Potent Adjuvant Effect Research Articles

Testing S. sonnei GMMA with and without Aluminium Salt-Based Adjuvants in Animal Models.

Shigellosis is one of the leading causes of diarrheal disease in low- and middle-income countries, particularly in young children, and is more often associated with antimicrobial resistance. Therefore, a preventive vaccine against shigellosis is an urgent medical need. We have proposed Generalised Modules for Membrane Antigens (GMMA) as an innovative delivery system for Shigella sonnei O-antigen, and an Alhydrogel formulation (1790GAHB) has been extensively tested in preclinical and clinical studies. Alhydrogel has been used as an adsorbent agent with the main purpose of reducing potential GMMA systemic reactogenicity. However, the immunogenicity and systemic reactogenicity of this GMMA-based vaccine formulated with or without Alhydrogel have never been compared. In this work, we investigated the potential adjuvant effect of aluminium salt-based adjuvants (Alhydrogel and AS37) on S. sonnei GMMA immunogenicity in mice and rabbits, and we found that S. sonnei GMMA alone resulted to be strongly immunogenic. The addition of neither Alhydrogel nor AS37 improved the magnitude or the functionality of vaccine-elicited antibodies. Interestingly, rabbits injected with either S. sonnei GMMA adsorbed on Alhydrogel or S. sonnei GMMA alone showed a limited and transient body temperature increase, returning to baseline values within 24 h after each vaccination. Overall, immunisation with unadsorbed GMMA did not raise any concern for animal health. We believe that these data support the clinical testing of GMMA formulated without Alhydrogel, which would allow for further simplification of GMMA-based vaccine manufacturing.

Adjuvant effects of β-defensin on DNA vaccine OmpC against edwardsiellosis in flounder (Paralichthys olivaceus)

β-defensin of flounder plays an important role in immunomodulation by recruiting immune cells and has a potential vaccine adjuvant effect in addition to its bactericidal activity. In this study, adjuvant effects of β-defensin on DNA vaccine OmpC against edwardsiellosis in flounder (Paralichthys olivaceus) were investigated. The bicistronic eukaryotic expression plasmid pBudCE4.1 plasmid vector with two independent coding regions was selected to construct DNA vaccine of p-OmpC which express only the gene for the outer membrane protein of Edwardsiella tarda and the vaccine of p-OmpC-βdefensin which express both the outer membrane protein of the bacterium and β-defensin of flounder. In vitro and in vivo studies have shown that the constructed plasmids can be expressed in flounder embryonic cell lines and injection sites of muscles. After vaccination by intramuscular injection, both p-OmpC and p-OmpC-βdefensin groups showed significant upregulation of immune-response. Compared to the pBbudCE4.1 and the p-OmpC vaccinated groups, the p-OmpC-βdefensin vaccinated group showed significantly more cell aggregation at the injection site and intense immune response. The proportion of sIgM+ cells, as well as the CD4-1+ and CD4-2+ cells in both spleen and kidney was significantly higher in the p-OmpC-βdefensin vaccinated group at peak time point than in the control groups. The relative survival rate of the p-OmpC-βdefensin vaccine was 74.17%, which was significantly higher than that of the p-OmpC vaccinated group 48.33%. The results in this study determined that β-defensin enhances the responses in cellular and humoral immunity and evokes a high degree of protection against E. tarda, which is a promising candidate for vaccine adjuvant.

Plant-derived galactolipids enhance specific antibody production and induce class-switch as vaccine adjuvant

Various plant-derived compounds can activate immune responses against bacterial infections, and this property contributes to them being developed as effective and safe adjuvants for vaccines. This study evaluated the potential adjuvant effects of a galactolipid-enriched fraction generated from the medicinal plant Crassocephalum rabens (designated CRA). Heat shock protein 60 of periodontal disease pathogen Actinobacillus actinomycetemcomitans (AaHSP60) was taken as an antigen and mixed with CRA. The AaHSP60/CRA mixture was then injected intraperitoneally into the BALB/c mice. Titers and affinity of specific antibodies were measured by ELISA. Cytokine profiles in mouse serum or culture media of AaHSP60/CRA-treated splenocytes were analyzed by cytokine multiplex assay and ELISA kits. B cell differentiation and macrophage activation were determined by phenotyping. CRA dramatically enhanced specific antibody titers and induced Ig class switch, as shown by increases in the IgG2a, IgG2b, and IgG3 proportions of total Ig in mouse serum. Furthermore, CRA-induced anti-AaHSP60 antibodies had cross-reactivity to other bacterial HSP60s. Cell-based and animal results demonstrated that CRA induced the release of IL-21 and B cell activating factor (BAFF), which stimulated B cell differentiation. CRA enhanced cell proliferation, uptake ability, and antigen presentation in mouse phagocytes. CRA served as a vaccine adjuvant that enhance mouse immunity against pathogenic antigens. CRA strengthened the activation and capabilities of phagocytes and B cells. Therefore, CRA may be a promising adjuvant for bacterial vaccines including periodontal disease.

Micronanoparticled risedronate exhibits potent vaccine adjuvant effects

Micro/Nano-scale particles are widely used as vaccine adjuvants to enhance immune response and improve antigen stability. While aluminum salt is one of the most common adjuvants approved for human use, its immunostimulatory capacity is suboptimal. In this study, we modified risedronate, an immunostimulant and anti-osteoporotic drug, to create zinc salt particle-based risedronate (Zn-RS), also termed particulate risedronate. Compared to soluble risedronate, micronanoparticled Zn-RS adjuvant demonstrated increased recruitment of innate cells, enhanced antigen uptake locally, and a similar antigen depot effect as aluminum salt. Furthermore, Zn-RS adjuvant directly and quickly stimulated immune cells, accelerated the formulation of germinal centers in lymph nodes, and facilitated the rapid production of antibodies. Importantly, Zn-RS adjuvant exhibited superior performance in both young and aged mice, effectively protecting against respiratory diseases such as SARS-CoV-2 challenge. Consequently, particulate risedronate showed great potential as an immune-enhancing vaccine adjuvant, particularly beneficial for vaccines targeting the susceptible elderly.

Diversification of a Novel α-Galactosyl Ceramide Hotspot Boosts the Adjuvant Properties in Parenteral and Mucosal Vaccines.

The development of potent adjuvants is an important step for improving the performance of subunit vaccines. CD1d agonists, such as the prototypical α-galactosyl ceramide (α-GalCer), are of special interest due to their ability to activate iNKT cells and trigger rapid dendritic cell maturation and B-cell activation. Herein, we introduce a novel derivatization hotspot at the α-GalCer skeleton, namely the N-substituent at the amide bond. The multicomponent diversification of this previously unexplored glycolipid chemotype space permitted the introduction of a variety of extra functionalities that can either potentiate the adjuvant properties or serve as handles for further conjugation to antigens toward the development of self-adjuvanting vaccines. This strategy led to the discovery of compounds eliciting enhanced antigen-specific T cell stimulation and a higher antibody response when delivered by either the parenteral or the mucosal route, as compared to a known potent CD1d agonist. Notably, various functionalized α-GalCer analogues showed a more potent adjuvant effect after intranasal immunization than a PEGylated α-GalCer analogue previously optimized for this purpose. Ultimately, this work could open multiple avenues of opportunity for the use of mucosal vaccines against microbial infections.

Diversification of a Novel α‐Galactosyl Ceramide Hotspot Boosts the Adjuvant Properties in Parenteral and Mucosal Vaccines

AbstractThe development of potent adjuvants is an important step for improving the performance of subunit vaccines. CD1d agonists, such as the prototypical α‐galactosyl ceramide (α‐GalCer), are of special interest due to their ability to activate iNKT cells and trigger rapid dendritic cell maturation and B‐cell activation. Herein, we introduce a novel derivatization hotspot at the α‐GalCer skeleton, namely the N‐substituent at the amide bond. The multicomponent diversification of this previously unexplored glycolipid chemotype space permitted the introduction of a variety of extra functionalities that can either potentiate the adjuvant properties or serve as handles for further conjugation to antigens toward the development of self‐adjuvanting vaccines. This strategy led to the discovery of compounds eliciting enhanced antigen‐specific T cell stimulation and a higher antibody response when delivered by either the parenteral or the mucosal route, as compared to a known potent CD1d agonist. Notably, various functionalized α‐GalCer analogues showed a more potent adjuvant effect after intranasal immunization than a PEGylated α‐GalCer analogue previously optimized for this purpose. Ultimately, this work could open multiple avenues of opportunity for the use of mucosal vaccines against microbial infections.

Nanoadjuvants Produced by Advanced Nanochelating Technology in the Inactivated-Severe Acute Respiratory Syndrome Coronavirus-2 Vaccine Formulation: Preliminary Results on Cytokines and IgG Responses.

Despite the great success of vaccines in various infectious diseases, most current vaccines are not effective enough, and on the contrary, clinically approved alum adjuvants cannot induce sufficient immune responses, including a potent cellular immune response to confer protection. In this study, we used Nanochelating Technology to develop novel nanoadjuvants to boost the potency of the alum-adjuvanted inactivated severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) vaccine. BALB/c mice were immunized twice over 2 weeks with different doses of adjuvanted-vaccine formulations and immune responses were assessed. The analysis results of IFN-γ and IL-17 cytokines demonstrated the effectiveness of the nanoadjuvants produced by the Nanochelating Technology in shifting the alum-based vaccine toward a stronger Th1 pattern. In addition, these nanoadjuvants improved IL-2 cytokine response, which shows the efficacy of these novel formulations in inducing specific T lymphocyte proliferation. Using these nanoadjuvants increased IL-10 cytokine secretion that may be representative of a better immunoregulatory impact and may also potentially prevent immunopathology responses. Moreover, specific IgG titer analysis revealed the potency of these nanoadjuvants in improving humoral immune responses. The enzyme-linked immunosorbent assay of receptor-binding domain (RBD)-specific IgG response showed that the developed novel formulations induced strong IgG responses against this protein. This study shows that the nanostructures produced by the Advanced Nanochelating Technology have potent adjuvant effects on alum-based SARS-CoV-2 vaccines to not only compensate for alum weakness in inducing the cellular immune responses by smart regulation of the immune system but also significantly improve the humoral and cellular immune responses simultaneously.

Intranasal administration of sodium nitroprusside augments antigen-specific mucosal and systemic antibody production in mice

The coronavirus disease 2019, i.e., the COVID-19 pandemic, caused by a highly virulent and transmissible pathogen, has profoundly impacted global society. One approach to combat infectious diseases caused by pathogenic microbes is using mucosal vaccines, which can induce antigen-specific immune responses at both the mucosal and systemic sites. Despite its potential, the clinical implementation of mucosal vaccination is hampered by the lack of safe and effective mucosal adjuvants. Therefore, developing safe and effective mucosal adjuvants is essential for the fight against infectious diseases and the widespread clinical use of mucosal vaccines. In this study, we demonstrated the potent mucosal adjuvant effects of intranasal administration of sodium nitroprusside (SNP), a known nitric oxide (NO) donor, in mice. The results showed that intranasal administration of ovalbumin (OVA) in combination with SNP induced the production of OVA-specific immunoglobulin A in the mucosa and increased serum immunoglobulin G1 levels, indicating a T helper-2 (Th2)-type immune response. However, an analog of SNP, sodium ferrocyanide, which does not generate NO, failed to show any adjuvant effects, suggesting the critical role of NO generation in activating an immune response. In addition, SNPs facilitated the delivery of antigens to the lamina propria, where antigen-presenting cells are located, when co-administered with antigens, and also transiently elicited the expression of interleukin-6, interleukin-1β, granulocyte colony-stimulating factor, C-X-C motif chemokine ligand 1, and C-X-C motif chemokine ligand 2 in nasal tissue. These result suggest that SNP is a dual-functional formulation with antigen delivery capabilities to the lamina propria and the capacity to activate innate immunity. In summary, these results demonstrate the ability of SNP to induce immune responses via an antigen-specific Th2-type response, making it a promising candidate for further development as a mucosal vaccine formulation against infectious diseases.

Silver nanoparticles combat Salmonella Typhimurium: Suppressing intracellular infection and activating dendritic cells

Salmonella Typhimurium (ST) can hide inside cells, avoid antibiotic therapy and being killed by host’s immune system to cause persistent infection in humans and animals. Metal nanoparticles are regarded as an alternative to overcome the above limitations, silver nanoparticles especially have been applied in combating drug-resistant bacteria. However, the therapeutic effects of silver nanoparticles against intracellular infection and their impacts on host immunity remain an area of further investigation. In this work, we synthesized Ganoderma extract-capped silver nanoparticles (Ag@Ge) and explored the therapeutic potential and immune adjuvant effects of Ag@Ge against intracellular ST. Firstly, Ag@Ge had a small particle size of 35.52±7.46 nm, good stability, and biocompatibility. Then, Ag@Ge effectively entered RAW 264.7 cells, suppressed intracellular ST infection. Furthermore, Ag@Ge activated mouse dendritic cells (DCs) in vitro, evidenced by increased phenotypic markers (CD80/CD86/CD40/major compatibility complex II (MHCII)) expression and cytokine and chemokine (interleukin-6 (IL-6), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), chemokine (C-C motif) ligand 2 (CCL-2), and chemokine (C-C motif) receptor-7 (CCR-7)) transcription. More notably, the combination of Ag@Ge with inactivated ST recruited intestinal DCs to mitigate ST infection in mice, evidenced by decreased body weight loss and bacterial loads in the tissues (liver, jejunum, and colon), and improved platelets count. The above findings indicate that Ag@Ge has the potential as an alternative nano-antibiotic against intracellular ST infection.

Lactobacillus rhamnosus GG cell-free supernatant as a novel anti-cancer adjuvant

BackgroundGut microbiota modulation has been demonstrated to be effective in protecting patients against detrimental effects of anti-cancer therapies, as well as to improve the efficacy of certain anti-cancer treatments. Among the most characterized probiotics, Lactobacillus rhamnosus GG (LGG) is currently utilized in clinics to alleviate diarrhea, mucositis or intestinal damage which might be associated with several triggers, including Clostridium difficile infections, inflammatory gut diseases, antibiotic consumption, chemotherapy or radiation therapy. Here, we investigate whether LGG cell-free supernatant (LGG-SN) might exert anti-proliferative activity toward colon cancer and metastatic melanoma cells. Moreover, we assess the potential adjuvant effect of LGG-SN in combination with anti-cancer drugs.MethodsLGG-SN alone or in combination with either 5-Fuorouracil and Irinotecan was used to treat human colon and human melanoma cancer cell lines. Dimethylimidazol-diphenyl tetrazolium bromide assay was employed to detect cellular viability. Trypan blue staining, anti-cleaved caspase-3 and anti-total versus anti-cleaved PARP western blots, and annexin V/propidium iodide flow cytometry analyses were used to assess cell death. Flow cytometry measurement of cellular DNA content (with propidium iodide staining) together with qPCR analysis of cyclins expression were used to assess cell cycle.ResultsWe demonstrate that LGG-SN is able to selectively reduce the viability of cancer cells in a concentration-dependent way. While LGG-SN does not exert any anti-proliferative activity on control fibroblasts. In cancer cells, the reduction in viability is not associated with apoptosis induction, but with a mitotic arrest in the G2/M phase of cell cycle. Additionally, LGG-SN sensitizes cancer cells to both 5-Fluorouracil and Irinotecan, thereby showing a positive synergistic action.ConclusionOverall, our results suggest that LGG-SN may contain one or more bioactive molecules with anti-cancer activity which sensitize cancer cells to chemotherapeutic drugs. Thus, LGG could be proposed as an ideal candidate for ground-breaking integrated approaches to be employed in oncology, to reduce chemotherapy-related side effects and overcome resistance or relapse issues, thus ameliorating the therapeutic response in cancer patients.

A Pilot Randomized Controlled Trial Evaluating Essential Oils for Chemotherapy-Induced Peripheral Neuropathy

BackgroundChemotherapy-induced peripheral neuropathy (CIPN) is a painful, debilitating consequence of cancer treatment affecting up to 60% of patients. Pharmacological approaches to CIPN are often ineffective and cause adverse effects. Essential oils are an underutilized non-pharmacological approach to pain reduction. AimsTo ascertain the efficacy of an essential oil intervention to reduce CIPN. DesignA single-blind, pilot randomized controlled trial. MethodsParticipants (n = 27) were stratified by baseline pain scores and randomized to intervention (n = 13) and placebo groups (n = 14). Participants topically-applied the essential oil intervention or placebo every eight hours for six weeks. Pain was assessed using the Short-Form-McGill Pain Questionnaire-2 weekly and the Visual Analogue Scale daily. Quality-of-life was assessed using the Quality-of-Life: CIPN-20 and Quality-of-Life Adult Cancer Survivor questionnaires. Data were analyzed in SPSS using generalized estimating equations. ResultsNo significant difference was observed between groups in pain or quality-of-life scores over seven weeks, but improvement was observed in both groups. Participants using the intervention with pain medications showed a significant reduction in pain compared to placebo (p = .001). Educational level (p = .041) and annual income (p = .005) were significant covariates mirroring these social determinates of pain. Older participants felt less negatively about their CIPN (p = .002). Positive placebo effect and spatiotemporal interactions were observed. ConclusionsThis pilot study demonstrated that participants adhered to the intervention for six weeks. Essential oils have potential direct and adjuvant pain-reducing effects and should be studied further.

Impact of Epichaperome Inhibitor PU-H71 on Anti-Tumor T Cell Responses and Its Implications for Immune and Cellular Therapy

The epichaperome is a tightly integrated network of proteins supporting critical roles in the tumor-associated microenvironment and cancer cell survival. The epichaperome inhibitor PU-H71 is under development in clinical trials. A patient with an unclassified myeloproliferative neoplasm harboring a novel PML-SYK fusion, who progressed to acute myeloid leukemia despite allogeneic stem cell transplantation was successfully treated using PU-H71 (Sugita et al. 2021, NPJ Precis Oncol). We reported changes in donor immune cells after PU-H71 treatment, suggesting that epichaperome inhibition was modulating antitumor immune responses and improving the efficacy of cellular immunotherapy. To investigate this hypothesis, we analyzed the effects of PU-H71 on antitumor immune responses, GvHD and CAR-T cells. We tested the effects of PU-H71 on C57BL/6 mice immune responses to ovalbumine (OVA) immunization. OVA-specific T- and B-cell responses were evaluated by ELISPOT and ELISA, respectively. When administered at the time of priming, PU-H71 significantly increased the number of IFNγ-secreting T cells as compared to controls (median 126 vs 42 per million of cells, p=0.0002; n=8), without an impact on T cell phenotype or anti-OVA antibody levels. In contrast, PU-H71 had no effect when administered once the OVA-specific T cell response was ongoing. Then, we tested the effect of PU-H71 on antitumor immune responses in OVA-primed mice challenged with E.G7 (OVA+) or parental EL4 lymphoma cell lines (OVAneg). PU-H71 improved in vivo anti-tumor immune responses in OVA-primed mice challenged with E.G7 but not EL4 cells (Mean survival 17.5 vs 14 days, p=0.0003, n=8 per group). Since PU-H71 augmented early but not established T cell responses, we next evaluated whether it would have an effect on a GvHD murine model. Lethally-irradiated B6C3F1 mice received bone marrow and splenocytes from C57BL/6 mice and were treated with PU-H71 or vehicle at the first signs of GvHD (D11). GvHD clinical scores and histological analysis (colon, small intestine, liver and skin) were assessed. PU-H71 did not aggravate GvHD at both clinical and histological levels. Therefore, PU-H71 augments antigen-specific mouse T cell responses by acting during priming. To better understand its effect on human T cells and its potential adjuvant effect during cellular therapy, we next evaluated in vitro and in vivo CAR-T cells phenotype and function in the presence of PU-H71. T cells were purified from healthy donor PBMCs and were transduced with a CD19.4-1Bb.3z lentiviral vector to obtain CART19 cells. First, we found that epichaperome indeed forms in CART19 cells upon activation with anti-CD3/CD28 beads vs control. In vitro cytotoxicity of CAR-T cells toward NALM6 B-ALL cells was significantly improved (55.39 vs 74.66%, p=0.0356) by PU-H71. Finally, the therapeutic effect of the combination of PU-H71 and CART19 was evaluated in a B-ALL NSG mouse model using NALM6-BLIV cells, with bioluminescence imaging and flow cytometry (CART19 and leukemic cells) monitoring. Results suggest that PU-H71 administration increases CART19 numbers in the bone marrow and improves their in vivo antitumor efficacy against NALM6. Together, our results show that PU-H71 augments antigen-specific T cell responses during priming including CAR-T cell cytotoxicity, with little effect on ongoing immune responses including GvHD. Our data suggest that PU-H71 improves the in vivo efficiency of CAR-T cells in a B-ALL mouse model. In addition to its previously demonstrated direct anti-tumor effect, PU-H71 also had an adjuvant effect on anti-tumor T cell responses while not aggravating GvHD, which supports its clinical evaluation in the post-transplant setting or in combination with immune therapies like CAR-T cells.

A novel built-in adjuvant metallothionein-3 aids protein antigens to induce rapid, robust, and durable immune responses.

Adjuvants are crucial components of vaccines that can enhance and modulate antigen-specific immune responses. Herein, we reported for the first time that human metallothionein-3 (MT3), a low molecular weight cysteine-rich metal-binding protein, was a novel promising adjuvant candidate that could help protein antigens to induce rapid, effective, and durable antigen-specific immune responses. In the present study, MT3 was fused to outer membrane protein 19 (Omp19) of Brucella abortus (MT3-Omp19, MO) and C fragment heavy chain (Hc) of tetanus neurotoxin (MT3-Hc, MH), respectively. The results showed that MT3 as a built-in adjuvant increased the Omp19- or Hc-specific antibody responses by 100-1000 folds in seven days after primary immunization. Compared to other commercially available adjuvants, MT3 could stimulate earlier (4 days after primary injection) and stronger (10-100 folds) antibody response with lower antigen dose, and its adjuvanticity relied on fusion to antigen. Although the mechanism was not clear yet, the fusion protein MO was observed to directly activate DCs, promote germinal center formation and improve the speed of Ig class switching. Interestingly, our subsequent study found that other members of the mammalian MT family (human MT1 or murine MT3 for examples) also had potential adjuvant effects, but their effects were lower than human MT3. Overall, this study explored a new function of human MT3 as a novel built-in adjuvant, which may have important clinical application potential in vaccine development against global pandemics.

Thymol as an Adjuvant to Restore Antibiotic Efficacy and Reduce Antimicrobial Resistance and Virulence Gene Expression in Enterotoxigenic Escherichia coli Strains.

The continuous spread of antimicrobial resistance is endangering the efficient control of enterotoxigenic Escherichia coli (ETEC), which is mainly responsible for post-weaning diarrhea onset in piglets. Thymol, the key constituent of thyme essential oil, is already used in animal nutrition for its antimicrobial action. The aim of this study was to investigate the potential adjuvant effect of thymol to re-establish antibiotic efficacy against highly resistant ETEC field strains. Secondly, we evaluated the modulation of virulence and antibiotic resistance genes. Thymol showed the capacity to control ETEC growth and, when combined with ineffective antibiotics, it increased their antimicrobial power. In particular, it showed significant effects when blended with colistin and tetracycline, suggesting that the adjuvant effects rely on the presence of complementary mechanisms of action between molecules, or the absence of resistance mechanisms that inactivate antibiotics and target sites. Furthermore, our findings demonstrate that, when added to antibiotics, thymol can help to further downregulate several virulence and antibiotic resistance genes, offering new insights on the potential mechanisms of action. Therefore, in a one-health approach, our study supports the beneficial effects of combining thymol with antibiotics to restore their efficacy, together with the possibility of targeting gene expression as a pioneering approach to manage ETEC pathogenicity.

Ultrasound-Assisted Extraction of Anthocyanins from Malus 'Royalty' Fruits: Optimization, Separation, and Antitumor Activity.

Red Malus ‘Royalty’ fruits are rich in anthocyanins. This study aimed to obtain the optimal parameters for the extraction and separation of anthocyanins from Malus ‘Royalty’ fruits and to evaluate the inhibitory effect of the enriched anthocyanin fraction on gastric cancer cells. Ultrasonic-assisted extraction was used for the extraction of the anthocyanins of Malus ‘Royalty’ fruit, and the extraction results showed that the optimum parameters were an extraction temperature of 20 °C, a solid–liquid ratio of 1:6 (g/mL), ethanol and formic acid contents of 70% and 0.4%, respectively, an extraction time of 40 min, and an ultrasonic power of 300 W. The optimum extraction parameters to achieve the highest anthocyanin yield by a single-factor experiment coupled with response surface methodology were identified. The separation results showed that the AB-8 macroporous resin was a better purifying material, with 60% ethanol as an adsorbent, and the adsorption–desorption equilibrium times were 6 h and 1 h, respectively. Cyanidin-3-galactoside was the main body composition separation of anthocyanins by a high-performance liquid chromatography-diode array detector. The antitumor activity results showed that the anthocyanins of Malus ‘Royalty’ fruits have a significant inhibitory effect on the gastric cancer cell line BGC-803. The in vitro cell viability test of CCK-8 showed that the inhibitory effect on tumor cells was more significant with the increased anthocyanin concentration, with a half maximal inhibitory concentration (IC50) value of 105.5 μg/mL. The cell morphology was observed by an inverted microscope, and it was found that the backbone of BGC-803 treated with a high concentration of anthocyanins was disintegrated and the nucleoplasm was concentrated. The mechanism of apoptosis was analyzed by Western blotting, and the results showed that with increasing anthocyanin concentration in the medium, the expression levels of the proapoptotic proteins Bax and Bak increased, and the expression levels of the antiapoptotic proteins Bcl-2 and Bcl-xL decreased, which coordinated the regulation of cell apoptosis. This research suggests that the enriched anthocyanin fraction from Malus ‘Royalty’ fruits have potential antitumor and adjuvant therapeutic effects on gastric cancer.

Human adenovirus type 26 basic biology and its usage as vaccine vector.

Due to their nature, adenoviruses have been recognised as promising candidates for vaccine vector development. Since they mimic natural infection, recombinant adenovirus vectors have been proven as ideal shuttles to deliver foreign transgenes aiming at inducing both humoral and cellular immune response. In addition, a potent adjuvant effect can be exerted due to the adenovirus inherent stimulation of various elements of innate and adaptive immunity. Due to its low seroprevalence in humans as well as induction of favourable immune response to inserted transgene, human adenovirus type 26 (HAdV-D26) has been recognised as a promising platform for vaccine vector development and is studied in number of completed or ongoing clinical studies. Very recently HAdV-D26 based Ebola and Covid-19 vaccines were approved for medical use. In this review, current state of the art regarding HAdV-D26 basic biology and itsusage as vaccine vector will be discussed.

Potent adjuvant effect elicited for tumor immunotherapy by a liposome conjugated pH-sensitive polymer and dendritic cell-targeting Toll-like-receptor ligand

The generation of DCs with augmented functions is a strategy for obtaining satisfactory clinical outcomes in tumor immunotherapy. We developed a novel synthetic adjuvant comprising a liposome conjugated with a DC-targeting Toll-like-receptor ligand and a pH-sensitive polymer for augmenting cross-presentation. In an in vitro study using mouse DCs, these liposomes were selectively incorporated into DCs, significantly enhanced DC function and activated immune responses to present an epitope of the incorporated antigen on the major histocompatibility complex class I molecules. Immunization of mice with liposomes encapsulating a tumor antigen significantly enhanced antigen-specific cytotoxicity. In tumor-bearing mice, vaccination with liposomes encapsulating a tumor antigen elicited complete tumor remission. Furthermore, vaccination significantly enhanced cytotoxicity, targeting not only the vaccinated antigen but also the other antigens of the tumor cell. These results indicate that liposomes are an ideal adjuvant to develop DCs with considerably high potential to elicit antigen-specific immune responses; they are a promising tool for cancer therapy with neoantigen vaccination.

Cell-Selective Encapsulation within Metal-Organic Framework Shells via Precursor-Functionalized Aptamer Identification for Whole-Cell Cancer Vaccine.

Single-cell encapsulation is an emerging technology to endow cells with various functions, of which developing new applications in vivo is in high demand. Currently, metal-organic frameworks (MOFs) that are used as nanometric shells to coat living cells, however, have not realized cell-selective encapsulation. Here, a biocompatible and selective cell encapsulation strategy based on precursor-functionalized nucleolin aptamer and in situ MOF mineralization on the aptamer-identified cancer cell surface are developed. After MOF coating, the encapsulated cancer cells undergo immunogenic cell death, which is found associated with the changed cell stiffness (indicated by Young's modulus). The immunogenic dead cancer cells are used as whole-cell cancer vaccines (WCCVs), forming the integral WCCV-in-shell structure with enhanced immunogenicity ascribing from the surface-exposed calreticulin to promote dendritic cell recruitment, antigen presentation, and T-cell activation. The major activation pathways in the immune response are identified including tumor necrosis factor signaling pathway, cytokine-cytokine receptor interaction, and Toll-like receptor signaling pathway, suggesting the potential adjuvant effect of the MOF shells. After vaccination, WCCV-in-shell shows much better tumor immunoprophylaxis than either the imperfectly coated cancer cells or the traditional WCCV. This strategy is promising for the universal and facile development of novel whole-cell vaccines.

Cry1Ac Protoxin Confers Antitumor Adjuvant Effect in a Triple-Negative Breast Cancer Mouse Model by Improving Tumor Immunity.

The Cry1Ac protoxin from Bacillus thuringiensis is a systemic and mucosal adjuvant, able to confer protective immunity in different infection murine models and induce both Th1 and TCD8+ cytotoxic lymphocyte responses, which are required to induce antitumor immunity. The Cry1Ac toxin, despite having not being characterized as an adjuvant, has also proved to be immunogenic and able to activate macrophages. Here, we investigated the potential antitumor adjuvant effect conferred by the Cry1Ac protoxin and Cry1Ac toxin in a triple negative breast cancer (TNBC) murine model. First, we evaluated the ability of Cry1Ac proteins to improve dendritic cell (DC) activation and cellular response through intraperitoneal (i.p.) coadministration with the 4T1 cellular lysate. Mice coadministered with the Cry1Ac protoxin showed an increase in the number and activation of CD11c+MHCII- and CD11c+MHCII+low in the peritoneal cavity and an increase in DC activation (CD11c+MHCII+) in the spleen. Cry1Ac protoxin increased the proliferation of TCD4+ and TCD8+ lymphocytes in the spleen and mesenteric lymph nodes (MLN), while the Cry1Ac toxin only increased the proliferation of TCD4+ and TCD8+ in the MLN. Remarkably, when tested in the in vivo TNBC mouse model, prophylactic immunizations with 4T1 lysates plus the Cry1Ac protoxin protected mice from developing tumors. The antitumor effect conferred by the Cry1Ac protoxin also increased specific cytotoxic T cell responses, and prevented the typical tumor-related decrease of T cells (TCD3+ and TCD4+) as well the increase of myeloid-derived suppressor cells (MDSC) in spleen. Also in the tumor microenvironment of mice coadministered twice with Cry1Ac protoxin immunological improvements were found such as reductions in immunosupressive populations (T regulatory lymphocytes and MDSC) along with increases in macrophages upregulating CD86. These results show a differential antitumor adjuvant capability of Cry1Ac proteins, highlighting the ability of Cry1Ac protoxin to enhance local and systemic tumor immunity in TNBC. Finally, using a therapeutic approach, we evaluated the coadministration of Cry1Ac protoxin with doxorubicin. A significant reduction in tumor volume and lung metastasis was found, with increased intratumoral levels of tumor necrosis factor-α and IL-6 with respect to the vehicle group, further supporting its antitumor applicability.

Analgesic and adjuvant properties of exercise with vaccinations in healthy young population

ABSTRACT Introduction: Exercise holds the potential to be beneficial if used during vaccination processes by 1)exercise-induced analgesia to reduce pain associated with vaccination, 2)immune-enhancing effects, improving antibody responses to the vaccine, and 3)reducing local and systemic adverse reactions to the vaccine. This study examines whether analgesic responses could be enhanced locally in the exercising limb to further benefit the use of exercise during influenza vaccination processes to minimize vaccine-related pain and improve antibody response to inactivated influenza vaccines. Methods: 57 participants (22.6 ± 3.2 years, 33 females) randomized into a control (n = 19) or one of two exercise groups: pre-vaccine arm (n = 19) or pre-vaccine leg (n = 19). Intervention groups performed exercise (15 minutes), prior to administration of the vaccine. Vaccine-related pain and pressure pain threshold (PPT) were measured at baseline and post-vaccination for all groups. Blood samples were taken on the day of vaccination and one month later to measure serum antibody titers to influenza. Results: No significant difference in vaccine-related pain or change in PPT was found with exercise, however, there was a trend in higher reports of vaccine-related pain in females compared to males(p = .06). Significantly higher fold increase (p = .02) of the B/Brisbane/60/2008 strain was found in the exercise group compared to the control group. Conclusion: The current study failed to observe an analgesic effect of exercise to improve vaccine-related pain in young adults. However, immune-enhancing effects in one of four strains suggest potential adjuvant effects of exercise. Importantly, the sex difference in pain sensitivity suggests the need for separate analysis, especially when examining pain perception. Australian New Zealand Clinical Trial Registry (ACTRN:12617000374369)