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•CMV seronegative girls between 12 and 17 years of age received CMV glycoprotein B (gB) vaccine with MF59 or saline placebo at 0, 1 and 6 months.•The vaccine was generally well tolerated, although local and systemic adverse events were significantly more common in the vaccine group.•In the per protocol population vaccine efficacy was 43% after 3 doses, p=0.20 and 45%, p=0.08 after 2 doses.•We conclude the vaccine was safe and immunogenic and although the efficacy did not reach significance, the results are consistent with a previous study in adult women (Pass et al NEJM 360:1191, 2009) using the same formulation. Cytomegalovirus (CMV) is a leading cause of congenital infection and an important target for vaccine development. CMV seronegative girls between 12 and 17 years of age received CMV glycoprotein B (gB) vaccine with MF59 or saline placebo at 0, 1 and 6 months. Blood and urine were collected throughout the study for evidence of CMV infection based on PCR and/or seroconversion to non-vaccine CMV antigens. 402 CMV seronegative subjects were vaccinated (195 vaccine, 207 placebo). The vaccine was generally well tolerated, although local and systemic adverse events were significantly more common in the vaccine group. The vaccine induced gB antibody in all vaccine recipients with a gB geometric mean titer of 13,400EU; 95%CI 11,436, 15,700, after 3 doses. Overall, 48 CMV infections were detected (21 vaccine, 27 placebo). In the per protocol population (124 vaccine, 125 placebo) vaccine efficacy was 43%; 95%CI: −36; 76, p=0.20. The most significant difference was after 2 doses, administered as per protocol; vaccine efficacy 45%, 95%CI: −9; 72, p=0.08. The vaccine was safe and immunogenic. Although the efficacy did not reach conventional levels of significance, the results are consistent with a previous study in adult women (Pass et al. N Engl J Med 2009;360:1191) using the same formulation.

In this report regarding an MF59-adjuvanted 2009 H1N1 monovalent vaccine, significant immune responses were elicited by the administration of one or two doses of vaccine (with or without the MF59 adjuvant) in most subjects within 2 to 3 weeks. Higher titers were seen in subjects who received the MF59-adjuvanted vaccine. In this report regarding a monovalent MF59-adjuvanted 2009 H1N1 vaccine, significant immune responses were elicited by the administration of one or two doses of vaccine (with or without the MF59 adjuvant) in most subjects within 2 to 3 weeks. Higher titers were seen in subjects who received the MF59-adjuvanted vaccine. The emergence of the 2009 pandemic influenza A (H1N1) virus demonstrates the unpredictable nature of influenza. 1 The virus has the potential to cause disease, death, and socioeconomic disruption, 2 , 3 and modeling suggests that the effect of the virus can be reduced by immunization. 4 The development of effective vaccines is a public health priority. Traditional seasonal influenza vaccines are produced from reassortant vaccine strains grown in hens' eggs. However, demand for vaccine against the 2009 H1N1 virus will most likely exceed the supply if this method of manufacturing is solely used. Cell culture provides an additional platform for the manufacture of . . .

While it remains impossible to predict the timing of the next influenza pandemic, novel avian influenza A viruses continue to be considered a significant threat. A Phase II study was conducted in healthy adults aged 18–64 years to assess the safety and immunogenicity of two intramuscular doses of pre-pandemic 2017 influenza A(H7N9) inactivated vaccine administered 21 days apart. Participants were randomized (n = 105 in each of Arms 1–3) to receive 3.75 μg, 7.5 μg or 15 μg of hemagglutinin (HA) with MF59® adjuvant, or 15 μg of HA unadjuvanted vaccine (n = 57, Arm 4). The three MF59 adjuvanted vaccines and the 15 μg unadjuvanted vaccine were safe and well-tolerated. Little antibody activity was detected against the A(H7N9) vaccine antigen after the first vaccination across study Arms. After second vaccination, the three adjuvanted Arms showed increases in hemagglutination inhibition (HAI), neutralizing (Neut), and neuraminidase inhibition (NAI) geometric mean titers (GMT), peaking at 21 days post second vaccination. The percentage of participants with titer ≥1:40 and seroconversion rates for HAI were 30–43 % and 0 for the adjuvanted Arms and the unadjuvanted Arm, respectively. Antibody responses against antigenically drifted A(H7N9) strains A/Shanghai/2/2013 and A/Guangdong/17SF003/2016 showed similar trends. Exploratory linear modeling of HAI and Neut responses post second vaccination revealed significantly lower log antibody titers among older participants (aged 35–49 and 50–64 years) compared to participants aged 18–34 years after adjusting for study vaccination, BMI, sex, and prior seasonal influenza vaccination. Post second vaccination, participants who received seasonal influenza vaccination in at least one of the two previous seasons had significantly lower log antibody titers than participants who did not. Adjuvanted doses of vaccine provided higher antibody responses, on average, than the 15 μg unadjuvanted vaccine. Proportion of participants achieving seroconversion and antibody titers ≥40 remained below 50 % in all study Arm. •The three dosages of MF59 adjuvanted A(H7N9) vaccine were safe and well-tolerated.•There were similar influenza antibody responses across the adjuvanted Arms.•MF59 adjuvant increased the magnitude and breadth of antibody responses.•Most participants in each Arm did not seroconvert against the vaccine strain.•There was little antibody response in the unadjuvanted 15 μg Arm.

Influenza poses a significant global healthcare burden, with up to 1 billion infections annually, and poorer outcomes in vulnerable populations such as older adults. Vaccination effectiveness is often lower in elderly individuals. By adding an adjuvant and using cell-based vaccine production methods, the MF59-adjuvanted quadrivalent cell-based influenza vaccine (aQIVc) may boost immunogenicity and vaccine effectiveness in this population. We report the results of a randomised proof-of-concept study, investigating the immunogenicity and safety of aQIVc. Eligible participants aged ≥50 years were randomised 1: 1:1:1 to receive aQIVc (n = 116), a non-adjuvanted quadrivalent cell-based influenza vaccine (QIVc; n = 119), an MF59-adjuvanted quadrivalent egg-based influenza vaccine (aQIV; n = 116), or a high-dose quadrivalent recombinant influenza vaccine (QIVr; n = 120). The primary objective was to assess immunogenicity of aQIVc vs the comparators by haemagglutination inhibition (HI) assay 28 days post-vaccination. Secondary objectives included immunogenicity of aQIVc vs comparators 28 days and 180 post-vaccination by microneutralisation assay and 180 days post-vaccination by HI assay; and reactogenicity and safety of all study vaccines. Compared with QIVc and aQIV, aQIVc elicited a higher immune response (adjusted geometric mean titre [GMT] ratio range 1.18–1.85) against all four influenza strains at Day 29. Against QIVr, aQIVc elicited lower responses against A strains (adjusted GMT ratio range 0.79–0.84), and higher responses against B strains (adjusted GMT ratio range 1.15–1.26). Estimated GMT ratios were generally higher in the subgroup of participants aged ≥65 years vs those aged 50–64 years. aQIVc was well tolerated, eliciting similar rates of solicited local adverse events (AE) and slightly higher rates of solicited systemic AE than aQIV, and a higher rate of all solicited AE than QIVc and QIVr. No safety concern was identified. These data support further investigation of additional formulations of aQIVc in adults aged ≥50 years. Clinical trial registry:NCT04576702 •We assessed an adjuvanted, cell-based influenza vaccine (aQIVc) in adults ≥50 years•aQIVc elicited higher immune responses than a non-adjuvanted comparator•aQIVc elicited higher immune responses than an egg-based adjuvanted comparator•aQIVc showed an acceptable safety profile; no safety concerns were identified•These data are proof-of-concept for seasonal aQIVc vaccination in adults ≥50 years

Immune responses to vaccination are tested in clinical trials. This process usually requires years especially when immune memory and persistence are analyzed. Markers able to quickly predict the immune response would be very useful, particularly when dealing with emerging diseases that require a rapid response, such as avian influenza. To address this question we vaccinated healthy adults at days 1, 22, and 202 with plain or MF59-adjuvanted H5N1 subunit vaccines and tested both cell-mediated and antibody responses up to day 382. Only the MF59-H5N1 vaccine induced high titers of neutralizing antibodies, a large pool of memory H5N1-specific B lymphocytes, and H5-CD4⁺ T cells broadly reactive with drifted H5. The CD4⁺ response was dominated by IL-2⁺ IFN-γ⁻ IL-13⁻ T cells. Remarkably, a 3-fold increase in the frequency of virus-specific total CD4⁺ T cells, measurable after 1 dose, accurately predicted the rise of neutralizing antibodies after booster immunization and their maintenance 6 months later. We suggest that CD4⁺ T cell priming might be used as an early predictor of the immunogenicity of prepandemic vaccines.

Polysorbate (PS) is routinely used in biopharmaceutical formulation to stabilize the active pharmaceutical ingredient.Trace amounts of hydrolytic host cell proteins (HCPs) persist in the downstream purification process and can degrade PS in the final drug product over time, compromising its stability.Genomic removal of PS-degrading enzymes in the Chinese hamster ovary (CHO) host cell line allowed their traceless removal from the bioprocess.The combined knockout (KO) of the genes encoding nine previously confirmed PS-degrading HCPs resulted in a viable CHO host cell line with strongly reduced PS degradation potential.This is the first report of successful excision of a gene cluster >1 Mb in size from the CHO genome.The final multi-hydrolase KO cell line yielded competitive monoclonal antibody titers with high product quality and significantly reduced hydrolytic activity. Enzymatic degradation of polysorbates (PS) in biologic drug formulations is often caused by hydrolytic host cell proteins (HCPs) and can lead to particle formation and reduced shelf-life. Here, we present a host cell line-engineering approach by removing nine Chinese hamster ovary (CHO) host cell hydrolases, which have previously been confirmed to degrade PS. Strikingly, the sequential genomic knockout (KO) of these hydrolases, including the genetic removal of two entire gene clusters of unprecedented size, yielded viable CHO host cell line variants. This novel host cell line was further optimized using the additional KO of the two key proapoptotic genes, Bax and Bak1. Thus, we generated a competitive multi-hydrolase KO CHO host cell line that was further shown to be highly suitable for the generation of recombinant therapeutic glycoproteins. Most importantly, PS degradation and hydrolytic activity were drastically reduced, providing an avenue toward future PS degradation-free biologics-manufacturing processes. Enzymatic degradation of polysorbates (PS) in biologic drug formulations is often caused by hydrolytic host cell proteins (HCPs) and can lead to particle formation and reduced shelf-life. Here, we present a host cell line-engineering approach by removing nine Chinese hamster ovary (CHO) host cell hydrolases, which have previously been confirmed to degrade PS. Strikingly, the sequential genomic knockout (KO) of these hydrolases, including the genetic removal of two entire gene clusters of unprecedented size, yielded viable CHO host cell line variants. This novel host cell line was further optimized using the additional KO of the two key proapoptotic genes, Bax and Bak1. Thus, we generated a competitive multi-hydrolase KO CHO host cell line that was further shown to be highly suitable for the generation of recombinant therapeutic glycoproteins. Most importantly, PS degradation and hydrolytic activity were drastically reduced, providing an avenue toward future PS degradation-free biologics-manufacturing processes. [Display omitted] In the opinion of the authors, the presented technology of knocking out multiple host cell hydrolases from the Chinese hamster ovary (CHO) host cell genome to reduce polysorbate (PS) degradation is ready for implementation. We proved key considerable aspects when generating a novel CHO host cell line, such as transfectability, metabolic selection, ability to endure a conventional fed-batch process without compromising productivity, and consistent product quality attributes compared with the parental host cell line. Given that we performed cultivations in small-scale bioreactors mimicking the real-world process, we suggest a technology readiness level (TRL) of 5 out of 9 (based on NASA’s TRL system). In terms of the recently published Biomanufacturing Readiness Level (BRL) developed by the National Institute for Innovation in Manufacturing Biopharmaceuticals (NIIMBL), we would classify our technology at BLR 4 out of 9, as we have performed initial tests with industry-relevant feedstocks. Nevertheless, full-scale implementation might harbor further implications. In contrast to the parental CHO host cell line, detailed characterization data on the multi-hydrolase KO host cell line are not available. To ensure its seamless application in an industrial bioprocess, further characterization experiments should test media fit, long-term stability of clonal production cell lines, suitability to express a variety of biologics (including complex antibody formats), and capability for large-scale bioreactor cultivations. In addition, we can only predict that the presented reduction of hydrolytic activity translates all the way down to the final drug product. A complete bioprocess including the final formulation steps followed by stability studies will ultimately demonstrate whether our technology has the ability to solve the PS degradation challenge entirely. Enzymatic degradation of polysorbates (PS) in biologic formulations is often caused by hydrolytic host cell proteins (HCPs) and can lead to particle formation and reduced shelf-life. Combining hydrolase knockouts in a single Chinese hamster ovary (CHO) host cell line enabled traceless enzyme removal and represented a big step toward the production of PS degradation-free biologics.

The dynamics and molecular mechanisms underlying vaccine immunity in early childhood remain poorly understood. Here we applied systems approaches to investigate the innate and adaptive responses to trivalent inactivated influenza vaccine (TIV) and MF59-adjuvanted TIV (ATIV) in 90 14- to 24-mo-old healthy children. MF59 enhanced the magnitude and kinetics of serum antibody titers following vaccination, and induced a greater frequency of vaccine specific, multicytokine-producing CD4⁺ T cells. Compared with transcriptional responses to TIV vaccination previously reported in adults, responses to TIV in infants were markedly attenuated, limited to genes regulating antiviral and antigen presentation pathways, and observed only in a subset of vaccinees. In contrast, transcriptional responses to ATIV boost were more homogenous and robust. Interestingly, a day 1 gene signature characteristic of the innate response (antiviral IFN genes, dendritic cell, and monocyte responses) correlated with hemagglutination at day 28. These findings demonstrate that MF59 enhances the magnitude, kinetics, and consistency of the innate and adaptive response to vaccination with the seasonal influenza vaccine during early childhood, and identify potential molecular correlates of antibody responses.

•No interference was observed with antibody responses to influenza or pneumococcal antigens.•MF59-adjuvanted influenza vaccine might have enhanced responses to some pneumococcal antigens.•Concomitant vaccination had no additive effects on adverse events. Concomitant administration of influenza and pneumococcal vaccines facilitates their uptake by older adults; however, data on immunogenicity and safety of concomitant administration of adjuvanted trivalent inactivated influenza vaccine (aIIV3) and 23-valent pneumococcal polysaccharide vaccine (PPSV23) have not been reported. Subjects aged ≥65 years (N=224) were randomized 1:1:1:1 to receive MF59-aIIV3 alone, MF59-aIIV3+PPSV23 in contralateral arms, MF59-aIIV3+PPSV23 in the same arm or PPSV23 alone (Clinical Trial Number – NCT02225327). Hemagglutination inhibition assay and multiplex opsonophagocytic killing assay were used to compare immunogenicity after single or concomitant vaccination. All groups met immunogenicity criteria for the influenza vaccine in older adults with similar seroconversion rates and geometric mean fold-increases, irrespective of concomitant vaccinations and injection site. For each pneumococcal serotype, opsonic index (OI) increased markedly after the PPSV23 vaccination, irrespective of the concomitant influenza vaccine. All subjects showed an OI≥8 for serotypes 6B, 18C and 19A post-vaccination, with a suggestion that the ipsilateral concomitant vaccination might be associated with higher OIs for some antigens. Local and systemic adverse events were more common in subjects receiving PPSV23 compared to those receiving aIIV3 alone. No interference was observed with antibody responses to influenza or pneumococcal antigens when aIIV3 and PPSV23 were administered concomitantly.

With the exception of alum, emulsion-based vaccine adjuvants have been administered to far more people than any other adjuvant, especially since the 2009 H1N1 influenza pandemic. The number of clinical safety and immunogenicity evaluations of vaccines containing emulsion adjuvants has correspondingly mushroomed. In this review, the authors introduce emulsion adjuvant composition and history before detailing the most recent findings from clinical and postmarketing data regarding the effects of emulsion adjuvants on vaccine immunogenicity and safety, with emphasis on the most widely distributed emulsion adjuvants, MF59® and AS03. The authors also present a summary of other emulsion adjuvants in clinical development and indicate promising avenues for future emulsion-based adjuvant development. Overall, emulsion adjuvants have demonstrated potent adjuvant activity across a number of disease indications along with acceptable safety profiles.

Emulsifying agents, which are common food additives in the human diet, induce low-grade inflammation and obesity/metabolic syndrome in mice, suggesting that further investigation into the potential impact of dietary emulsifiers on the gut microbiota and human heath are warranted. Harmful effects of dietary emulsifiers Non-genetic factors are important contributors to the pathogenesis of inflammatory conditions such as such as inflammatory bowel disease and metabolic syndrome. Here the authors find that mice on a diet containing emulsifying agents develop low-grade inflammation and obesity/metabolic syndrome. These conditions correlate with a decrease in gut microbiota–epithelial distance through degradation of mucus layers, altered species composition and pro-inflammatory potential. Emulsifying agents are common food additives in the human diet, and these findings suggest that further investigations are warranted into their potential impact on gut microbiota and human health. The intestinal tract is inhabited by a large and diverse community of microbes collectively referred to as the gut microbiota. While the gut microbiota provides important benefits to its host, especially in metabolism and immune development, disturbance of the microbiota–host relationship is associated with numerous chronic inflammatory diseases, including inflammatory bowel disease and the group of obesity-associated diseases collectively referred to as metabolic syndrome. A primary means by which the intestine is protected from its microbiota is via multi-layered mucus structures that cover the intestinal surface, thereby allowing the vast majority of gut bacteria to be kept at a safe distance from epithelial cells that line the intestine 1 . Thus, agents that disrupt mucus–bacterial interactions might have the potential to promote diseases associated with gut inflammation. Consequently, it has been hypothesized that emulsifiers, detergent-like molecules that are a ubiquitous component of processed foods and that can increase bacterial translocation across epithelia in vitro 2 , might be promoting the increase in inflammatory bowel disease observed since the mid-twentieth century 3 . Here we report that, in mice, relatively low concentrations of two commonly used emulsifiers, namely carboxymethylcellulose and polysorbate-80, induced low-grade inflammation and obesity/metabolic syndrome in wild-type hosts and promoted robust colitis in mice predisposed to this disorder. Emulsifier-induced metabolic syndrome was associated with microbiota encroachment, altered species composition and increased pro-inflammatory potential. Use of germ-free mice and faecal transplants indicated that such changes in microbiota were necessary and sufficient for both low-grade inflammation and metabolic syndrome. These results support the emerging concept that perturbed host–microbiota interactions resulting in low-grade inflammation can promote adiposity and its associated metabolic effects. Moreover, they suggest that the broad use of emulsifying agents might be contributing to an increased societal incidence of obesity/metabolic syndrome and other chronic inflammatory diseases.