in free flight, at rest, and during feeding to assess the role of the kidney in maintaining water balance during migration. We used FITC-inulin and one- and two-phase exponential decay models to first validate a technique and then measure GFR in the Swainson's thrush, a small (∼30 g) songbird. Single-phase exponential decay models and the modified slope intercept method overestimated GFR by 26% compared with two-phase exponential decay models. We found no differences in GFR among fed, resting and flying birds, but FWR was significantly higher in resting and flying birds relative to feeding birds. There was no effect of the rate of respiratory water loss on GFR or FWR in flight. These data support the idea that birds in flight do not dramatically alter GFR but rely on increased FWR to minimize excretory water losses.

Synthesis, characterization and self-assembly of biosurfactants based on hydrophobically modified inulins.

Biosurfactants have been synthesized using a low energy, environmentally friendly process by the derivatization of inulin with octenyl (OSA) and dodecenyl (DDSA) succinic anhydrides in aqueous solution. The inulin and its derivatives have been characterized using gel permeation chromatography/multi angle light scattering (GPC/MALLS), high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD), Fourier transform infrared spectroscopy (FTIR), and NMR, and the reaction efficiency was found to be between 59 and 95%. The efficiency was generally higher for OSA derivatives compared to DDSA derivatives. The hydrophobic derivatives were found to aggregate in solution and the critical aggregation concentration (CAC) was determined using dye solubilization, surface tension, dynamic light scattering (DLS), and conductivity. There was reasonable agreement in the CAC values obtained by the different techniques except for conductivity. It was found that the CAC decreased with increasing alkenyl chain length and degree of modification, and the values were significantly lower for the DDSA derivatives compared to the OSA derivatives. GPC elution profiles for the DDSA-inulin using 12 mol % reagent confirmed the presence of aggregates with a molecular mass of ∼2.5 × 10(6) g/mol and a radius of gyration of ∼25 nm corresponding to ∼550 inulin molecules. DLS study was undertaken to determine the hydrodynamic radius, and values obtained for the DDSA (12%) derivative were 30 nm in both water and 0.1 M sodium nitrate, while for the OSA (12%) derivative values of 13 and 7 nm were obtained. The derivatives have potential application in the stabilization of particulate dispersions and emulsions and also in the encapsulation and delivery of drugs.

[Relationship between the anticoagulant activity of sulfated plant polysaccharides and the area of their precipitation with polycations during biospecific electrophoresis].

Polyanions (in an amount within 1.5 - 6.0 mg), including cellulose sulfates (excreted from Gossipium hirsutum L., molecular weight 22.0 kDa, degree of sulfation within 0.8 - 1.8), inulin sulfates (excreted from Helianthus tuberosus, molecular weight 8.0 kDa, degree of sulfation within 0.6 - 1.6), pectin sulfates (excreted from Abies sibirica L., molecular weight 24.0 kDa, degree of sulfation within 0.8 - 1.1), give rise to peaks of precipitation with polycations of protamine sulfate. Only cellulose sulfates (in amount within 0.38 - 6.00 mg) give the peaks of precipitation with chitosan polycations (molecular weight 10 kDa, degree of deacetylation 85%) during horizontal biospecific electrophoresis. The height of the peak of precipitation with protamin sulfate was found to grow with increasing antithrombin activity of cellulose sulfates and pectin sulfate (for polyanions in an amount within 1.5 - 6 mg). The size of the area of precipitation with chitosan was found to decrease with increasing antithrombin activity of cellulose sulfates.

[Dependence of the anticoagulant activity of starch and inulin on their degree of sulfonation].

We have studied a relationship between the degree of sulfonation and anticoagulant activity of starch from Solanum tuberosum (molecular weight, 25000-30000 Da; sulfonation degree, 0.4-2.5) and inulin from Helianthus tuberosus (molecular weight, 7000-8000 Da; sulfonation degree, 0.6-1.6). Starch and inulin sulfates (i) increased the time of appearance of fibrin clots in plasma in coagulometric tests and (ii) reduced (via antithrombin) the rate of thrombin-induced hydrolysis of a chromogen substrate. The antithrombin (aIIa) activity of starch sulfates reached 16.8-70.0 IU/mg and the activity against factor Xa (aXa activity) was 2.3-16.6 IU/mg. The antithrombin activity of inulin sulfates was within 5.5-11.4 IU/mg and the activity against factor Xa (aXa activity) was within 0-1.4 IU/mg. An increase in the degree of sulfonation led to a growth in the anticoagulant activity of starch sulfates. The anticoagulant activity of starch sulfates and inulin sulfate with sulfonation degree 1.0 is mediated by antithrombin, which is the plasma inhibitor of serine proteases.

Evaluation of the safety and efficacy of AdvaxTM as an adjuvant: A systematic review and meta-analysis.

PURPOSE: Developing a vaccine with improved immunogenicity is still a growing priority for many diseases. Different types of adjuvants may be beneficial to initiate and maintain the long-lasting immunogenicity of vaccines. Evidence has shown that polysaccharide adjuvants are efficient in improving immunological mechanisms with their biocompatibility and biodegradability characteristics. In this study, we aimed to investigate the safety and efficacy of AdvaxTM an adjuvant derived from delta inulin. METHODS: A systematic research was performed in Pubmed, Web of Science, and Scopus databases for the following keywords; "AdvaxTM" OR "delta inulin" until December 14th, 2020. RevMan 5.4.1 software was used for cumulative meta-analysis and bias analysis. We also used GraphPad Prism 6 software for the figures. RESULTS: In the cumulative meta-analysis, it was found that seroconversion and geometric mean titers (GMT) levels significantly increased in AdvaxTM-adjuvanted group (mean difference: 12.31, 95% Cl [4.14, 20.47], p â€‹= â€‹0.003; 17.10, 95% Cl [4.35, 29.85], p â€‹= â€‹0.009, respectively). We also observed that AdvaxTM could be effective in improving immunogenicity by inducing T-cell responses and plasmablast generation in viral vaccines. CONCLUSIONS: In this study, it was shown that AdvaxTM is a safe and well-tolerated adjuvant. AdvaxTM could be a potent adjuvant in increasing the protection and immunogenicity of different vaccines without safety issues. However, further studies are needed to verify these effects of AdvaxTM adjuvant.

A typhoid fever protein capsular matrix vaccine candidate formulated with Advax-CpG adjuvant induces a robust and durable anti-typhoid Vi polysaccharide antibody response in mice, rabbits and nonhuman primates.

Typhax is an investigational typhoid fever vaccine candidate that is comprised of Vi polysaccharide from Salmonella enterica serovar typhi (S. Typhi) non-covalently entrapped in a glutaraldehyde catalyzed, cross-linked α-poly-L-lysine and CRM197 protein matrix. A previous Phase 1 trial of an aluminum phosphate adjuvanted Typhax formulation showed it induced Vi IgG after a single dose but that subsequent doses failed to further boost Vi IgG levels. The current study asked whether Advax-CpG adjuvant might instead be able to overcome polysaccharide-induced immune inhibition and improve Typhax immunogenicity. Advax-CpG adjuvanted Typhax elicited high and sustained Vi IgG responses in mice, rabbits and non-human primates (NHP) with levels being boosted by repeated immunization. High Vi antibody responses were lost in CD4 + T cell depleted mice confirming that despite the lack of conjugation of the polysaccharide to the carrier protein, Typhax nevertheless acts in a T cell dependent manner, explaining its ability to induce long-term B cell memory responses to Vi capable of being boosted. In NHP, Advax-CpG adjuvanted Typhax induced up to 100-fold higher Vi IgG levels than the commercial Typhim Vi polysaccharide vaccine. Typhax induced high and sustained serum bactericidal activity against S. Typhi and stimulated robust Vi IgG responses even in animals previously primed with a pure polysaccharide vaccine. Hence Advax-CpG adjuvanted Typhax vaccine is a highly promising candidate to provide robust and durable protection against typhoid fever.

Covax-19/Spikogen® vaccine based on recombinant spike protein extracellular domain with Advax-CpG55.2 adjuvant provides single dose protection against SARS-CoV-2 infection in hamsters.

COVID-19 presents an ongoing global health crisis. Protein-based COVID-19 vaccines that are well-tolerated, safe, highly-protective and convenient to manufacture remain of major interest. We therefore sought to compare the immunogenicity and protective efficacy of a number of recombinant SARS-CoV-2 spike protein candidates expressed in insect cells. By comparison to a full length (FL) spike protein detergent-extracted nanoparticle antigen, the soluble secreted spike protein extracellular domain (ECD) generated higher protein yields per liter of culture and when formulated with either Alum-CpG55.2 or Advax-CpG55.2 combination adjuvants elicited robust antigen-specific humoral and cellular immunity in mice. In hamsters, the spike ECD when formulated with either adjuvant induced high serum neutralizing antibody titers even after a single dose. When challenged with the homologous SARS-CoV-2 virus, hamsters immunized with the adjuvanted spike ECD exhibited reduced viral load in day 1-3 oropharyngeal swabs and day 3 nasal turbinate tissue and had no recoverable infectious virus in day 3 lung tissue. The reduction in lung viral load correlated with less weight loss and lower lung pathology scores. The formulations of spike ECD with Alum-CpG55.2 or Advax-CpG55.2 were protective even after just a single dose, although the 2-dose regimen performed better overall and required only half the total amount of antigen. Pre-challenge serum neutralizing antibody levels showed a strong correlation with lung protection, with a weaker correlation seen with nasal or oropharyngeal protection. This suggests that serum neutralizing antibody levels may correlate more closely with systemic, rather than mucosal, protection. The spike protein ECD with Advax-CpG55.2 formulation (Covax-19® vaccine) was selected for human clinical development.

Safety and immunogenicity of SpikoGen®, an Advax-CpG55.2-adjuvanted SARS-CoV-2 spike protein vaccine: a phase 2 randomized placebo-controlled trial in both seropositive and seronegative populations.

OBJECTIVE: We aimed to investigate the immunogenicity and safety of SpikoGen®, a subunit COVID-19 vaccine composed of a recombinant prefusion-stabilized SARS-CoV-2 spike protein combined with the Advax-CpG55.2™ adjuvant, in seronegative and seropositive populations as primary vaccination. METHODS: This randomized, placebo-controlled, double-blind phase 2 trial was conducted on 400 participants randomized 3:1 to receive two doses of 25 µg of SpikoGen® 3 weeks apart or the placebo. The primary safety outcomes were the incidence of solicited adverse events up to 7 days after each dose and unsolicited adverse events up to 28 days after the second dose. The primary immunogenicity outcomes were seroconversion against the S1 protein and the geometric mean concentration of S1 antibodies by days 21 and 35. RESULTS: The SpikoGen® vaccine was well tolerated and no serious adverse events were recorded. The most common solicited adverse events were injection site pain and fatigue, largely graded as mild and transient. By day 35 (2 weeks post second dose), the seroconversion rate against S1 was 63.55 (95% CI: 57.81-69.01) in the SpikoGen® group versus 7.23 (95% CI: 2.7-15.07) in the placebo group. The geometric mean concentration of S1 antibodies was 29.12 (95% CI: 24.32-34.87) in the SpikoGen® group versus 5.53 (95% CI: 4.39-6.97) in the placebo group. Previously infected seropositive volunteers showed a large SARS-CoV-2 humoral response after a single SpikoGen® dose. DISCUSSION: SpikoGen® had an acceptable safety profile and induced promising humoral and cellular immune responses against SARS-CoV-2.

Prebiotic oligosaccharides reduce proinflammatory cytokines in intestinal Caco-2 cells via activation of PPARγ and peptidoglycan recognition protein 3.

Prebiotic oligosaccharides modulate the intestinal microbiota and beneficially affect the human body by reducing intestinal inflammation. This immunomodulatory effect was assumed to be bacterial in origin. However, some observations suggest that oligosaccharides may exert an antiinflammatory effect per se. We hypothesized that oligosaccharides affect the intestinal immunity via activation of peptidoglycan recognition protein 3 (PGlyRP3), which reduces the expression of proinflammatory cytokines. Caco-2 cells were treated with the oligosaccharides, α3-sialyllactose, or fructooligosaccharides (Raftilose p95), and the effects of these treatments on PGlyRP3 and PPARγ expression, the release and expression of some proinflammatory cytokines, and NF-κB translocation were tested. Both oligosaccharides had antiinflammatory activity; they significantly reduced IL-12 secretion in Caco-2 cells and gene expression of IL-12p35, IL-8, and TNFα. They also reduced the gene expression and nuclear translocation of NF-κB. Both oligosaccharides dose and time dependently induced the production of PGlyRP3, the silencing of which by transfection of Caco-2 cells with specific small interfering RNA targeting PGlyRP3 abolished the antiinflammatory role of both oligosaccharides. Incubation of Caco-2 cells with both oligosaccharides induced PPARγ. Antagonizing PPARγ by culturing the cells with GW9662 for 24 h inhibited the oligosaccharide-induced PGlyRP3 production and the antiinflammatory effect of the oligosaccharides. We conclude that oligosaccharides may exert an antiinflammatory effect by inducing the nuclear receptor PPARγ, which regulates the antiinflammatory PGlyRP3.

Advax, as a Co-adjuvant, in Combination with Poly(I:C) Elicits Enhanced Th1 Immune Responses and Parasite Growth-Inhibitory Antibodies Against Plasmodium Falciparum Merozoite Surface Protein-1 (PfMSP-142) in BALB/c Mice.

BACKGROUND: One of the main challenges in protein-based vaccines is the poor immunogenicity of antigens, which can be solved by the use of adjuvants. Advax is a novel microparticle polysaccharide adjuvant that in combination with antigens can induce both cellular and humoral immunity based on the intrinsic features of the antigen. It has been shown that poly(I:C) can be a suitable adjuvant for the PfMSP-142-based malaria vaccine. Advax is a suitable co-adjuvant for poly(I:C) to increase its half-life and reduce dose-dependent toxicity. OBJECTIVES: To investigate whether advax alone or advax /poly(I:C) combination can enhance the immunogenicity with increased parasite inhibitory anti-PfMSP-142 antibodies in comparison to poly(I:C). METHODS: Mice groups were inoculated with rPfMSP-142 alone or formulated in poly(I:C), poly(I:C)/advax, or advax. Then, humoral and cellular immune responses, the ratio of Th1/Th2 and growth inhibitory activity of induced antibodies were analyzed. RESULTS: Poly(I:C)/advax formulated PfMSP-142 induced higher levels of anti-PfMSP-142 IgG, IgG2a, and IgG2b antibodies relative to poly(I:C)-formulated PfMSP-142. The maximum ratio of IFN-?/IL-4 (50.13) and IgG2a/IgG1 (2.65), was induced in mice receivedadvax-formulated PfMSP-142. Besides, poly(I:C)/advax formulated PfMSP-142 induced a higher ratio of IFN-?/IL-4 (25.33) and IgG2a/IgG1 (1.89) when compared with poly(I:C) alone. Strong growth inhibitory activity was observed in antibodies induced in mice received poly(I:C)/advax-formulated PfMSP-142. CONCLUSION: These findings indicate that advax is a favorable adjuvant to be combined with poly(I:C), and this combination of adjuvants could induce Th1 immune responses and growth inhibitory antibodies against rPfMSP-142.

Advax adjuvant formulations promote protective immunity against aerosol Mycobacterium tuberculosis in the absence of deleterious inflammation and reactogenicity.

The development of safe and effective adjuvants is a critical goal of vaccine development programs. In this report, we defined the immunostimulatory profile and protective effect against aerosol Mycobacterium tuberculosis infection of vaccine formulations incorporating the semi-crystalline adjuvant δ-inulin (Advax). Advax formulated with CpG oligonucleotide and the QS-21 saponin (AdvaxCpQS) was the most effective combination, demonstrated by the capacity of CysVac2/AdvaxCpQS to significantly reduce the bacterial burden in the lungs of M. tuberculosis-infected mice. CysVac2/AdvaxCpQS protection was associated with rapid influx of neutrophils, macrophages and monocytes to the site of vaccination and the induction of antigen-specific IFN-γ+/IL-2+/TNF+ polyfunctional CD4+ T cells in the lung. When compared to the highly potent adjuvant combination of monophosphoryl lipid A and dimethyldioctadecylammonium bromide (MPL/DDA), AdvaxCpQS imparted a similar level of protective efficacy yet without the profound stimulation of inflammatory cytokines and vaccination site ulceration observed with MPL/DDA. Addition of DDA to CysVac2/AdvaxCpQS further improved the protective effect of the vaccine, which correlated with increased polyfunctional CD4+ T cells in the lung but with no increase in vaccine reactogenicity. The data demonstrate that Advax formulations can decouple protective tuberculosis immunity from reactogenicity, making them ideal candidates for human application.