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Since the introduction of allergen immunotherapy (AIT) over 100 years ago, focus has been on standardization of allergen extracts, with reliable molecular composition of allergens receiving the highest attention. While adjuvants play a major role in European AIT, they have been less well studied. In this Position Paper, we summarize current unmet needs of adjuvants in AIT citing current evidence. Four adjuvants are used in products marketed in Europe: aluminium hydroxide (Al(OH)(3)) is the most frequently used adjuvant, with microcrystalline tyrosine (MCT), monophosphoryl lipid A (MPLA) and calcium phosphate (CaP) used less frequently. Recent studies on humans, and using mouse models, have characterized in part the mechanisms of action of adjuvants on pre-existing immune responses. AIT differs from prophylactic vaccines that provoke immunity to infectious agents, as in allergy the patient is presensitized to the antigen. The intended mode of action of adjuvants is to simultaneously enhance the immunogenicity of the allergen, while precipitating the allergen at the injection site to reduce the risk of anaphylaxis. Contrasting immune effects are seen with different adjuvants. Aluminium hydroxide initially boosts Th2 responses, while the other adjuvants utilized in AIT redirect the Th2 immune response towards Th1 immunity. After varying lengths of time, each of the adjuvants supports tolerance. Further studies of the mechanisms of action of adjuvants may advise shorter treatment periods than the current three-to-five-year regimens, enhancing patient adherence. Improved lead compounds from the adjuvant pipeline are under development and are explored for their capacity to fill this unmet need.

Thymic epithelial tumors (TETs) are a heterogenous group of rare tumors, with a complex histopatological classification. Furthermore, the recent introduction of the first TNM staging system, that is scheduled to replace the Masaoka-Koga system, may create further difficulties in TET management, that remains challenging. Several guidelines for treatment of TETs are available and provide recommendations based mainly on non randomized trials and retrospective or limited series. Often the lack of evidence leads to formulation of indications based on expert opinions. As for other rare cancers it is crucial to create networks to coordinate the work among centres involved in treatment of these diseases in order to offer the best diagnostic and therapeutic tools. For this purpose, in 2014 a network named TYME (ThYmic MalignanciEs), was founded in Italy with the aim of improving care and research in TETs. In September 2017 a panel of multidisciplinary experts from TYME network and from other Italian centres strongly involved in TET diagnosis and treatment convened a first Italian Expert meeting together with representatives of association for patients affected by rare thoracic cancers Tu.To.R, to explore how these tumors are managed in the different centres of Italy compared to ESMO guidelines. In this paper we summarize the issues discussed during that meeting and we propose recommandations based on Masaoka Koga and the new TNM staging system.

Immunization is highly effective in preventing infectious diseases and therefore an indispensable public health measure. Allergic patients deserve access to the same publicly recommended immunizations as nonallergic patients unless risks associated with vaccination outweigh the gains. Whereas the number of reported possible allergic reactions to vaccines is high, confirmed vaccine-triggered allergic reactions are rare. Anaphylaxis following vaccination is rare, affecting less than 1/100,000, but can occur in any patient. Some patient groups, notably those with a previous allergic reaction to a vaccine or its components, are at heightened risk of allergic reaction and require special precautions. Allergic reactions, however, may occur in patients without known risk factors and cannot be predicted by currently available tools. Unwarranted fear and uncertainty can result in incomplete vaccination coverage for children and adults with or without allergy. In addition to concerns about an allergic reaction to the vaccine itself, there is fear that routine childhood immunization may promote the development of allergic sensitization and disease. Thus, although there is no evidence that routine childhood immunization increases the risk of allergy development, such risks need to be discussed. This article is protected by copyright. All rights reserved.

Chronic Hepatitis B virus (CHB) infection is a global public health problem. Oligodeoxynucleotides (ODNs) containing class C unmethylated cytosine-guanine dinucleotide (CpG-C) motifs may provide potential adjuvants for the immunotherapeutic strategy against CHB, since CpG-C ODNs stimulate both B cell and dendritic cell (DC) activation. However, the efficacy of CpG-C ODN as an anti-HBV vaccine adjuvant remains unclear. In this study, we demonstrated that CpG M362 (CpG-C ODN) as an adjuvant in anti-HBV vaccine (cHBV-vaccine) successfully and safely eliminated the virus in HBV-carrier mice. The cHBV-vaccine enhanced DC maturation both in vivo and in vitro, overcame immune tolerance, and recovered exhausted T cells in HBV-carrier mice. Furthermore, the cHBV-vaccine elicited robust hepatic HBV-specific CD8+ and CD4+ T cell responses, with increased cellular proliferation and IFN-γ secretion. Additionally, the cHBV-vaccine invoked a long-lasting follicular CXCR5+ CD8+ T cell response following HBV re-challenge. Taken together, CpG M362 in combination with rHBVvac cleared persistent HBV and achieved long-term virological control, making it a promising candidate for treating CHB.

Tumors of the intestinal tract are among the most common tumor diseases in humans, but, like many other tumor entities, show an unsatisfactory prognosis with a need for effective therapies. To test whether nutritional interventions and a combination with a targeted therapy can effectively cure these cancers, we used the fruit fly Drosophila as a model. In this system, we induced tumors by EGFR overexpression in intestinal stem cells. Limiting the amount of protein in the diet restored life span to that of control animals. In combination with a specific EGFR inhibitor, all major tumor-associated phenotypes could be rescued. This form of treatment was also successful in a real treatment scenario, which means when they started after the full tumor phenotype was expressed. In conclusion, reduced protein administration can be a very promising form of adjuvant cancer therapy.

“Immune normalization” has emerged as a new paradigm in immunotherapy, which is proposed in cancer patients instead of conventional “immune-enhancement” therapy. Immune normalization may also be implemented in cancer prevention of “sub-healthy” individuals. We established in vitro cultured mixed-natural killer (NKM) cells to achieve immune normalization. The in vitro cytotoxicity of NKM cells was tenfold higher than that of peripheral blood mononuclear cells (PBMCs). The cytotoxicity of NKM cells was negatively correlated with the proportion of T-helper cells (cluster of differentiation: CD3+CD4+ T), and positively correlated with the proportion of NK cells (especially CD56brightCD16bright NK cells). Then, we defined “sub-healthy individuals” after measuring Programmed cell death protein-1 (PD-1) expression in PBMCs from 95 donors aged > 50 years. Furthermore, we evaluated the potential clinical application of NKM-cell therapy in 11 patients with malignant lymphoma, one patient with pancreatic cancer, and four sub-healthy individuals. NKM-cell therapy elicited good tolerance and side-effects were not found. In sub-healthy individuals, the proportion of CD3+PD-1+ T cells and CD3+CD8+PD-1+ T cells was reduced significantly after NKM-cell treatment. We demonstrated that a new method using NKM cells was safe and efficacious as adjuvant treatment for cancer patients as well as therapy for sub-healthy individuals. Normalization of the peripheral immune system through NKM-cell therapy could expand its scope of application in different disorders.

Adjuvant concurrent chemoradiotherapy (CCRT) is the standard care for patients with resected advanced gastric cancer, but its survival benefits remain undetermined in patients undergoing D2 lymph node dissection (D2 dissection). We evaluated safety and efficacy of adjuvant CCRT with 5-fluorouracil (5-FU) versus chemotherapy alone in 110 gastric cancer patients with D2 dissection treated in Taiwan between January 2009 and January 2013. All the 71 patients receiving adjuvant CCRT were treated with daily infusional 5-FU and radiotherapy. Adjuvant CCRT was associated with higher risks of major hematologic (56.3% vs. 23.8%, p = 0.002) and gastrointestinal (46.9% vs. 14.3%, p = 0.027) toxicities and death (12.5% vs. 0.0%, p = 0.041) in patients above 70 years old, but this was not the case in those ≤70 years of age. Univariate Cox proportional regressions identified adjuvant CCRT as a factor for better overall survival (OS) (hazard ratio [HR]=0.52; 95% confidence interval [CI]: 0.27–0.99) and disease-free survival (DFS) (HR=0.46, 95% CI: 0.24–0.88), but it was not a significant factor for OS or DFS after adjusting for other factors in the multivariate analysis. However, in stratified analyses by age, we found adjuvant CCRT was an independent prognostic factor for better OS (HR=0.07; 95% CI: 0.01–0.38) in patients ≤70 years old, but not in those above 70 years of age. Therefore, it was concluded that age may to be a modifier of the effects of adjuvant CCRT.

Purpose Cutaneous B-cell lymphomas (CBCLs) are a rare group of diseases. External beam radiation therapy is recommended to treat CBCLs in all subtypes for locally advanced cases. However, there are no reports on high-dose-rate brachytherapy (HDR-BT) exclusively dedicated to CBCLs. The purpose of this paper was to report the first case series of CBCLs treated with HDR-BT. Material and methods Seven patients were treated between 2011 and 2019, with 12 skin lesions histopathologically proven as CBCLs. There were four T1a and eight T2a lesions. HDR-BT was prescribed as the first-line treatment for all cases, as the second-line treatment for recurrences after surgical failure for 4 patients, and as an adjuvant treatment for 1 case. The median total dose was 36 Gy (range, 30-40 Gy) in 10 fractions (range, 6-10 fractions), with a median overall treatment time of 11 days (range, 4-11 days). Treatment toxicity was assessed accordingly to the RTOG scale. Results The mean follow-up was 41 months. Local control was 100%. The rates of early toxicity were as follows: erythema (G1) - 33%, patchy epidermal desquamation (G2) - 25%, confluent epidermal desquamation (G3) - 25%, and minor bleeding (G4) - 17%. The reported rates of late toxicity included slight depigmentation (G1) - 59%, small telangiectasia (G2) - 8%, massive telangiectasia (G3) - 25%, and small ulceration (G4) in one site irradiated interstitially (8%). Conclusions HDR-BT allows for achieving high local control of CBCLs with relatively low-late toxicity in the form of skin discoloration in most patients.

Dengue virus (DENV) is a mosquito-borne virus that poses an incomparable public health problem, particularly in tropical and subtropical areas. Vaccination remains the most rational measure for controlling DENV infection. In this study, an ultraviolet irradiation (UV)-inactivated DENV-2 carried by N,N,N-trimethyl chitosan nanoparticles (UV-inactivated DENV2 TMC NPs) was investigated as a potential non-replicating dengue vaccine candidate. Using a human ex vivo model, the human monocyte-derived dendritic cells (MoDCs), we showed that TMC served as both a vaccine vehicle and a potent adjuvant. TMC NPs not only efficiently enhanced UV-inactivated DENV2 internalization into MoDCs but also greatly increased the breadth of UV-inactivated DENV2 immunogenicity to drive the maturation of MoDCs. Moreover, UV-inactivated DENV2 TMC NPs were highly immunogenic in mice, inducing greater levels of antibodies (total IgG, IgG1, IgG2a and neutralizing antibodies) and T cells (activated CD4⁺ and CD8⁺ T cells) against DENV-2 compared to soluble DENV-2 immunogens. Notably, the neutralizing activity of sera from mice immunized with UV-inactivated DENV2 TMC NPs was significantly augmented in the presence of complement activation, leading to the strong elimination of both DENV-2 particles and infected cells. We further showed that the immunogenicity of an inactivated dengue-based vaccine was significantly improved in a concentration-dependent manner. These positive results warrant further investigations of this platform of vaccine delivery for tetravalent vaccines or monovalent vaccines in sequential immunizations.

Objective: The study was designed to investigate some plasma markers which help us to decide the use of adjuvant corticosteroid therapy in bronchopulmonary dysplasia (BPD) of premature infants. Methods: Thirty BPD infants were treated by dexamethasone. Among these cases, dexamethasone was significant effective in 10 cases, and no significant effective in 20 cases. These patients were divided into two groups as the significant effect (SE) group (n=10) and the non-significant effect (NE) group (n=20) according to the curative effect of dexamethasone. Fifteen non-BPD infants with gestational age and gender matching were selected as the control group. Plasma samples before and after dexamethasone treatment were collected from three infants chosen randomly from SEG for the data-independent acquisition (DIA) analysis. ELISA was further used to detect the levels of differential proteins LRP1 and S100A8 in all individuals, including SE, NE and control groups. Results: DIA analysis results showed that after dexamethasone treatment, there were a total of 52 plasma proteins that showed significant differences, of which 43 proteins were down-regulated and 9 proteins were up-regulated. LRP1 and S100A8 were two plasma proteins that were significantly changed after dexamethasone treatment. Compared with the control group, plasma LRP1 was significantly increased in BPD. Interestingly, the plasma concentration of LRP1 in the NE group was significantly higher than that in the SE group. S100A8, as an indicator of plasma inflammation, was significantly higher in BPD than the control group. Unlike LRP1, there was no significantly difference between the SE and NE group (P=0.279) before dexamethasone treatment. Conclusion: Elevated plasma LRP1 and S100A8 in BPD infants are two indicators that correlated with the efficacy of dexamethasone, and might be used as biomarkers for deciding the use of adjuvant corticosteroids therapy in the BPD.

Flavin adenine dinucleotide (FAD) is engaged in several metabolic diseases. Its main role is being a cofactor essential for the activity of many flavoproteins, which play a crucial role in electron transport pathways in living systems. The aim of this study was to apply a pegylated flavins formulation named FAD-PEG diacide complex as theranostic pathway in cancer therapy. For this purpose, a mouse liver cancer model induced by Hepa1-6 cells was used to evaluate the therapeutic efficacy of FAD (named NP1) and FAD-PEG diacide complex (named NP2). The cytokines were applied to screen the serum inflammatory factors, to establish the blood cell content of different groups of nude mice. The highlights follows that FAD formulations (NP1; NP2) significantly suppressed the tumor growth and reduced the tumor index without effects on the body weight of mice. Furthermore, NP2 significantly reduced the serum levels of cytokines IL-6, TNF-α and IL-12 (P70). The reported results provide the proof-of-concept for the synthesis of a smart adjuvant for liver cancer therapy and support their further development in the field of nanomedicine.

Mycobacterium avium subsp. paratuberculosis (MAP) is the etiological agent of Paratuberculosis, a contagious, untreatable, and chronic granulomatous enteritis that results in diarrhea, emaciation, and death in farmed ruminants (i.e., cattle, sheep, and goats). In this study, the Ag85B antigen from MAP was expressed in transgenic alfalfa as an attractive vaccine candidate. Agrobacterium-mediated transformation allowed the rescue of 56 putative transformed plants and transgenesis was confirmed in 19 lines by detection of the Ag85B gene (MAP1609c) by PCR. Line number 20 showed the highest Ag85B expression [840 ng Ag85B per gram of dry weight leaf tissue, 0.062% Total Soluble Protein (TSP)]. Antigenicity of the plant-made Ag85B was evidenced by its reactivity with a panel of sera from naturally MAP-infected animals, whereas immunogenicity was assessed in mice immunized by either oral or subcutaneous routes. The plant-made Ag85B antigen elicited humoral responses by the oral route when co-administered with cholera toxin as adjuvant; significant levels of anti-85B antibodies were induced in serum (IgG) and feces (IgA). Long-lasting immunity was evidenced at day 180 days post-first oral immunization. The obtained alfalfa lines expressing Ag85B constitute the first model of a plant-based vaccine targeting MAP. The initial immunogenicity assessment conducted in this study opens the path for a detailed characterization of the properties of this vaccine candidate. Supplementary Information The online version contains supplementary material available at 10.1007/s12033-021-00307-w.

Current human papillomavirus (HPV) vaccines provide substantial protection against the most common HPV types responsible for oral and anogenital cancers, but many circulating cancer-causing types remain for which vaccine coverage is lacking. In addition, all current HPV vaccines rely on aluminum salt-based adjuvant formulations that function through unclear mechanisms with few substitutes available. In an effort to expand the toolbox of available adjuvants suitable for HPV vaccines, we compared the immunogenicity of the RG1-VLP (virus-like particle) vaccine in BALB/c mice when formulated with either the aluminum hydroxide adjuvant Alhydrogel or the novel polyphosphazene macromolecular adjuvant poly[di (carboxylatoethylphenoxy) phosphazene] (PCEP). PCEP-formulated RG1-VLPs routinely outperformed VLP/Alhydrogel in several measurements of VLP-specific humoral immunity, including consistent improvements in the magnitude of antibody (Ab) responses to both HPV16-L1 and the L2 RG1 epitope as well as neutralizing titers to HPV16 and cross-neutralization of pseudovirion (PsV) types HPV18 and HPV39. Dose-sparing studies indicated that RG1-VLPs could be reduced in dose by 75% and the presence of PCEP ensured activity comparable to a full VLP dose adjuvanted by Alhydrogel. In addition, levels of HPV16-L1 and -L2-specific Abs were achieved after two vaccinations with PCEP as adjuvant that were equivalent to or greater than levels achieved with three vaccinations with Alhydrogel alone, indicating that the presence of PCEP resulted in accelerated immune responses that could allow for a decreased dose schedule. Given the extensive clinical track record of polyphosphazenes, these data suggest that substitution of alum-based adjuvants with PCEP for the RG1-VLP vaccine could lead to rapid seropositivity requiring fewer boosts, the dose-sparing of commercial VLP-based vaccines, and the establishment of longer-lasting humoral responses to HPV.

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).

Introduction: Large and consistent evidence supports the use of eribulin mesylate in clinical practice in third or later line treatment of metastatic triple negative breast cancer (mTNBC). Conversely, there is paucity of data on eribulin efficacy in second line treatment. Methods: We investigated outcomes of 44 mTNBC patients treated from 2013 through 2019 with second line eribulin mesylate in a multicentre retrospective study involving 14 Italian oncologic centres. Results: Median age was 51 years, with 11.4% of these patients being metastatic at diagnosis. Median overall survival (OS) and progression free survival (PFS) from eribulin starting were 11.9 (95%CI: 8.4-15.5) and 3.5 months (95%CI: 1.7-5.3), respectively. We observed 8 (18.2%) partial responses and 10 (22.7%) patients had stable disease as best response. A longer PFS on previous first line treatment predicted a better OS (HR=0.87, 95%CI: 0.77-0.99, p= 0.038) and a longer PFS on eribulin treatment (HR=0.92, 95%CI: 0.85-0.98, p=0.018). Progression free survival to eribulin was also favorably influenced by prior adjuvant chemotherapy (HR=0.44, 95%CI: 0.22-0.88, p=0.02). Eribulin was generally well tolerated, with grade 3-4 adverse events being recorded in 15.9% of patients. Conclusions: The outcomes described for our cohort are consistent with those reported in the pivotal Study301 and subsequent observational studies. Further data from adequately-sized, ad hoc trials on eribulin use in second line for mTNBC are warranted to confirm our findings.

Objectives A potent HIV vaccine should overcome some limitations such as polymorphism of human HLA, the diversity of HIV-1 virus, and the lack of an effective delivery system. In this study, a DNA construct encoding Nef60–84, Nef126–144, Vpr34–47, Vpr60–75, Gp16030–53, Gp160308–323, and P248–151 epitopes was designed using bioinformatics tools. The pcDNA3.1-nef-vpr-gp160-p24 and pcDNA3.1-nef constructs were prepared in large scale as endotoxin-free form. Moreover, the recombinant Nef-Vpr-Gp160-p24 polypeptide and Nef protein were generated inE. coli. These constructs were delivered using cell penetrating peptides (CPPs) in vivo, and immune responses were assessed for different modalities in BALB/c mice. Results The recombinant DNA constructs were confirmed as the ~ 867 bp and ~ 648 bp bands related tonef-vpr-gp160-p24 andnef genes on agarose gel. Moreover, the purified Nef-Vpr-Gp160-p24 polypeptide and Nef protein showed the ~ 32 kDa and ~ 30 kDa bands on SDS-PAGE, respectively. The results of immune responses indicated that the heterologous prime/boost regimens using both Nef-Vpr-Gp160-P24 and Nef antigens induced significantly the secretion of IgG2a, IgG2b, IFN-γ and Granzyme B compared to other groups. The levels of Granzyme B in mice immunized with Nef antigen were higher than those immunized with Nef-Vpr-Gp160-P24 antigen. The CPPs showed the same potency with Montanide adjuvant for eliciting immune responses. Conclusions The heterologous prime/boost regimens for both antigens could significantly direct immune responses toward Th1 and CTL activity compared to other regimens. Comparing the efficiency of Nef-Vpr-Gp160-P24 and Nef constructs, the Nef-Vpr-Gp160-P24 constructs delivered by CPPs showed promising results as an HIV vaccine candidate.

Currently, the cancer immunotherapy has made great progress while antitumor vaccine attracts substantial attention. Still, the selection of adjuvants as well as antigens are always the most crucial issues for better vaccination. In this study, we proposed a biomimetic antitumor nanovaccine based on biocompatible nanocarriers and tumor cell membrane antigens. Briefly, endogenous calcium pyrophosphate nanogranules with possible immune potentiating effect are designed and engineered, both as delivery vehicles and adjuvants. Then, these nanocarriers are coated with lipids and B16-OVA tumor cell membranes, so the biomembrane proteins can serve as tumor-specific antigens. It was found that calcium pyrophosphate nanogranules themselves were compatible and possessed adjuvant effect, while membrane proteins including tumor associated antigen were transferred onto the nanocarriers. It was demonstrated that such a biomimetic nanovaccine could be well endocytosed by dendritic cells, promote their maturation and antigen-presentation, facilitate lymph retention, and trigger obvious immune response. It was confirmed that the biomimetic vaccine could induce strong T-cell response, exhibit excellent tumor therapy and prophylactic effects, and simultaneously possess nice biocompatibility. In general, the present investigation might provide insights for the further design and application of antitumor vaccines., Graphical abstract Image, graphical abstractA biomimetic calcium pyrophosphate nanovaccine coated with tumor cell membrane was constructed. After subcutaneous administration, the biomimetic nanovaccine could accumulate in the drainage lymph nodes and activate effective specific cytotoxic T cells, then exerting a strong anti-tumor effect.

Objectives: Atopic dermatitis (AD) is one of the most common skin disorders in infants and children and is often aggravated by increased Staphylococcus aureus (S. aureus) colonization. An inhibitory effect of a specific egg yolk antibody (IgY) on S. aureus growth was demonstrated in this study. Furthermore, the effects of water- or oil-based adjuvants on the preparation of anti-S. aureus IgY and hen immunization were compared. Methods: Hens were immunized intramuscularly with formalin-killed S. aureus mixed with either a water-soluble polysaccharide λ-carrageenan, oil-based Freund's complete adjuvant (FCA), or Freund's incomplete adjuvant (FIA). Anti-S. aureus IgYs (FIA-IgY, FCA/FIA-IgY, and λCarra-IgY) were purified from the egg yolk of immunized hen eggs, and the activity of the IgY against S. aureus antigen was measured by ELISA. The proportion of each IgY that was absorbed by S. aureus was also determined. Then, the effect of purified anti-S. aureus IgY on S. aureus growth inhibition was investigated in vitro. Results: The yolk of eggs and purified FIA-IgY from the FIA group showed the highest antibody activity, followed by FCA/FIA-IgY and λCarra-IgY. The proportion of each IgY that was absorbed by S. aureus antigen was as follows: FIA-IgY (18.1%), FCA/FIA-IgY (12.9%), and λCarra-IgY (7.0%). Only FIA-IgY significantly inhibited S. aureus growth in liquid medium. Conclusion: A specific IgY that was produced using the FIA adjutant inhibited S. aureus growth. Although water-soluble λ-carrageenan showed an adjuvant effect on anti-S. aureus IgY induction in egg yolk, but did not inhibit S. aureus growth. The use of the oil adjuvant FIA was necessary in the preparation of anti-S. aureus IgY as a treatment for AD symptoms.

Objectives: Despite the inclusion of extranodal extension (ENE) in the recent staging system, the presence of ENE alone is not sufficient to depict all clinical situations, as ENE is frequently found in multiple nodes. Thus, the purpose of this study was to evaluate the surgery-based treatment outcomes and clinicopathological features of oral cavity squamous cell carcinoma (OCSCC) patients with ENE found in bilateral multiple cervical metastases. Materials and methods: A retrospective single-institutional study of OCSCC patients with bilateral ENE nodes was performed from January 2011 to December 2018. OCSCC patients of different admission statuses (with primary lesions (PL), recurrent lesions (RL) and isolated neck metastases (INM)) were included for subgroup comparisons. All patients received surgical treatment with/without adjuvant therapies and had complete follow-up data. Disease-free survival (DFS) was regarded as the main outcome. Time-to-relapse data were also collected for comparison. Results: A total of 128 patients were included, of whom 97 (75.8%) were male. The mean follow-up period reached 15 months. Among the patients, 85 (66.4%) were treated for PLs, followed by 26 (20.3%) treated for RLs after failed prior therapy and 17 (13.3%) treated for INMs (concurrent or sequential). The DFS rate was merely 35.2%. Treatment-related factors such as surgical margin (p=0.003), postoperative adjuvant therapy (p=0.014) and perioperative complications (p=0.036) were significantly associated with patient outcomes. In addition, oral lesion-related variables such as oral subsites (p=0.037), T classification (p=0.026) and skull base involvement (p=0.040) were indicators of a worse prognosis. For bilateral ENE features, ENE subclassification (p=0.036), maximum size of ENE nodes (p=0.039) and arterial nodal encasement (p=0.025) tended to predict the surgery-based treatment outcomes of these patients. Conclusions: Bilateral cervical metastases with ENE features, though uncommon, are a serious regional burden, and these patients have lower-than-expected treatment outcomes, especially those with RLs or INMs. A fairly large number of OCSCC patients with advanced oral lesions gain little benefit from intensified salvage surgical treatment. Such treatment should instead be offered to select patients with smaller bilateral ENE nodes (

Dendritic cells (DCs) can process the antigens of cancer vaccine and thus stimulate the CD8+ T cells to recognize and kill the tumor cells that express these antigens. However, lack of promising carriers for presenting the antigens to DCs is one of the main barriers to the development of clinically effective cancer vaccines. Another limitation is the risk of inflammatory side effects induced by the adjuvants. It is still unclear how we can develop ideal adjuvant-free DC vaccine carriers without adjuvants. Methods: A 12-mer peptide carrier (CBP-12) with high affinity for Clec9a expressed on DCs was developed using an in silico rational optimization method. The therapeutic effects of the adjuvant-free vaccine comprising CBP-12 and exogenous or endogenous antigenic peptides were investigated in terms of antigen cross-presentation efficacy, specific cytotoxic T lymphocyte response, and antitumor activity. We also explored the mechanism involved in the antitumor effects of the adjuvant-free CBP-12 vaccine. Finally, we assessed the effects of the CBP-12 conjugated peptide vaccine combined with radiotherapy. Results: Here, we developed CBP-12 as a vaccine carrier that enhanced the uptake and cross-presentation of the antigens, thus inducing strong CD8+ T cell responses and antitumor effects in both anti-PD-1-responsive (B16-OVA) and -resistant (B16) models, even in adjuvant-free conditions. CBP-12 bound to and activated Clec9a, thereby stimulating Clec9a+ DC to product IL-21, but not IL-12 by activating of Syk. The antitumor effects of the CBP-12 conjugated peptide vaccines could be blocked by an IL-21 neutralizing antibody. We also observed the synergistic antitumor effects of the CBP-12 conjugated peptide vaccine combined with radiotherapy. Conclusions: CBP-12 could serve as an adjuvant-free peptide vaccine carrier for cancer immunotherapy.

Antibiotic resistance among gram-negative bacteria continues to rise globally at an alarming rate. New vaccines that prevent bacterial infections and reduce antibiotic use could provide a potential solution to these problems. This study focused on development of an investigational vaccine to prevent recurrent urinary traction infections (UTI) caused by gram-negative bacteria that use type 1 pili to adhere to, invade, and colonize human bladders. The vaccine antigen is FimH, an adhesin protein on the tip of type 1 pili with a lectin binding domain that enables attachment to glycoproteins on mammalian bladders. This was a phase 1, open-label, dose escalation study evaluating the vaccine in 67 healthy women with and without histories of recurrent UTI. The objectives of the study were to evaluate the safety, tolerability, and immunogenicity of different dosages of the antigen and adjuvant of the vaccine. All dosages were well-tolerated and a low incidence of systemic reactions occurred. No serious adverse events related to the vaccine were reported. The vaccine induced both binding and functional antibodies. The women with histories of recurrent UTI demonstrated greater than 150-fold increases in antibodies against the N-terminal region of FimH. Based on the results of this phase 1 study, this vaccine is proceeding to a double-blind, randomized, placebo-controlled phase 2 study. If this vaccine is successful in future studies, it could potentially prevent millions of recurrent UTI globally and reduce the development of antibiotic resistance.

Purpose To evaluate dosimetric differences between point-based 2-dimensional (2D) vaginal brachytherapy (VBT) treatment planning technique and volume-based 3-dimensional (3D) VBT method for endometrial cancer (EC). Material and methods Ten patients with uterine-confined EC treated with VBT were included in this study. All patients received 27.5 Gy in 5 fractions. Three different treatment plans were performed for each patient: plan A for dose prescribed to the entire vaginal wall thickness delineated via computed tomography guidance, plan B for dose prescribed to the vaginal mucosa/cylinder surface, and plan C for dose prescribed to 5 mm beyond the vaginal mucosa/cylinder surface. Dose-volume histograms (DVH) of treatment volumes and organs at risk (OARs) were evaluated and compared. Results DVH analysis of target volume doses (D100, D95, and D90) showed a significant difference between plan A and plan B (p = 0.005), and plan B was found lower. D100 for plan C was significantly higher than plan A (p = 0.009), but for D95 and D90, no statistically significant difference was found (p = 0.028 and p = 0.028, respectively). In terms of OARs doses, including vagina, rectum, bladder, and sigmoid, D2cm3 doses were significantly higher in plan A than plan B (p = 0.009, p = 0.009, p = 0.005, and p = 0.005, respectively). All these doses were also significantly lower than in plan C (p = 0.005, p = 0.012, and p = 0.013, respectively), except for sigmoid (p = 0.155). Conclusions In this dosimetric analysis, we have shown that the volume-based 3D VBT technique provides the ability to balance the target dose against the sparing of OARs. Therefore, in the new modern 3D treatment era, instead of normalization of the dose to standard reference points, customized 3D volume-based VBT planning should be recommended.

Background and purpose Temporomandibular disorders (TMD) and migraine can be co-morbid. This can be a significant factor in exacerbating and increasing the prevalence of migraine-like symptoms. However, the underlying mechanisms involved are unknown. Our objective was to investigate these neural mechanisms and the role of CGRP as a key modulator in this co-morbidity. Experimental approach We combined experimental approaches using CGRP, which triggers a migraine-like response in patients, with that of masseteric muscle injection of complete Freund's adjuvant (CFA), to model myofascial TMD-like inflammation. Using validated electrophysiological methods to assess each of the above approaches independently or in combination, we examined their effects on the response properties of migraine-like dural-trigeminocervical neurons. Key results Independently, in ~2/3 of animals (rats) each approach caused delayed migraine-like activation and sensitisation of dural-trigeminocervical neurons. The response to masseteric-CFA was attenuated by a selective CGRP receptor antagonist. The combination approach caused a migraine-like neuronal response in all animals tested, with somatosensory-evoked cranial hypersensitivity significantly exacerbated. Conclusion and implications The data demonstrate a neuronal phenotype that translates to the exacerbated clinical co-morbid phenotype, supporting this combination approach as a relevant model to study the mechanisms involved. It provides a pathophysiological rationale for this exacerbated phenotype, strongly implicating the involvement of CGRP. The results provide support for targeting the CGRP pathway as a novel monotherapy approach for treating this co-morbid condition. This has key implications into our understanding of this co-morbid condition, as well as potentially addressing the major unmet need for novel and effective therapeutic approaches.

Supramolecular peptide hydrogel has shown promising potential in vaccine development largely because of its ability to function both as antigen depot and immune adjuvant. Nap‐GdFdFdY, a tetrapeptide hydrogel that has been previously reported to exhibit adjuvant effect, is inadvertently found to contain conserved peptide sequence for insulin, proinsulin, and glutamic acid decarboxylase, 3 major autoantigens for the autoimmune type 1 diabetes (T1D). At present, despite being managed clinically with insulin replacement therapy, T1D remains a major health threat with rapidly increasing incidences, especially in children and young adults, and antigen‐specific immune tolerance induction has been proposed as a feasible approach to prevent or delay T1D progression at an early stage. Here, it is reported that innoculation of Nap‐GdFdFdY leads to complete protection of nonobese diabetic (NOD) mice from T1D development till the age of 36 weeks. Better maintenance of pancreatic islet morphology with minimal immune cell infiltration is also observed from mice exposed to Nap‐GdFdFdY. This beneficial impact is mainly due to its facilitative role on enhancing peripheral T regulatory cell (Treg) population, shown as increased splenic Treg percentage, and function, demonstrated by maintenance of circulating TGF‐β1 level. Serum cytokine microarray data further implicate a “buffering” role of Nap‐GdFdFdY on systemic inflammatory tone in NOD mice. Thus, with its versatility, applicability, and excellent potency, Nap‐GdFdFdY is posited as a novel therapeutic intervention for T1D., A hydrogel of Nap‐GdFdFdY containing conserved sequence of autoantigens including insulin, proinsulin, and glutamic acid decarboxylase as a novel therapeutic intervention for the autoimmune type 1 diabetes. Better pancreatic islet morphology with minimal immune cell infiltration is observed from mice with hydrogel because of enhancing peripheral T regulatory cell population and maintenance of circulating TGF‐β1 level.

Purpose Radiotherapy (RT), as part of trimodal therapy, is an attractive alternative treatment in patients with urothelial muscle-invasive bladder cancer (MIBC). There is accumulating evidence suggesting the immunomodulatory effects of RT and its potential synergy when combined with immunotherapy. The aim of this review was to report on the most recent advances on this combination, including the mechanisms of RT immunomodulation, practical approach to combining RT and immunotherapy, and ongoing clinical trials in bladder cancer. Methods Using the PubMed database, we identified articles published between March 2004 and April 2020 on the combination of RT with immunotherapy in localized or metastatic MIBC. A search of the Clinicaltrials.gov and Clinicaltrialsregister.eu/ retrieved ongoing clinical trials on the topic as well. Results Combination of RT with immunotherapy leads to immunogenic cell death and an increase in immune markers thus leading to improved tumor control. For localized MIBC, there are safety concerns related to the use of concurrent immunotherapy with hypofractionated RT, thus neoadjuvant or adjuvant immunotherapy is preferred. In the metastatic setting, the combination of multi-site RT with SBRT-like doses (≥ 6 Gy per fraction) and concurrent immunotherapy seems most efficacious at harnessing the abscopal effect. At least 25 clinical trials combining immunotherapy and RT in MIBC are currently ongoing and will answer pending questions on safety, efficacy, and practical considerations on RT scheduling, fractionation, and targets volumes. Conclusion RT has the potential to synergize with immunotherapy to improve oncological outcomes in patient with localized or metastatic MIBC. Clinical trials results are eagerly awaited.

Archaeosomes are liposomes formulated using total polar lipids (TPLs) or semi-synthetic glycolipids derived from archaea. Conventional archaeosomes with entrapped antigen exhibit robust adjuvant activity as demonstrated by increased antigen-specific humoral and cell-mediated responses and enhanced protective immunity in various murine infection and cancer models. However, antigen entrapment efficiency can vary greatly resulting in antigen loss during formulation and variable antigen:lipid ratios. In order to circumvent this, we recently developed an admixed archaeosome formulation composed of a single semi-synthetic archaeal lipid (SLA, sulfated lactosylarchaeol) which can induce similarly robust adjuvant activity as an encapsulated formulation. Herein, we evaluate and compare the mechanisms involved in the induction of early innate and antigen-specific responses by both admixed (Adm) and encapsulated (Enc) SLA archaeosomes. We demonstrate that both archaeosome formulations result in increased immune cell infiltration, enhanced antigen retention at injection site and increased antigen uptake by antigen-presenting cells and other immune cell types, including neutrophils and monocytes following intramuscular injection to mice using ovalbumin as a model antigen. In vitro studies demonstrate SLA in either formulation is preferentially taken up by macrophages. Although the encapsulated formulation was better able to induce antigen-specific CD8+ T cell activation by dendritic cells in vitro, both encapsulated and admixed formulations gave equivalently enhanced protection from tumor challenge when tested in vivo using a B16-OVA melanoma model. Despite some differences in the immunostimulatory profile relative to the SLA (Enc) formulation, SLA (Adm) induces strong in vivo immunogenicity and efficacy, while offering an ease of formulation.

Objectives We recently developed a novel peptide, AJP001, that possesses both a mouse T-cell epitope and adjuvant action. Direct conjugation to the antigen is useful for peptide vaccines without the addition of adjuvants. In this study, the efficacy of an angiotensin (Ang) II and AJP001-conjugated peptide vaccine (AJ-Ang II) was evaluated in mice. Methods The anti-Ang II antibody titer was measured in Balb/C mice following three injections of AJ-Ang II at 2-week intervals. SBP was measured during vaccination of Balb/C mice treated with Ang II infusion (1 μg/kg per min). Results AJ-Ang II treatment resulted in an increase in the anti-Ang II antibody titer in a dose-dependent manner without the addition of adjuvants. In the analysis of the humoral immune response, AJ-Ang II mainly elicited IgG1 antibodies and IL-4 and IL-10 production, as measured by an enzyme-linked immune absorbent spot assay, which suggests the induction of a Th2 response. Importantly, cotreatment with purified antibodies attenuated Ang II-induced extracellular signal-regulated kinase phosphorylation and nuclear factor (NF)-κB activation in cultured vascular smooth muscle cells. The SBP in immunized mice was significantly lower than that in nonimmunized mice (135.9 ± 8.5 vs. 154.9 ± 16.8 mmHg, P = 0.02). Furthermore, Ang II-induced perivascular fibrosis in the heart was significantly attenuated in immunized mice, which also exhibited decreased mRNA expression of collagen I/III and transforming growth factor-β. Conclusion AJ-Ang II may be a simple and useful therapeutic peptide vaccine without the addition of any adjuvants.

Traditional non-living vaccines are often least effective in the populations that need them most, such as neonates and elderly adults. Vaccine adjuvants are one approach to boost the immunogenicity of antigens in populations with reduced immunity. Ideally, vaccine adjuvants will increase the seroconversion rates across the population, lead to stronger immune responses, and enable the administration of fewer vaccine doses. We previously demonstrated that a cationic liposomal formulation of the commercial influenza split virus vaccine (CCS/C-HA) enhanced cellular and humoral immunity to the virus, increased seroconversion rates, and improved survival after live virus challenge in a preclinical model, as compared to the commercial vaccine as is (F-HA). We now evaluated vaccine efficacy in different strains and sexes of mice and determined the role of innate immunity in the mechanism of action of the CCS/C adjuvant by testing the response of mice deficient in Toll-like receptors or the TLR/IL-1 adaptor protein MyD88 following immunization with CCS/C-HA vs. F-HA. Although TLR2- and TLR4-deficient mice responded to F-HA immunization, F-HA immunization failed to engender a significant immune response in the absence of MyD88. In contrast, immunization with the CCS/C-HA vaccine overcame the requirement for MyD88 in the response to the commercial vaccine and improved the immune responses and seroconversion rates in all strains of mice tested, including those deficient in TLR2 and TLR4.

BACKGROUND AND PURPOSE: Intravenous thrombolysis (IVT) after stroke enhances C3a generation, which may abrogate the benefits of reperfusion. The C3aR antagonist SB290157 is neuroprotective following transient but not permanent middle cerebral artery occlusion (MCAo). SB290157 remains untested in thromboembolic (TE) models, which better approximate human stroke and also facilitate testing in combination with IVT. We hypothesized SB290157 would confer neuroprotection in TE stroke with and without “late” IVT. EXPERIMENTAL APPROACH: We used two different models of TE stroke to examine the efficacy of SB290157 alone and in combination with late IVT. We evaluated the benefit of SB290157 in attenuating post‐ischaemic behavioural deficits, infarction, brain oedema and haemorrhage. KEY RESULTS: Plasma C3a was elevated 6 hr after TE stroke alongside increased cerebrovascular C3aR expression, which was sustained to 4 weeks. Increased C3aR expression also was visualized in human ischaemic brain. In a photothrombotic (PT) stroke model, which exhibits rapid spontaneous reperfusion, SB290157 given at 1 hr post‐PT significantly improved neurofunction and reduced infarction at 48 hr. In an embolic (eMCAo) model, SB290157 administered at 2 hr improved histological and functional outcomes. Conversely, late IVT administered 4.5 hr post‐eMCAo was ineffective likely due to increased haemorrhage and brain oedema. However, SB290157 administered prior to late IVT ameliorated haemorrhage and oedema and improved outcomes. CONCLUSIONS AND IMPLICATIONS: We conclude that SB290157 is safe and effective with and without late IVT following TE stroke. Therefore, C3a receptor antagonist therapy represents a promising candidate for clinical translation in stroke, particularly as an adjuvant to IVT.

Introduction This study assesses the efficacy and tolerability of two cycles of adjuvant chemotherapy (AC) with gemcitabine and cisplatin after radical cystectomy in patients with a high risk of progression of muscle-invasive urothelial bladder cancer as compared to chemotherapy at relapse, in a prospective randomized study. Material and methods From 2008 to 2013, all patients after radical cystectomy at our institution for primary or recurrent urothelial bladder cancer with stage pT3-4 and/or pN+ on histopathology and without contraindications to combination cisplatin-based chemotherapy, were randomized either to two cycles of gemcitabine and cisplatin chemotherapy or to follow-up and chemotherapy at the time of relapse. The study endpoints were overall, cancer-specific, and disease-free survival. Results The study included 100 patients, of whom 53 received AC and the other 47 were assigned to the control arm. Out of 53 allocated to AC arm, 16 patients did not start chemotherapy or received only one cycle of AC. The median follow-up for patients in the AC and control arms was 88 and 86 months, respectively. In the AC arm the hazard ratio for death from any cause, death from bladder cancer, and disease relapse were 0.70 (95% CI 0.45-1.11; p = 0.13), 0.84 (95% CI 0.50-1.41; p = 0.51), and 0.77 (95% CI 0.46-1.28; p = 0.31), respectively. Conclusions Two cycles of AC with gemcitabine and cisplatin in patients with high-risk urothelial bladder cancer after radical cystectomy does not improve overall, cancer-specific, and disease-free survival. Only 53% of patients randomized to AC received the entire planned treatment.

More effective rotavirus vaccines are essential for preventing extensive diarrheal morbidity and mortality in children under five years of age in low-resource regions. Nonreplicating rotavirus vaccines (NRRV) administered parenterally provide an alternate vaccination method to the current licensed oral vaccine. Live attenuated vaccines and may generate increased efficacy in low-resource settings because the parenteral administration route bypasses some of the challenges associated with oral administration, including differences in intestinal environments. Work described here supports development of a trivalent NRRV vaccine for parenteral administration to avoid complications of the gastrointestinal route. Recombinant VP8* subunit proteins representing some of the most prevalent strains of rotavirus infecting humans – DS-1 (P[4]), 1076 (P[6]), and Wa (P[8]) – were combined with an aluminum adjuvant and the P2 epitope of tetanus toxoid to enhance the immune response to this NRRV antigen. Vaccine formulation development included selection of aluminum hydroxide (Alhydrogel®) as an appropriate adjuvant as well as an optimal buffer to maintain antigen stability and optimize antigen binding to the adjuvant. Characterization assays were used to select the lead vaccine formulation and monitor formulation stability. The NRRV liquid formulation was stable for one year at 2°C to 8°C and four weeks at 37°C. Immunogenicity of the NRRV formulation was evaluated using a guinea pig model, where we demonstrated that the adjuvant provided a 20-fold increase in neutralization titer against a homologous antigen and that the P2-fusion also enhanced the serum neutralizing antibody responses. This vaccine candidate is currently being evaluated in human clinical trials.

Background and Aims: The prognosis of intrahepatic cholangiocarcinoma (ICC) after radical resection is far from satisfactory, but the effect of postoperative transarterial chemoembolization (p-TACE) remains controversial. This multi-center retrospective study was to evaluate the clinical value of p-TACE and identify the selected patients who would benefit from p-TACE. Methods: Data of ICC patients who underwent radical resection with/without p-TACE therapy was obtained from 12 hepatobiliary centers in China between Jan 2014 and Jan 2017. Overall survival (OS) was set as the primary endpoint, which was analyzed by the Kaplan-Meier method before and after propensity score matching (PSM). Subgroup analysis was conducted based on the established staging system and survival risk stratification. Results: A total of 335 patients were enrolled in this study, including 39 patients in the p-TACE group and 296 patients in the non-TACE group. Median OS in the p-TACE group was longer than that in the non-TACE group (63.0 months vs. 18.0 months, P=0.041), which was confirmed after 1:1 PSM (P=0.009). According to the 8th TNM staging system, patients with stage II and stage III stage would be benefited from p-TACE (P=0.021). Subgroup analysis stratified by risk factors showed that p-TACE could only benefit patients with risk factors

Adjuvants are common vaccine components. Novel adjuvants may improve the protective immunity conferred by vaccines against poultry diseases. Here, a less-hemolytic saponin, platycodin D (PD), isolated from the root of Platycodon grandiflorum was investigated as a potential alternative adjuvant. PD was tested as an adjuvant in the infectious bronchitis (IB) vaccine, because the existing IB vaccine has often failed to induce effective immune responses. The adjuvant activity of PD in conjunction with IB vaccine was evaluated in this study. Compared to control treatment, PD treatment significantly increased the proliferation of chicken peripheral blood mononuclear cells, concentration of interferon-γ in culture supernatants, and anti-IB antibody titer. In chickens pre-challenged with the Mass 41 infectious bronchitis virus (IBV), PD administration resulted in fewer and less severe clinical signs, lower mortality rate, and higher protection compared to control treatment. Histopathological examination showed that the lungs and kidneys of PD-treated chickens displayed fewer pathological lesions than those of control chickens. Our results also demonstrated that this new vaccine adjuvant improved chicken humoral and cellular immune responses without any side effects. Hence, our findings suggest that PD might serve as an effective adjuvant in IBV vaccines.

Background: To explore the effects of postoperative adjuvant transarterial chemoembolization (PA-TACE) on the prognosis of HCC patients with Portal Vein Tumor Thrombus (PVTT) undergoing resection, and to develop a PA-TACE-related nomogram for predicting survival individually. Patients and Methods: Two hundred and ninety-three consecutive HCC patients with PVTT under R0 hepatectomy were recruited. Forty-seven cases had recurrence within one month after surgery. The remaining 246 cases consisted of 90 PA-TACE and 156 non-PA-TACE cases. COX regression analysis was performed for overall survival (OS) or recurrence-free survival (RFS) of these 246 cases, allowing the derivation of independent factors that were integrated into the nomogram. C-index, calibration curves, and risk stratification were performed to evaluate the performance and discriminative power of the nomograms. Results: In 246 patients without recurrence within one month after surgery, the OS and RFS for the PA-TACE group were significantly better than those for the non-PA-TACE group (P

Background: Human cytomegalovirus (HCMV) causes a ubiquitous infection which can pose a significant threat for immunocompromised individuals, such as those undergoing solid organ transplant (SOT). Arguably, the most successful vaccine studied to date is the recombinant glycoprotein-B (gB) with MF59 adjuvant which, in 3 Phase II trials, demonstrated 43–50% efficacy in preventing HCMV acquisition in seronegative healthy women or adolescents and reduction in virological parameters after SOT. However, the mechanism of vaccine protection in seronegative recipients remains undefined. Methods: We evaluated samples from the cohort of seronegative SOT patients enroled in the Phase II glycoprotein-B/MF59 vaccine trial who received organs from seropositive donors. Samples after SOT (0–90 days) were tested by real-time quantitative PCR for HCMV DNA. Anti-gB antibody levels were measured by ELISA. Neutralization was measured as a decrease in infectivity for fibroblast cell cultures revealed by expression of immediate-early antigens. Findings: Serological analyses revealed a more rapid increase in the humoral response against gB post transplant in vaccine recipients than in those randomised to receive placebo. Importantly, a number of patient sera displayed HCMV neutralising responses – neutralisation which was abrogated by pre-absorbing the sera with recombinant gB. Interpretation: We hypothesise that the vaccine primed the immune system of seronegative recipients which, when further challenged with virus at time of transplant, allowed the host to mount rapid immunological humoral responses even under conditions of T cell immune suppression during transplantation. Keywords: Cytomegalovirus, Vaccination, Antibody responses, Prime-boost

Background: Disease-free survival (DFS) did not reflect accurate individual prognosis after initial diagnosis. As conditional DFS (CDFS) could provide dynamic prognostic information, we evaluated CDFS in these patients treated with or without sunitinib. Results: A total of 1329 patients with median follow-up 6.54 years were enrolled. CDFS improved continuously with disease-free survivorship increasing in both sunitinib and placebo group with minimal difference. In placebo arm, the CDFS of surviving to five year after living 1, 2, 3, and 4 years were 65%, 78%, 87%, and 95% (observed 5-year DFS: 51%). Dynamic changes of HR showed adjuvant sunitinib decrease relapse risks during the first 1.5 years after surgery (P < 0.03). Conclusions: Our study provided contemporary data of CDFS and change of relapse HR in high-risk ccRCC patients after adjuvant sunitinib or placebo. The remarkable improvement in CDFS highlighted the importance of disease-free interval as a strong indicator in patient counseling and surveillance planning. Materials and Methods: The primary end point was CDFS and the second end point was smooth hazard ratios (HR) for the prediction of relapses. The differences of conditional survival were compared with the calculation of d value.

The human commensal Staphylococcus aureus (SA) is a leading cause of skin/soft tissue and surgical-site infections, and bacteremia. Functional antibodies and T-cell-mediated immunity, particularly Th1/Th17 responses, are thought to mediate protection. Vaccine development may be hindered by modulation of vaccine-induced T cells by pathogen-activated immunoregulatory responses, e.g., via IL-10. We screened SA proteins for CD4+ T-cell-activating and IL-10/IL-17-inducing capacities using healthy donor-derived PBMCs. Responses were characterized (Th1/Th17/Th22/immunosuppressive IL-10-producing cells) using intracellular cytokine staining and flow cytometry. Phenotypic plasticity of Th1/Th17 cells was evaluated under pro- or anti-inflammatory conditions using modulatory cytokines. The impact of vaccination on SA-specific memory responses was assessed using samples from a clinical trial evaluating AS03-adjuvanted and non-adjuvanted multicomponent (CPS5/CPS8/α-toxin/ClfA) vaccines (NCT01160172). The donors exhibited SA-specific memory T-cell responses, indicative of pre-existing immunity to SA. We identified effective activators of Th1 responses (EbhA/IsaA/SdrE/MntC/Aaa/α-toxin), and Th17 and Th1/Th17 responses (EbhA/IsaA/SdrE and, to a lesser extent, α-toxin), but not of Th22 responses or IL-10 production. MRPII, IsdA, and ClfA were inefficient CD4+ T-cell activators in our assays. IL-10, likely produced by innate immune cells, influenced mainly Th1 cells by suppressing IFN-γ production. The memory CD4+ T-cells observed after long-term stimulation with α-toxin and ClfA indicated that vaccination with these proteins had induced expansion of pre-existing Th1 but not Th17 responses, without apparent adjuvant effect, confirming the trial data. The Th1/Th17-driving proteins (EbhA/IsaA/SdrE) shared low IL-10-promoting abilities and restricted phenotypic plasticity under pro- and anti-inflammatory conditions. Given the complex immunopathology and multiple virulence factors, identification of Th1/Th17-driving antigens, adjuvants and administration routes, and delineation of the role of memory responses, may advance vaccine development.

Transarterial chemoembolization (TACE) is a commonly used treatment modality in hepatocellular carcinoma (HCC). The ability to identify patients who will respond to TACE represents an important clinical need, and tumor gene expression patterns may be associated with TACE response. We investigated whether tumor transcriptome is associated with TACE response in patients with HCC. We analyzed transcriptome data of treatment-naive tumor tissues from a Chinese cohort of 191 HCC patients, including 105 patients who underwent TACE following resection with curative intent. We then developed a gene signature, TACE Navigator, which was associated with improved survival in patients that received either adjuvant or post-relapse TACE. To validate our findings, we applied our signature in a blinded manner to three independent cohorts comprising an additional 130 patients with diverse ethnic backgrounds enrolled in three different hospitals who received either adjuvant TACE or palliative TACE. TACE Navigator stratified patients into Responders and Non-Responders which was associated with improved survival following TACE in our test cohort (Responders: 67 months vs Non-Responders: 39.5 months, p

Equine influenza is a leading cause for respiratory illness in equines. Major control measures involve vaccination which requires continuous harmonization owing to antigenic drift. The present study focused on assessing the protective efficacy of an inactivated recombinant equine influenza virus (rgEIV) vaccine candidate adjuvanted with MontanideTM Pet Gel in murine model. The rgEIV was generated using reverse genetics by incorporating HA and NA segments from EIV/H3N8, clade 2-Florida sublineage in an A/WSN/33 /H1N1 backbone and inactivated by formalin. The vaccine was prepared by mixing inactivated rgEIV with MontanideTM Pet Gel adjuvant followed by intranasal inoculation into BALB/c mice intranasally. The immune responses and protective efficacy of the vaccine was evaluated by measurement of antibody titer, immunoglobulin subtyping, cytokines, clinical signs and pathological lesions after immunization and challenge with wild EIV. Serology and cytokine expression pattern indicated that the vaccine activated mixed Th1- and Th2-like responses of vaccine. Booster immunization stimulated strong antibody responses (HAI titre: 192 ± 28.6) at 42 days post immunization and the predominant antibody subtype was IgG1. Upregulation of interferon (IFN)-gamma, interleukin (IL)-12 and IL-2 levels indicates effective induction of Th1 type response. We found that vaccination has protected mice against equine influenza virus challenge as adjudged through a lack of nonappearance of visible clinical signs of disease, no loss of body weight loss, reduced pathology in the lungs and markedly reduced virus shedding from the respiratory tract. Therefore, we conclude that recombinant EIV vaccine candidate adjuvanted with MontanideTM Pet Gel could aid in quick harmonization of the vaccines through replacement of HA and NA genes for control of EIV outbreaks.

Highlights • Adoption of adjuvant bisphosphonates for early breast cancer into standard clinical practice. • UK and Australian experience of adjuvant bisphosphonates in early breast cancer. • Pathway taken for adjuvant bisphosphonates implementation in the UK. • Steps to increase update of adjuvant bisphosphonates in early breast cancer. • Improve the care of women with early breast cancer., International guidelines recommend adjuvant bisphosphonates (BPs) for post-menopausal women with early breast cancer to reduce recurrence and mortality. However, globally, wide variation exists in their adoption. In the UK, adjuvant BPs were a recommendation in the breast cancer Clinical Reference Group service specification and were included as a priority for implementation by the national oncologists group UK Breast Cancer Group in November 2015, promoting national uptake, guidance and funding arrangements. In 2018, adjuvant BPs were recommended by the UKs National Institute for Health and Care Excellence. In Australia, adjuvant BPs are still ‘off-label’ and do not receive national reimbursement or endorsement. To date there has been no research into the prescribing habits of these agents in Australia. With the aim to gather data on adjuvant BPs prescribing practices, online surveys were developed and disseminated to breast oncologists in both countries between December 2018 and June 2019. Almost all of the UK oncologists prescribed adjuvant BPs, demonstrating that education, endorsement from professional bodies, presence of national guidelines and funding decisions have been critical to implementation. In contrast, only 48% of the Australian responders prescribed adjuvant BPs, while 83% reported that they would prescribe them if funding was available. Lack of local protocol guidance was also seen as a major barrier. This study was intended to assess the pathway taken for adjuvant BP implementation in the UK and how it might inform changes in Australian practice and also guide other countries with similar issues with the ultimate aim of improving the care of women with early breast cancer globally.

Synthetic vaccines, based on antigenic peptides that comprise MHC−I and MHC‐II T‐cell epitopes expressed by tumors, show great promise for the immunotherapy of cancer. For optimal immunogenicity, the synthetic peptides (SPs) should be adjuvanted with suitable immunostimulatory additives. Previously, we have shown that improved immunogenicity in vivo is obtained with vaccine modalities in which an SP is covalently connected to an adjuvanting moiety, typically a ligand to Toll‐like receptor 2 (TLR2). SPs were covalently attached to UPam, which is a derivative of the classic TLR2 ligand Pam3CysSK4. A disadvantage of the triply palmitoylated UPam is its high lipophilicity, which precludes universal adoption of this adjuvant for covalent modification of various antigenic peptides as it renders the synthetic vaccine insoluble in several cases. Here, we report a novel conjugatable TLR2 ligand, mini‐UPam, which contains only one palmitoyl chain, rather than three, and therefore has less impact on the solubility and other physicochemical properties of a synthetic peptide. In this study, we used SPs that contain the clinically relevant neoepitopes identified in a melanoma patient who completely recovered after T‐cell therapy. Homogeneous mini‐UPam‐SP conjugates have been prepared in good yields by stepwise solid‐phase synthesis that employed a mini‐UPam building block pre‐prepared in solution and the standard set of Fmoc‐amino acids. The immunogenicity of the novel mini‐UPam‐SP conjugates was demonstrated by using the cancer patient's T‐cells., Less lipophilic ligands: A novel TLR2 ligand has been designed and synthesized to be subsequently conjugated with synthetic peptides containing clinically relevant neoepitopes. By treating HEK‐TLR2 cells, human moDCs and antigen specific human T‐cells with these new constructs, the immunogenic potential of the new TLR2 ligand and the conjugation of TLR ligands with neoepitopes was assessed.

This study aimed to evaluate the influence of maltodextrin addition on the flow properties of cocoa pulp powder obtained by spray and freeze drying. Cocoa pulp samples received 15% and 30% (m m-1) maltodextrin DE20. Two drying methods were used, spray and freeze drying. Powder morphology was evaluated through scanning electron microscopy (SEM). Wall friction angle, bulk density and tapped density were determined. Carr index (CI), Hausner ratio (HR) and flow index (FI) were used to evaluate powder flow. The particles powders obtained by spray drying showed rounded shapes, whereas the freeze dried powder showed irregular shapes. Increased maltodextrin concentration in the samples altered the powder particle size by spray drying and powder particle surface by freeze drying. The powder by spray drying of the sample with 30% of maltodextrin showed smallest wall friction angles, 13.4 to 14.9. The powder by freeze drying of the samples with 15% and 30% of maltodextrin showed wall friction angles between 14.0 and 20.6. Regarding flow, the powders by spray drying containing 30% of maltodextrin showed the best CI, HR and FI, 24.88, 1.33 and 4.88, respectively, being considered an acceptable flow. According to CI, HR and FI values, samples with 15% of maltodextrin produced powder classified as difficult flow in both methods applied. The higher maltodextrin concentration in cocoa pulp, the lower agglomeration in the powder by spray drying and the smoother particles surfaces in the powder by freeze drying. The addition of maltodextrin to the samples, for both drying methods, improve the powder flow and decrease the powder cohesion. Resumo O objetivo deste estudo foi avaliar a influência da adição de maltodextrina nas propriedades de escoamento do pó de polpa de cacau obtido em secagem por aspersão e liofilização. Amostras da polpa de cacau foram adicionadas com 15% e 30% (m m-1) de maltodextrina DE20. Foram utilizados dois métodos de secagem: por aspersão e liofilização. A morfologia do pó foi avaliada por microscopia eletrônica de varredura (MEV). Foram determinados o ângulo de atrito com a parede, a densidade aparente e a de compactação. O índice de Carr (CI), a razão de Hausner (HR) e o índice de fluxo (FI) foram determinados e utilizados para avaliar a fluidez dos pós. As partículas do pó obtido por aspersão apresentaram formato arredondado, enquanto as do liofilizado apresentaram formas irregulares. O aumento da concentração de maltodextrina nas amostras alterou o tamanho das partículas do pó obtido por spray dryer e a superfície das partículas daquele obtido por liofilização. O pó obtido por spray dryer, a partir da amostra com 30% de maltodextrina, apresentou os menores ângulos de atrito com a parede, entre 13,4 e 14,9. Os pós obtidos por liofilização das amostras com 15% e 30% de maltodextrina apresentaram ângulos de fricção entre 14,0 e 20,6. Em relação à fluidez, os pós obtidos por spray dryer contendo 30% de maltodextrina apresentaram os melhores CI, HR e FI, com valores de 24,88, 1,33 e 4,88, respectivamente, classificando-os como de aceitável escoamento. De acordo com os valores de CI, HR e FI, amostras contendo 15% de maltodextrina produziram pós classificados como de difícil escoamento em ambas as secagens utilizadas. Quanto maior a concentração de maltodextrina na polpa de cacau, menor a aglomeração nos pós obtidos por spray dryer e mais lisas as superfícies das partículas do pó obtido por liofilização. A adição de maltodextrina nas amostras, em ambos os métodos de secagem, melhora a fluidez e diminui a coesão dos pós.

Adjuvants are key immunostimulatory components in vaccine formulations, which improve the immune response to the co‐administered antigen. The saponin natural product QS‐21 is one of the most promising immunoadjuvants in the development of vaccines against cancer and infectious diseases but suffers from limitations that have hampered its widespread human use. Previous structure–activity relationship studies have identified simplified saponin variants with truncated carbohydrate chains, but have not focused on the influence of the linear oligosaccharide domain of QS‐21 in adjuvant activity. Herein, an expeditious 15‐step synthesis of new linear trisaccharide variants of simplified QS‐21‐derived adjuvants is reported, in which the complex terminal xylose‐rhamnose moiety has been replaced with commercially available, simpler lactose and cellobiose disaccharides in a β‐anomeric configuration. In vivo immunological evaluation of the synthetic saponins showed attenuated antibody responses, highlighting the negative impact of such carbohydrate modifications on adjuvant activity, which could be associated with higher saponin conformational flexibility., Shorter route to vaccine adjuvants: These multidisciplinary studies have provided streamlined access to new terminal disaccharide variants of truncated, QS‐21 inspired saponin vaccine adjuvants through an expeditious synthetic route, and have identified a critical role for the native rhamnose‐xylose moiety in adjuvant activity in connection with saponin conformation.

Aims To generate different larval stages of Strongylus vulgaris and to study cytokine responses in cultures of eqPBMC exposed to defined larval stages of S. vulgaris and cyathostomins with the aim to understand the early immune reaction to these parasites. Methods and results EqPBMC were exposed to S. vulgaris larvae (L3, exsheated L3 and L4) and cyathostomin L3 and analysed for cytokine gene expression. Procedures for decontamination, culturing and attenuation of larvae were established. Transcription of IL‐4, IL‐5 and IL‐13 was induced by both S. vulgaris and cyathostomin L3. Moulting of S. vulgaris from L3 to L4 stage was accompanied by a shift to high expression of IL‐5 and IL‐9 (exsheated L3 and L4) and IFN‐γ (L4 only). In parallel, the adjuvant G3 modified the cytokine profile induced by both parasites by reducing the expression of IL‐4, IL‐5 and IL‐10 while concomitantly enhancing the expression of IFN‐γ. Conclusion The L4 stage of S. vulgaris generated a cytokine profile different from that induced by the earlier L3 stage of S. vulgaris and cyathostomins. This diversity depending on the life cycle stage will have implications for the choice of antigen and adjuvant in future vaccine design.

7 páginas, 3 figuras, 4 tablas., The experimental vaccine for bovine malignant catarrhal fever consists of viable attenuated alcelaphine herpesvirus 1 (AlHV-1) derived‘ by extensive culture passage, combined with an oil-in-water adjuvant, delivered intramuscularly. This immunisation strategy was over 80% effective in previous experimental and field trials and protection appeared to be associated with induction of virus-neutralising antibodies. Whether the vaccine virus is required to be viable at the point of immunisation and whether adjuvant is required to induce the appropriate immune responses remains unclear. To address these issues two studies were performed, firstly to analyse immune responses in the presence and absence of adjuvant and secondly, to investigate immune responses to vaccines containing adjuvant plus viable or inactivated AlHV-1. The first study showed that viable attenuated AlHV-1 in the absence of adjuvant induced virus-specific antibodies but the titres of virus-neutralising antibodies were significantly lower than those induced by vaccine containing viable virus and adjuvant, suggesting adjuvant was required for optimal responses. In contrast, the second study found that the vaccine containing inactivated (>99.9%) AlHV-1 induced similar levels of virus-neutralising antibody to the equivalent formulation containing viable AlHV-1. Together these studies suggest that the MCF vaccine acts as an antigen depot for induction of immune responses, requiring adjuvant and a suitable antigen source, which need not be viable virus. These obser- vations may help in directing the development of alternative MCF vaccine formulations for distribution in the absence of an extensive cold chain., This work was supported by the Scottish Government Rural and Environment Science and Analytical Services (RESAS) Strategic Research Programme; the Department for International Develop- ment and the Biotechnology and Biological Sciences Research Council, grant BB/H008950/1; and GALVmed with funding from Bill & Melinda Gates Foundation and UKAID, grant MRI-R34A0985

Objective This study evaluated the effects of mesenchymal stem cell-extracellular vesicles (MSC-EVs) on chondrocyte proliferation in vitro and on cartilage repair in vivo following bone marrow stimulation (BMS) of focal chondral defects of the knee. Methods Six adult Göttingen minipigs received 2 chondral defects in each knee. The pigs were randomized to treatment with either BMS combined with MSC-EVs or BMS combined with phosphate-buffered saline (PBS). Intraarticular injections MSC-EVs or PBS were performed immediately after closure of the surgical incisions, and at 2 and 4 weeks postoperatively. Repair was evaluated after 6 months with gross examination, histology, histomorphometry, immunohistochemistry, and micro-computed tomography (µCT) analysis of the trabecular bone beneath the defect. Results Defects treated with MSC-EVs had more bone in the cartilage defect area than the PBS-treated defects (7.9% vs. 1.5%, P = 0.02). Less than 1% of the repair tissue in both groups was hyaline cartilage. International Cartilage and Joint Preservation Society II histological scoring showed that defects treated with MSC-EVs scored lower on “matrix staining” (20.8 vs. 50.0, P = 0.03), “cell morphology” (35.4 vs. 53.8, P = 0.04), and “overall assessment” (30.8 vs. 52.9, P = 0.03). Consistently, defects treated with MSC-EVs had lower collagen II and higher collagen I areal deposition. Defects treated with MSC-EVs had subchondral bone with significantly higher tissue mineral densities than PBS-treated defects (860 mg HA/cm3 vs. 838 mg HA/cm3, P = 0.02). Conclusion Intraarticular injections of MSC-EVs in conjunction with BMS led to osseous ingrowth that impaired optimal cartilage repair, while enhancing subchondral bone healing.

Objective: The aim of this study was to compare the clinical efficacy of neoadjuvant chemoradiotherapy (NACRT) combined with postoperative adjuvant XELOX (Oxaliplatin +Capecitabine) chemotherapy and postoperative adjuvant chemotherapy (ACT) with XELOX for local advanced gastric cancer (LAGC). Methods: In this prospectively randomized trial, we investigated the effect of NACRT combined with postoperative ACT for LAGC. 60 patients were randomly divided into NACRT group and ACT group, with 30 patients in each group. Patients in NACRT group were given three-dimensional conformal radiotherapy (45 Gy/1.8 Gy/f) accompanied by synchronous XELOX of two cycles, followed by surgery, and then postoperative adjuvant XELOX chemotherapy of four cycles was performed. Patients in ACT group received surgery in advance, and then XELOX chemotherapy of six cycles was given. Results: The objective response rate of NACRT was 76.7%. The overall incidence of postoperative complications in NACRT group was not significantly different from that in ACT group (23.1% vs 30.0%, p = 0.560). The 1 year, 2 years, and 3 years progression-free survival (PFS）and overall survival (OS) in NACRT and ACT groups were 80.0% vs 56.7%, 73.3% vs 46.7%, 60.0% vs 33.3%, and 86.7% vs 80.0%, 76.7% vs 66.7%, 63.3% vs 50.0%, respectively. Patients in NACRT group showed a significantly higher R0 resection rate (84.6% vs 56.7%, p = 0.029),lower loco-regional recurrence rate (36.7% vs 11.5%, p = 0.039), longer PFS (p = 0.019) and freedom from locoregional progression(FFLP) (p = 0.004) than patients in ACT group, while there was no difference in OS (p = 0.215) and in toxicity incidence (p > 0.05). Conclusions: NACRT combined with postoperative adjuvant XELOX chemotherapy can improve R0 resection rate, reduce loco-regional recurrence, prolong PFS and FFLP without increasing the incidence of postoperative complications in patients with LAGC. Advances in knowledge: Compared with postoperative adjuvant chemotherapy, locally advanced gastric cancer patients may benefit from neoadjuvant chemoradiotherapy, and toxicity associated with chemoradiotherapy was tolerant and manageable.

Prevention and intervention methods are urgently needed to curb the global pandemic of coronavirus disease‐19 caused by severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2). Herein, a general pro‐antigen strategy for subunit vaccine development based on the reversibly formulated receptor binding domain of SARS‐CoV‐2 spike protein (S‐RBD) is reported. Since the poor lymph node targeting and uptake of S‐RBD by antigen‐presenting cells prevent effective immune responses, S‐RBD protein is formulated into a reversible nanogel (S‐RBD‐NG), which serves as a pro‐antigen with enhanced lymph node targeting and dendritic cell and macrophage accumulation. Synchronized release of S‐RBD monomers from the internalized S‐RBD‐NG pro‐antigen triggers more potent immune responses in vivo. In addition, by optimizing the adjuvant used, the potency of S‐RBD‐NG is further improved, which may provide a generally applicable, safer, and more effective strategy for subunit vaccine development against SARS‐CoV‐2 as well as other viruses., The receptor binding domain of severe acute respiratory syndrome coronavirus‐2 spike protein is reversibly formulated into a protein nanogel, which serves as a pro‐antigen and elicites more potent and rapid protective immune responses as a potential candidate for subunit vaccine development against severe acute respiratory syndrome coronavirus‐2.