Dioctadecyldimethylammonium Bromide Research Articles

Ordered porous films of single-walled carbon nanotubes using an ionic exchange reaction

Ordered porous films of single walled carbon nanotubes (SWCNTs) modified with a cationic surfactant, dimethyldioctadecylammonium bromide (DODMAB), are prepared at the air/water interface. We determined the parameters for tuning the well-defined reproducible porous films adjusting the casting conditions, the solvent polarity and DODMAB concentration, etc. Double-chain cations, DODMA+, can be completely removed through an ionic exchange reaction to obtain the ordered porous films of SWCNTs only. Compared with SWCNTs, the ordered porous SWCNTs films exhibit excellent antibacterial activity and electric conductivity. The ordered porous SWCNTs films were used for self-patterning of Ag nanoparticles attached to SWCNTs films which efficiently possess the catalytic reduction of organic dyes. This work may provide new avenues for the preparation of ordered functional carbon materials and is also envisaged to find applications in catalytic chemistry.

Incorporation of aptamer in mixed Langmuir-Blodgett films between lipid and graphene oxide sheets

Abstract Interaction between different molecules during formation of complex nanostructured films, which can be used as matrixes for fixing of macromolecules, plays important role in formation of sensitive films for biosensors. The Langmuir-Blodgett technique was used to obtain mixed films of dimethyldioctadecylammonium bromide (DODAB) and graphene oxide sheets (GO). Specific aptamers were subsequently attached. The obtained results after characterization of films on the air/water interface show that incorporation of GO sheets between lipid monolayers takes place. Transfer rates for mixed films indicates the formation of Z-type films after GO incorporation. Quartz Crystal Microbalance measurements of films density clearly demonstrated the presence of GO sheets in films. After drop casting method with the solution of aptamer marked with Cy5 fluorophores on the surface of the mixed films, measurements of fluorescence microscopy and contact angle were performed demonstrating the presence of these macromolecules even after cleaning procedures.

Probing relaxation dynamics of a cationic lipid based non-viral carrier: a time-resolved fluorescence study.

Lipid vesicles composed of cationic dioctadecyldimethylammonium bromide (DODAB) and neutral 1-monooleoyl-rac-glycerol (MO) are promising non-viral carriers of nucleic acids for delivery into cells. Among them, higher cell transfection efficiency was displayed by DODAB-rich vesicles than those enriched with MO. Structural relaxation of these mixed lipid vesicles plays a key role in their cell transfection efficiency because structural organization of the DODAB-rich vesicles are different from that of the MO-rich vesicles. Polarization-gated anisotropy in conjunction with picosecond resolved emission transients of a novel fluorophore 6-acetyl-(2-((4-hydroxycyclohexyl)(methyl)amino)naphthalene) (ACYMAN) has been employed to probe relaxation dynamics in pure DODAB vesicles, and in mixed vesicles of DODAB with varying content of MO. Both orientational relaxation of ACYMAN and relaxation dynamics of its local environment are retarded significantly in mixed lipid vesicles with increasing MO content, from a mole fraction (χMO) of 0.2 to that of 0.8 which is consistent with increased rigidity of the MO-rich (χMO > 0.5) vesicles relative to the DODAB-rich (χMO < 0.5) vesicles. Therefore, higher structural rigidity of the MO-rich vesicles (χMO > 0.5) gives rise to their lower cell transfection efficiency than the more flexible DODAB-rich (χMO < 0.5) vesicles as observed in previous in vivo studies (Biochim. Biophys. Acta, Biomembr., 2014, 1838, 2555–2567).

Short communication: Activity of nisin, lipid bilayer fragments and cationic nisin-lipid nanoparticles against multidrug-resistant Staphylococcus spp. isolated from bovine mastitis

Staphylococci are the main etiological agents of bovine mastitis. Bacteriocins and nanoparticles have emerged as promising alternatives for the future development of antimicrobial agents. This study evaluated the activity of the bacteriocin nisin and bicelles of the synthetic cationic lipid dioctadecyldimethylammonium bromide, alone and in combination, against multidrug-resistant Staphylococcus spp. strains isolated from bovine mastitis. In summary, cationic nisin/dioctadecyldimethylammonium bromide nanoparticles are shown to be a promising alternative for the control of mastitis caused by multidrug-resistant Staphylococcus spp.

Effects of NSAIDs on the Dynamics and Phase Behavior of DODAB Bilayers.

Despite well-known side effects, nonsteroidal anti-inflammatory drugs (NSAIDs) are one of the most prescribed drugs worldwide for their anti-inflammatory and antipyretic properties. Here, we report the effects of two NSAIDs, aspirin and indomethacin, on the thermotropic phase behavior and the dynamics of a dioctadecyldimethylammonium bromide (DODAB) lipid bilayer as studied using neutron scattering techniques. Elastic fixed window scans showed that the addition of aspirin and indomethacin affects the phase behavior of a DODAB bilayer in both heating and cooling cycles. Upon heating, there is a change in the coagel- to fluid-phase transition temperature from 327 K for pure DODAB bilayer to 321 and 323 K in the presence of aspirin and indomethacin, respectively. More strikingly, upon cooling, the addition of NSAIDs suppresses the formation of the intermediate gel phase observed in pure DODAB. The suppression of the gel phase on addition of the NSAIDs evidences the synchronous ordering of a lipid headgroup and chain. Analysis of quasi-elastic neutron scattering data showed that only localized internal motion exists in the coagel phase, whereas both internal and lateral motions exist in the fluid phase. The internal motion is described by a fractional uniaxial rotational diffusion model in the coagel phase and by a localized translation diffusion model in the fluid phase. In the coagel phase, the rotational diffusion coefficient of DODAB is found to be almost twice for the addition of the drugs, whereas the mobility fraction did not change for indomethacin but becomes twice for aspirin. In the fluid phase, the lateral motion, described well by a continuous diffusion model, is found to be slower by about ∼30% for indomethacin but almost no change for aspirin. For the internal motion, addition of aspirin leads to enhancement of the internal motion, whereas indomethacin did not show significant effect. This study shows that the effect of different NSAIDs on the dynamics of the lipid membrane is not the same; hence, one must consider these NSAIDs individually while studying their action mechanism on the cell membrane.

Dual targeted magnetic photosensitive liposomes for photothermal/photodynamic tumor therapy

Abstract In this work, we prepared a nano-vehicle for dual targeted (magnetic and ligand) and dual mode (photothermal/photodynamic) cancer therapy. For this purpose, magnetic photosensitive liposomes (MPLs) from 1,2-distearoyl-sn-glycero-3-phosphocholine, dimethyldioctadecyl ammonium bromide (DDAB) and cholesterol were prepared using solvent evaporation/hydration technique to encapsulate the photosensitizer indocyanine green (ICG) and citric acid-coated MNPs (CMNPs). Hyaluronic acid-polyethylene glycol (HA-PEG) was coated to the MPLs by self-assembly of HA-PEG on liposome surface through ionic interactions between negatively charged HA and positively charged lipid DDAB to fabricate HA-PEG-MPLs with ∼220 nm particle size. The HA-PEG-MPLs aqueous solution showed highly efficient photothermal effects as the solution temperature reaches 45 °C in 3 min after exposure to near infrared (NIR) 808 nm laser at 2 W/cm2. The in vitro cell culture experiments after treating human glioblastoma cells (U-87MG) with HA-PEG-MPLs confirmed enhanced cytotoxicity after 4 min exposure to NIR laser. A xenograft tumor model from subcutaneously implanted U87MG cells in nude mice demonstrated accumulation of HA-PEG-MPLs at tumor sites. By combination with successive NIR laser treatment, tumor growth could be prevented and the tumor volume at the end of treatment was 12.7% that of the control. Excellent and consistent anti-tumor efficacy with laser + HA-PEG-MPLs treatment was also demonstrated from the survival rates of animals, in vivo bioluminescence imaging and histology of tumor sections.

Lipid conjugation of TLR7 agonist Resiquimod ensures co-delivery with the liposomal Cationic Adjuvant Formulation 01 (CAF01) but does not enhance immunopotentiation compared to non-conjugated Resiquimod+CAF01

Pattern recognition receptors, including the Toll-like receptors (TLRs), are important in the induction and activation of two critical arms of the host defence to pathogens and microorganisms: the rapid innate immune response (as characterised by the production of Th1 promoting cytokines and type 1 interferons) and the adaptive immune response. Through this activation, ligands and agonists of TLRs can enhance immunotherapeutic efficacy. Resiquimod is a small (water-soluble) agonist of the endosome-located Toll-like receptors 7 and 8 (TLR7/8). However due to its molecular attributes it rapidly distributes throughout the body after injection. To circumvent this, these TLR agonists can be incorporated within delivery systems, such as liposomes, to promote the co-delivery of both antigen and agonists to antigen presenting cells. In this present study, resiquimod has been chemically conjugated to a lipid to form a lipid-TLR7/8 agonist conjugate which can be incorporated within immunogenic cationic liposomes composed of dimethyldioctadecylammonium bromide (DDA) and the immunostimulatory glycolipid trehalose 6,6′ – dibehenate (TDB). This DDA:TDB-TLR7/8 formulation offers similar vesicle characteristics to DDA:TDB (size and charge) and offers high retention of both resiquimod and the electrostatically adsorbed TB subunit antigen Ag85B-ESAT6-Rv2660c (H56). Following immunisation through the intramuscular (i.m.) route, these cationic DDA:TDB-TLR7/8 liposomes form a vaccine depot at the injection site. However, immunisation studies have shown that this biodistribution does not translate into notably increased antibody nor Th1 responses at the spleen and draining popliteal lymph node compared to DDA:TDB liposomes. This work demonstrates that the conjugation of TLR7/8 agonists to cationic liposomes can promote co-delivery but the immune responses stimulated do not merit the added complexity considerations of the formulation.

Antimicrobial Coatings from Hybrid Nanoparticles of Biocompatible and Antimicrobial Polymers.

Hybrid nanoparticles of poly(methylmethacrylate) synthesized in the presence of poly (diallyldimethyl ammonium) chloride by emulsion polymerization exhibited good colloidal stability, physical properties, and antimicrobial activity but their synthesis yielded poor conversion. Here we create antimicrobial coatings from casting and drying of the nanoparticles dispersions onto model surfaces such as those of silicon wafers, glass coverslips, or polystyrene sheets and optimize conversion using additional stabilizers such as cetyltrimethyl ammonium bromide, dioctadecyldimethyl ammonium bromide, or soybean lecithin during nanoparticles synthesis. Methodology included dynamic light scattering, determination of wettability, ellipsometry of spin-coated films, scanning electron microscopy, and determination of colony forming unities (log CFU/mL) of bacteria after 1 h interaction with the coatings. The additional lipids and surfactants indeed improved nanoparticle synthesis, substantially increasing the conversion rates by stabilizing the monomer droplets in dispersion during the polymerization. The coatings obtained by spin-coating or casting of the nanoparticles dispersions onto silicon wafers were hydrophilic with contact angles increasing with the amount of the cationic polymer in the nanoparticles. Against Escherichia coli and Staphylococcus aureus, bacteria cell counts were reduced by approximately 7 logs upon interaction with the coatings, revealing their potential for several biotechnological and biomedical applications.

Solid separation from the heavy oil sludge produced from Liaohe Oilfield

Heavy oil sludge usually exists in the form of stable W/O emulsion, in which the water droplets were stabilized by natural surfactants and small solid particles. The oil recovery from the oil sludge is valuable, but the demulsification by using regular demulsifiers could be hindered by the solids. Therefore, the solid separation is important for oil recovery. In this paper, firstly, properties of heavy oil sludge produced from the Liaohe Oilfield were analyzed. Then, a multistage chemical treatment was proposed and chemicals were screened for solid separation. Limonene, sodium sulphate solution, (dioctadecyl dimethyl ammonium bromide + citric acid) were chosen for different purposes at different stages, respectively. At last, the rational separation conditions were investigated. After the treatment, the solid content in oil decreased from 18.57% to be 0.9% by mass. In addition, the mixture of recovered oil and fresh crude oil can be dehydrated easily by using regular demulsifiers.

Design of Bacterial Inclusion Bodies as Antigen Carrier Systems

AbstractSynthetic subunit vaccines hold great promise to prevent infectious diseases, but they often induce only weak immune responses. Here bioengineering techniques to develop immunogenic antigen particles comprised of self‐assembling antigenic peptides are exploited. A bioparticle platform is developed to present repetitive copies of subunit antigens, which mimic the host‐pathogen surface interaction to provide enhanced immunogenicity. Herein, mycobacterial fusion proteins H4 (consisting of Ag88B‐TB10.4) and H28 (containing Ag88B‐TB10.4‐Rv2660c) are bioengineered to assemble H4 and/or H28 antigens into particulate vaccines inside an endotoxin‐free Escherichia coli strain, to serve as a novel vaccine against tuberculosis (TB). A bioprocess for particle isolation is developed. The physicochemical properties of the particles are investigated. Antigen particles formulated with the adjuvant dimethyl dioctadecyl ammonium bromide are used to subcutaneously immunize C57BL/6 mice. Both soluble and particulate vaccines elicit production of functional and specific antibodies, which only interact with target antigens and not production host cell proteins. Particulate vaccines induce strong production of interferon gamma (IFNγ) and interleukin 17A (IL17A) cytokines. When compared to soluble antigens, the antigen particle vaccines show increased immunogenicity. Overall, this study provides proof of concept that selected antigens can be engineered to self‐assemble into inclusion bodies to serve as particulate vaccines with improved immunological properties compared to the respective soluble antigens.

Modulation of Solvation and Molecular Recognition of a Lipid Bilayer under Dynamical Phase Transition.

It is well accepted in contemporary biology that an ∼30 Å thick lipid bilayer film around living cells is a matter of life and death as the film typically delimits the environments that serve as a crucial margin. The dynamic organization of lipid molecules both across the lipid bilayer and in the lateral dimension are known to be crucial for cellular transport and molecular recognition by important biological macromolecules. Here, we study dilute (20 mM) Dioctadecyldimethylammonium bromide (DODAB) vesicles at different temperatures in aqueous dispersion with well-defined phases namely liquid crystalline, gel and subgel. The spectroscopic studies on two fluorescent probes 8-anilino-1-naphthalene sulfonic acid ammonium salt (ANS) and Coumarin 500 (C500), former in the head group region of the lipid-water interface and later located deeper in the lipid bilayer follow dynamics (solvation and fluidity) of their local environments in the vesicles. Binding of an anti-tuberculosis drug rifampicin has also been studied employing Förster resonance energy transfer (FRET) technique. The molecular insight concerning the effect of dynamical organization of the lipid molecules on the local dynamics of aqueous environments in different phases leading to molecular recognition becomes evident in our study.

Heterogeneity in Dynamics of Dioctadecyldimethylammonium Bromide Bilayers: Molecular Dynamics Simulation and Neutron Scattering Study

Molecular dynamics (MD) simulations have been carried out to investigate the structural and dynamical aspects of dioctadecyldimethylammonium bromide (DODAB) bilayer which exists in various phases. ...

High potency of lipid conjugated TLR7 agonist requires nanoparticulate or liposomal formulation

Conjugation of small molecule agonists of Toll-like receptor 7 (TLR7) to proteins, lipids, or polymers is known to modulate potency, and the physical form or formulation of these conjugates is likely to have a major effect on their immunostimulatory activity. Here, we studied the effect of formulation on potency of a 1,2‑di‑(9Z‑octadecenoyl)‑sn‑glycero‑3‑phosphoethanolamine (DOPE) conjugated TLR7 agonist (DOPE-TLR7a) alongside assessing physical form using Dynamic Light Scattering (DLS), Nanosight Particle Tracking (NTA) analysis and Small Angle X-ray Scattering (SAXS). A very high potency of DOPE-TLR7a conjugate (EC50 around 9 nM) was observed either when prepared by direct dilution from DMSO or when formulated into 400–700 nm large multilamella liposomes containing dimethyldioctadecylammonium bromide salt (DDA) and DOPE. When prepared by dissolution in DMSO followed by dilution in aqueous culture medium, 93 ± 5 nm nanoparticles were formed. Without dilution from solution in DMSO, no nanoparticles were observed and no immunostimulatory activity could be detected without this formulation step. SAXS analysis of the conjugate after DMSO dissolution/water dilution revealed a lamellar order with a layer spacing of 68.7 Å, which correlates with arrangement in groups of 3 bilayers. The addition of another immunostimulatory glycolipid, trehalose‑6,6‑dibehenate (TDB), to DOPE:DDA liposomes gave no further increase in immunostimulatory activity beyond that provided by incorporating DOPE-TLR7a. Given the importance of nanoparticle or liposomal formulation for activity, we conclude that the major mechanism for increased potency when TLR7 agonists are conjugated to macromolecules is through alteration of physical form.

Induction of Cytotoxic T-Lymphocyte Responses Upon Subcutaneous Administration of a Subunit Vaccine Adjuvanted With an Emulsion Containing the Toll-Like Receptor 3 Ligand Poly(I:C)

There is an unmet medical need for new subunit vaccines that induce cytotoxic T-lymphocyte (CTL) responses to prevent infection with a number of pathogens. However, stimulation of CTL responses via clinically acceptable subcutaneous (s.c.) and intramuscular (i.m.) injection is challenging. Recently, we designed a liposomal adjuvant [cationic adjuvant formulation (CAF)09] composed of the cationic lipid dimethyldioctadecylammonium (DDA) bromide, a synthetic monomycoloyl glycerol analog and polyinosinic:polycytidylic acid, which induce strong CTL responses to peptide and protein antigens after intraperitoneal administration. By contrast, CAF09 does not stimulate CTL responses upon s.c. or i.m. injection because the vaccine forms a depot that remains at the injection site. Hence, we engineered a series of nanoemulsions (CAF24a–c) based on the active components of CAF09. The oil phase consisted of biodegradable squalane, and the surface charge was varied systematically by replacing DDA with zwitterionic distearoylphosphoethanolamine. We hypothesized that the nanoemulsions drain to the lymph nodes to a larger extent than CAF09, upon s.c. co-administration with the model antigen chicken egg ovalbumin (OVA). This results in an increased dose fraction that reaches the draining lymph nodes (dLNs) and subsequently activates cross-presenting dendritic cells (DCs), which can prime CTL responses. Indeed, the nanoemulsions induced antigen-specific CD8+ T-cell responses, which were significantly higher than those stimulated by OVA adjuvanted with CAF09. We explain this by the observed rapid localization of CAF24a in the dLNs and the subsequent association with conventional DCs, which promotes induction of CTL responses. Uptake of CAF24a was not specific for DCs, because CAF24a was also detected with B cells and macrophages. No measurable dose fraction of CAF09 was detected in the dLNs within the study period, and CAF09 formed a depot at the site of injection. Importantly, s.c. vaccination with OVA adjuvanted with CAF24a induced significant levels of specific lysis of antigen-pulsed splenocytes were induced after, which was not observed for OVA adjuvanted with CAF09. Thus, CAF24a is a promising adjuvant for induction of CTL responses upon s.c. and i.m. immunization, and it offers interesting perspectives for the design of vaccines against pathogens for which CTL responses are required to prevent infection.

Interaction of Hydrophobic Ionic Liquids with Lipids in Langmuir Monolayers.

The interaction of two ionic liquids, trihexyl(tetradecyl)phosphonium bis(trifluoromethylsulfonyl)-imide and trihexyl(tetradecyl)phosphonium dicyanamide, [P66614][Ntf2] and [P66614]/[N(CN)2], with several long-chained lipids with a different net charge at the hydrophilic group, a cationic surfactant, dioctadecyldimethylammonium bromide (DODAB), a zwitterionic phospholipid (DPPC), an anionic phospholipid (DPPG), and the neutral stearic acid (SA), was investigated at the air-water interface using the Langmuir trough technique. The experimental surface pressure-area (π- A) isotherms obtained for selected compositions of each binary system reveal distinct interfacial behavior. The degree and the nature of the IL-lipid interaction strongly depend on the charge distribution in the lipid polar group. Miscibility, or immiscibility, at the monolayer was inferred from the comparison of the experimental π- A isotherm with the theoretical curve calculated for the corresponding ideal mixture based on the π- A isotherms of the pure components. Phase separation and partial miscibility occurred in IL/DODAB and IL/DPPC systems, respectively. In both the IL/DPPG and the IL/SA systems, a new catanionic complex was found. For the IL/SA system, the catanionic complex formation varies with the subphase pH.

Carbon Nanotubes and Algal Polysaccharides To Enhance the Enzymatic Properties of Urease in Lipid Langmuir-Blodgett Films.

Algal polysaccharides (extracellular polysaccharides) and carbon nanotubes (CNTs) were adsorbed on dioctadecyldimethylammonium bromide Langmuir monolayers to serve as a matrix for the incorporation of urease. The physicochemical properties of the supramolecular system as a monolayer at the air-water interface were investigated by surface pressure-area isotherms, surface potential-area isotherms, interfacial shear rheology, vibrational spectroscopy, and Brewster angle microscopy. The floating monolayers were transferred to hydrophilic solid supports, quartz, mica, or capacitive electrolyte-insulator-semiconductor (EIS) devices, through the Langmuir-Blodgett (LB) technique, forming mixed films, which were investigated by quartz crystal microbalance, fluorescence spectroscopy, and field emission gun scanning electron microscopy. The enzyme activity was studied with UV-vis spectroscopy, and the feasibility of the thin film as a urea sensor was essayed in an EIS sensor device. The presence of CNT in the enzyme-lipid LB film not only tuned the catalytic activity of urease but also helped to conserve its enzyme activity. Viability as a urease sensor was demonstrated with capacitance-voltage and constant capacitance measurements, exhibiting regular and distinctive output signals over all concentrations used in this work. These results are related to the synergism between the compounds on the active layer, leading to a surface morphology that allowed fast analyte diffusion owing to an adequate molecular accommodation, which also preserved the urease activity. This work demonstrates the feasibility of employing LB films composed of lipids, CNT, algal polysaccharides, and enzymes as EIS devices for biosensing applications.

Dynamical Transitions and Diffusion Mechanism in DODAB Bilayer

Dioctadecyldimethylammonium bromide (DODAB), a potential candidate for applications in drug transport or DNA transfection, forms bilayer in aqueous media exhibiting a rich phase behavior. Here, we report the detailed dynamical features of DODAB bilayer in their different phases (coagel, gel and fluid) as studied by neutron scattering techniques. Elastic intensity scans show dynamical transitions at 327 K in the heating and at 311 K and 299 K during cooling cycle. These results are consistent with calorimetric studies, identified as coagel-fluid phase transition during heating, and fluid-gel and gel-coagel phase transitions during cooling. Quasielastic Neutron Scattering (QENS) data analysis showed presence of only localized internal motion in the coagel phase. However, in the gel and fluid phases, two distinct motions appear, namely lateral motion of the DODAB monomers and a faster localized internal motion of the monomers. The lateral motion of the DODAB molecule is described by a continuous diffusion model and is found to be about an order of magnitude slower in the gel phase than in the fluid phase. To gain molecular insights, molecular dynamics simulations of DODAB bilayer have also been carried out and the results are found to be in agreement with the experiment.

Dioctadecyldimethylammonium bromide (DODAB-BF) as a new adjuvant for maternal-fetal immunization in mice against Neisseria meningitidis: evaluation of humoral response.

Neisseria meningitidis bacterium is a Gram-negative diplococcus. Among their serogroups, the B is one of the main causes of invasive meningococcal disease. Newborns and children are particularly susceptible to this infection because of their immune systems that are still maturing and relatively inexperienced. Thus, further studies on the use of maternal immunization for protection against this disease are needed. The purpose of this study was to evaluate the potential immunogenic antigens from the outer membrane of N. meningitidis serogroup B in outbred mice and the influence of maternal immunization in the offspring, and analyze the adjuvant effect of bilayer fragments of dioctadecyldimethylamonium bromide (DODAB-BF) and hydroxide aluminium (alum) in enhancing antibodies production and transference to offspring. IgG and IgG1, IgG2a and IgG2b subclasses of antibodies in serum from immunized mice and controls were quantified and compared. Immunization by subcutaneous and intramuscular routes exhibited evidence of IgG, and both adjuvants promoted the production of IgG1 and IgG2b that were transferred to the offspring. These antibodies also showed specificity with the outer membrane vesicles from homologous strain and were capable to cross react with different strains. The use of DODAB-BF seems to enhance immune response on mothers and offspring and may have immunological advantages.

Application of Pharmacokinetics Modelling to Predict Human Exposure of a Cationic Liposomal Subunit Antigen Vaccine System.

The pharmacokinetics of a liposomal subunit antigen vaccine system composed of the cationic lipid dimethyldioctadecylammonium bromide (DDA) and the immunostimulatory agent trehalose 6,6-dibehenate (TDB) (8:1 molar ratio) combined with the Ag85B-ESAT-6 (H1) antigen were modelled using mouse in-vivo data. Compartment modelling and physiologically based pharmacokinetics (PBPK) were used to predict the administration site (muscle) and target site (lymph) temporal concentration profiles and factors governing these. Initial estimates using compartmental modelling established that quadriceps pharmacokinetics for the liposome demonstrated a long half-life (22.6 days) compared to the associated antigen (2.62 days). A mouse minimal-PBPK model was developed and successfully predicted quadriceps liposome and antigen pharmacokinetics. Predictions for the popliteal lymph node (PLN) aligned well at earlier time-points. A local sensitivity analysis highlighted that the predicted AUCmuscle was sensitive to the antigen degradation constant kdeg (resulting in a 3-log change) more so than the fraction escaping the quadriceps (fe) (resulting in a 10-fold change), and the predicted AUCPLN was highly sensitive to fe. A global sensitivity analysis of the antigen in the muscle demonstrated that model predictions were within the 50th percentile for predictions and showed acceptable fits. To further translate in-vitro data previously generated by our group, the mouse minimal-PBPK model was extrapolated to humans and predictions made for antigen pharmacokinetics in muscle and PLN. Global analysis demonstrated that both kdeg and fe had a minimal impact on the resulting simulations in the muscle but a greater impact in the PLN. In summary, this study has predicted the in-vivo fate of DDA:TDB:H1 in humans and demonstrated the roles that formulation degradation and fraction escaping the depot site can play upon the overall depot effect within the site of administration.

A strong adjuvant based on glycol-chitosan-coated lipid-polymer hybrid nanoparticles potentiates mucosal immune responses against the recombinant Chlamydia trachomatis fusion antigen CTH522

Induction of mucosal immunity with vaccines is attractive for the immunological protection against pathogen entry directly at the site of infection. An example is infection with Chlamydia trachomatis (Ct), which is the most common sexually transmitted infection in the world, and there is an unmet medical need for an effective vaccine. A vaccine against Ct should elicit protective humoral and cell-mediated immune (CMI) responses in the genital tract mucosa. We previously designed an antibody- and CMI-inducing adjuvant based on poly(dl-lactic-co-glycolic acid) (PLGA) nanoparticles modified with the cationic surfactant dimethyldioctadecylammonium bromide and the immunopotentiator trehalose-6,6′-dibehenate. Here we show that immunization with these lipid-polymer hybrid nanoparticles (LPNs) coated with the mucoadhesive polymer chitosan enhances mucosal immune responses. Glycol chitosan (GC)-modified LPNs were engineered using an oil-in-water single emulsion solvent evaporation method. The nanoparticle design was optimized in a highly systematic way by using a quality-by-design approach to define the optimal operating space and to gain maximal mechanistic information about the GC coating of the LPNs. Cryo-transmission electron microscopy revealed a PLGA core coated with one or several concentric lipid bilayers. The GC coating of the surface was identified as a saturable, GC concentration-dependent increase in particle size and a reduction of the zeta-potential, and the coating layer could be compressed upon addition of salt. Increased antigen-specific mucosal immune responses were induced in the lungs and the genital tract with the optimized GC-coated LPN adjuvant upon nasal immunization of mice with the recombinant Ct fusion antigen CTH522. The mucosal responses were characterized by CTH522-specific IgG/IgA antibodies, together with CTH522-specific interferon γ-producing Th1 cells. This study demonstrates that mucosal administration of CTH522 adjuvanted with chitosan-coated LPNs represents a promising strategy to modulate the magnitude of mucosal vaccine responses.