The effects of cellulose powder and water activity (aw) on the equilibrium water content, glass transition temperature (Tg), mechanical relaxation, and caking of freeze-dried amorphous carbohydrate ...blend powders (dextrin-glucose mixture) were investigated. Water sorption isotherm and Tg-curve were obtained, and critical water activity (awc) was determined as the aw at Tg = 25 °C. There was a minor effect of cellulose powder on the awc. The degree of caking was negligible below the awc (glassy state), but increased remarkably above the awc (rubbery state). Cellulose powder played a dispersive role in the powder, and thus diminished the degree of caking. The caking behavior could be described by a stretched exponential function, and an experimental formula to predict the degree of caking as a function of aw/awc was obtained. The results for a food powder, with a more complex composition, largely deviated from the formula. Mechanical relaxation was evaluated as a force-reduction during isothermal compression. It was found that the relationship between the degree of caking and force-reduction could be described as a linear function independent of aw and cellulose powder content. The results for the food powder partially followed the linear relationship.
•Cellulose reduced the water sorption of dextrin-glucose blends at constant aw.•Critical water activity (awc) was evaluated at glass transition temperature of 25 °C.•Degree of caking and mechanical relaxation (ΔF) were reduced by cellulose above awc.•Caking was described by a stretched exponential function respect to aw/awc.•Caking increased linearly with ΔF independent of aw and cellulose content.
Effect of maltodextrin on the water sorption behavior and glass transition temperature (Tg) of freeze-dried mango solute (water-soluble mango material) was investigated. In addition, the caking ...property of freeze-dried mango puree and mango solute with and without maltodextrin was investigated. The Tg of mango solute (32 °C) increased discontinuously with maltodextrin weight fraction with a step-point between 0.6 and 0.7. The pulp of mango decreased the hygroscopicity and increased the Tg of mango powder to 49 °C. The degree of caking was reduced by the additions of maltodextrin and pulp because of the increased Tg. The powder samples except for maltodextrin showed a small degree of caking (below 13%) even below their Tg. An empirical model to predict the effect of water content on Tg was applied to the mango solute-maltodextrin mixtures. The predicted Tg values were in good agreement with the measured values (coefficient of determination = 0.919).
•Tg of mango solute (MS) increased with decreasing water and increasing maltodextrin (MD).•The pulp of mango decreased the hygroscopicity and increased the Tg of mango powder.•The degree of caking was reduced by the additions of MD and pulp because of the increased Tg.•The powder samples except for MD showed a small degree of caking even below their Tg.•An empirical model was effective for the prediction of water plasticizing to MS-MD mixtures.
Zika virus (ZIKV) has recently emerged as a pandemic associated with severe neuropathology in newborns and adults. There are no ZIKV-specific treatments or preventatives. Therefore, the development ...of a safe and effective vaccine is a high priority. Messenger RNA (mRNA) has emerged as a versatile and highly effective platform to deliver vaccine antigens and therapeutic proteins. Here we demonstrate that a single low-dose intradermal immunization with lipid-nanoparticle-encapsulated nucleoside-modified mRNA (mRNA-LNP) encoding the pre-membrane and envelope glycoproteins of a strain from the ZIKV outbreak in 2013 elicited potent and durable neutralizing antibody responses in mice and non-human primates. Immunization with 30 μg of nucleoside-modified ZIKV mRNA-LNP protected mice against ZIKV challenges at 2 weeks or 5 months after vaccination, and a single dose of 50 μg was sufficient to protect non-human primates against a challenge at 5 weeks after vaccination. These data demonstrate that nucleoside-modified mRNA-LNP elicits rapid and durable protective immunity and therefore represents a new and promising vaccine candidate for the global fight against ZIKV.
Glass transition temperature (
T
g
) and degree of caking for maltodextrin with added plasticizers (glucose, maltose, and sorbitol) were investigated. Onset and offset
T
g
s were evaluated by ...differential scanning calorimetry. From the onset and offset
T
g
-curves and water sorption isotherm, each critical water activity (
a
wc
) value was determined (water activity at each
T
g
= 25 °C). The
a
wc
range between onset
T
g
and offset
T
g
was extended by the addition of plasticizers. The degree of caking increased at a water activity just above
a
wc
of onset
T
g
, and completely caked at a water activity just below
a
wc
of offset
T
g
. Effect of
a
w
on the degree of caking became more gradual with the addition of plasticizers because of the extended
a
wc
range. According to the dependence of viscosity on temperature and water activity, a predictive model for the caking of amorphous powders was proposed based on the onset and offset
T
g
curves.
Water sorption isotherm, glass transition temperature (
T
g
), and caking properties of maca (
Lepidium meyenii Walpers
) powder were investigated. A broad endothermic shift reflecting continuously ...distributed glass transition was detected by differential scanning calorimetry, which prompted the evaluation of onset
T
g
and offset
T
g
. From the onset and offset
T
g
curves and water sorption isotherm, critical water activity (water activity at glass transition temperature of 25 °C) was determined to be 0.258 for the onset
T
g
and 0.480 for the offset
T
g
. The critical water activity values were validated by isothermal mechanical relaxation measurement. Caking properties of maca powder were compared with maltodextrin. The degree of caking and cake hardness for maltodextrin increased at a water activity just above critical water activity for onset
T
g
. Maca powder, on the other hand, showed a small degree of caking in the water activity range of 0.225 to 0.576, and the degree of caking drastically increased at a higher water activity. The hardness of maca cakes increased gradually at water activity values greater than 0.432. Since maca powder showed a continuously distributed glass transition, the molecular mobility required for caking was considered to have been provided incrementally by the increase in water activity above critical water activity for the onset
T
g
.
Whole tumor cell lysates consist of a mixture of tumor antigens and danger associated molecular patterns (DAMPs) that can be used for dendritic cell maturation and consequently for the activation of ...a polyclonal T cell-specific tumor response. We evaluated the in vitro efficacy of three different preparations of canine transmissible venereal tumor (TVT) cell lysates: hypochlorous acid-whole tumor cell lysates (HOCl-L), heat shock-whole tumor cell lysates (HS-L), and freeze-thaw cycles-whole tumor cell lysates (FT-L) for the maturation of canine-derived dendritic cells. Our results showed calreticulin, HSP70, and HSP90 release in the three tumor lysates preparations (HOCl-L, HS-L, and FT-L); however, HMGB1 was detected only in HOCl-L and FT-L. Additionally, the uptake by HOCl-L pulsed dendritic cell (DC) increased compared to HS-L and FT-L pulsed DC; and dendritic cell maturation was confirmed by the appropriate cell surface markers (CD11c, CD80, CD83, and MHCII). Furthermore, dendritic cells pulsed with HOCl-L, HS-L or FT-L were cultured with canine lymphocytes. There was an increase of Th1-type cytokines (IL-12, TNF-α, and IFN-γ), in all the tumor cell lysates co-cultures, this correlates with T lymphocyte activation and cytotoxic response. Our data confirm that TVT cell lysates can induce functional canine-DC and that HOCl-L is the most effective one. This preparation of TVT cell lysates with HOCl is an attractive approach that allows the recognition of neoantigens as potential tumor targets and DC priming and therefore could be used for cancer immunotherapy against TVT.
Ebola (EBOV), Marburg (MARV) and Sudan (SUDV) viruses are the three filoviruses which have caused the most fatalities in humans. Transmission from animals into the human population typically causes ...outbreaks of limited scale in endemic regions. In contrast, the 2013-16 outbreak in several West African countries claimed more than 11,000 lives revealing the true epidemic potential of filoviruses. This is further emphasized by the difficulty seen with controlling the 2018-2020 outbreak of EBOV in the Democratic Republic of Congo (DRC), despite the availability of two emergency use-approved vaccines and several experimental therapeutics targeting EBOV. Moreover, there are currently no vaccine options to protect against the other epidemic filoviruses. Protection of a monovalent EBOV vaccine against other filoviruses has never been demonstrated in primate challenge studies substantiating a significant void in capability should a MARV or SUDV outbreak of similar magnitude occur. Herein we show progress on developing vaccines based on recombinant filovirus glycoproteins (GP) from EBOV, MARV and SUDV produced using the
S2 platform. The highly purified recombinant subunit vaccines formulated with CoVaccine HT™ adjuvant have not caused any safety concerns (no adverse reactions or clinical chemistry abnormalities) in preclinical testing. Candidate formulations elicit potent immune responses in mice, guinea pigs and non-human primates (NHPs) and consistently produce high antigen-specific IgG titers. Three doses of an EBOV candidate vaccine elicit full protection against lethal EBOV infection in the cynomolgus challenge model while one of four animals infected after only two doses showed delayed onset of Ebola Virus Disease (EVD) and eventually succumbed to infection while the other three animals survived challenge. The monovalent MARV or SUDV vaccine candidates completely protected cynomolgus macaques from infection with lethal doses of MARV or SUDV. It was further demonstrated that combinations of MARV or SUDV with the EBOV vaccine can be formulated yielding bivalent vaccines retaining full efficacy. The recombinant subunit vaccine platform should therefore allow the development of a safe and efficacious multivalent vaccine candidate for protection against Ebola, Marburg and Sudan Virus Disease.
The browning, gelatinization of starch, water sorption, glass transition, and caking properties of freeze-dried maca (Lepidium meyenii Walpers) powders were investigated and compared with a ...commercial maca powder. The freeze-dried maca powders had lower optical density (browning) and higher enthalpy change for starch gelatinization than the commercial maca. This resulted from a difference in thermal history. The equilibrium water contents of the freeze-dried maca powders were higher than those of commercial maca at each water activity (aw) because of differences in amorphous part. The glass transition temperature (Tg) was evaluated by differential scanning calorimetry. There was a negligible difference in the anhydrous Tg (79.5–80.2 ºC) among the samples. The Tg-depression of freeze-dried maca powders induced by water sorption was more gradual than that of the commercial maca due to a difference in water insoluble material content. From the results, critical water activity (awc) was determined as the aw at which Tg becomes 25 ºC. There was negligible caking below aw = 0.328. At higher aw, the degree of caking remarkably increased with a large variation depending on the samples. The degree of caking could be described uniformly as a function of aw/awc. From these results, we propose an empirical approach to predict the caking of maca powders.
Context: Exosomes secreted by tumor cells are a good source of cellular components that stimulate the immune response, such as alarmins (mRNA, tetraspanins (CD9, CD63, CD81), heat-shock proteins, ...major histocompatibility complex class I molecules) and tumor-associated antigens. These properties permit to pulsed dendritic cells in the immunotherapy for many cancers types. The aim of this study was to demonstrate the use of exosomes derived from canine transmissible venereal tumor (CTVT) as an antigen to pulsed dendritic cells and its administration in dogs with CTVT as treatment against this disease.
Material and methods: From primary culture of CTVT cells the exosomes were isolated and characterized by scanning electron microscopy assay, dot blot and protein quantification. The monocytes of each patient were differentiated to dendritic cells (DC) and pulsed with CTVT exosomes (CTVTE). Phagocytosis, tumor size, populations of lymphocytes and IFN-c levels were evaluated.
Results: The CTVTE showed a size around 90 nm. CD81, CD63, CD9 and Hsp70 were expressed. Monocytes showed an expression of 85.71% for CD14
+
, 12.3% for CD80
+
, 0.1% for CD83
+
and 0.8% for DLA-II. In DC 5.1% for CD14
+
, 86.7% for CD80
+
, 90.1% for CD83
+
and 92.6% for DLA-II and a phagocytosis of 63% was obtained by FITC Dextran test. No side effects were observed in the experimental groups with our therapy. Tumor regression was of 100% at the seventh week, as well as an increase in the level of IFN-γ (142 pg/ml), and CD4
+
(28%) and CD8
+
(34%) cell percentage.
Discusion and conclusion: These results have shown that DC pulsed with tumor exosomes induce regression of the TVT in dogs.
Zaire ebolavirus (EBOV), Sudan ebolavirus (SUDV), and Marburg marburgvirus (MARV) are the most prevalent and pathogenic species of filovirus. Previously, we showed that glycoprotein antigens from ...each virus could be lyophilized to create thermostable monovalent subunit vaccines. However, cross-protection is not expected from the monovalent vaccines and therefore developing a trivalent filovirus vaccine would be desirable. Subunit protein vaccines often require the addition of an adjuvant to sufficiently boost the immunogenicity. Typically, liquid suspensions or emulsions of adjuvants and lyophilized antigens are stored in separate vials to avoid destabilizing interactions and are only mixed immediately before administration. Herein, we describe the development and characterization of monovalent and trivalent filovirus vaccines that are co-lyophilized with a squalane-in-water emulsion adjuvant. We found that the single-vial presentation retained adjuvant particle diameter and zeta potential after lyophilization and reconstitution. Furthermore, the trivalent vaccines elicited high antibody levels against all three antigens in mice and non-human primates. These results advance the prospect of developing a single-vial trivalent filovirus vaccine, which would enable easier distribution and administration of the vaccine to resource-poor areas.
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