Systematic screening of photopolymer resins for stereolithography (SLA) 3D printing of solid oral dosage forms: Investigation of formulation factors on printability outcomes.

Pharmaceutical three-dimensional printing (3DP) is now in its golden age. Recent years have seen a dramatic increase in the research in 3D printed pharmaceuticals due to their potential to deliver highly personalised medicines, thus revolutionising the way medicines are designed, manufactured, and dispensed. A particularly attractive 3DP technology used to manufacture medicines is stereolithography (SLA), which features key advantages in terms of printing resolution and compatibility with thermolabile drugs. Nevertheless, the enthusiasm for pharmaceutical SLA has not been followed by the introduction of novel excipients specifically designed for the fabrication of medicines; hence, the choice of biocompatible polymers and photoinitiators available is limited. This work provides an insight on how to maximise the usefulness of the limited materials available by evaluating how different formulation factors affect printability outcomes of SLA 3D printed medicines. 156 photopolymer formulations were systematically screened to evaluate the influence of factors including photoinitiator amount, photopolymer molecular size, and type and amount of liquid filler on the printability outcomes. Collectively, these factors were found highly influential in modulating the print quality of the final dosage forms. Findings provide enhanced understanding of formulation parameters informing the future of SLA 3D printed medicines and the personalised medicines revolution.

Virtual Clinical Trials Guided Design of an Age-Appropriate Formulation and Dosing Strategy of Nifedipine for Paediatric Use.

The rapid onset of action of nifedipine causes a precipitous reduction in blood pressure leading to adverse effects associated with reflex sympathetic nervous system (SNS) activation, including tachycardia and worsening myocardial and cerebrovascular ischemia. As a result, short acting nifedipine preparations are not recommended. However, importantly, there are no modified release preparations of nifedipine authorised for paediatric use, and hence a paucity of clinical studies reporting pharmacokinetics data in paediatrics. Pharmacokinetic parameters may differ significantly between children and adults due to anatomical and physiological differences, often resulting in sub therapeutic and/or toxic plasma concentrations of medication. However, in the field of paediatric pharmacokinetics, the use of pharmacokinetic modelling, particularly physiological-based pharmacokinetics (PBPK), has revolutionised the ability to extrapolate drug pharmacokinetics across age groups, allowing for pragmatic determination of paediatric plasma concentrations to support drug licensing and clinical dosing. In order to pragmatically assess the translation of resultant dissolution profiles to the paediatric populations, virtual clinical trials simulations were conducted. In the context of formulation development, the use of PBPK modelling allowed the determination of optimised formulations that achieved plasma concentrations within the target therapeutic window throughout the dosing strategy. A 5 mg sustained release mini-tablet was successfully developed with the duration of release extending over 24 h and an informed optimised dosing strategy of 450 µg/kg twice daily. The resulting formulation provides flexible dosing opportunities, improves patient adherence by reducing frequent administration burden and enhances patient safety profiles by maintaining efficacious levels of consistent drug plasma levels over a sustained period of time.

Stereolithography Apparatus Evolution: Enhancing Throughput and Efficiency of Pharmaceutical Formulation Development.

Pharmaceutical applications of 3D printing technologies are growing rapidly. Among these, vat photopolymerisation (VP) techniques, including Stereolithography (SLA) hold much promise for their potential to deliver personalised medicines on-demand. SLA 3D printing offers advantageous features for pharmaceutical production, such as operating at room temperature and offering an unrivaled printing resolution. However, since conventional SLA apparatus are designed to operate with large volumes of a single photopolymer resin, significant throughput limitations remain. This, coupled with the limited choice of biocompatible polymers and photoinitiators available, hold back the pharmaceutical development using such technologies. Hence, the aim of this work was to develop a novel SLA apparatus specifically designed to allow rapid and efficient screening of pharmaceutical photopolymer formulations. A commercially available SLA apparatus was modified by designing and fabricating a novel resin tank and build platform able to 3D print up to 12 different formulations at a single time, reducing the amount of sample resin required by 20-fold. The novel SLA apparatus was subsequently used to conduct a high throughput screening of 156 placebo photopolymer formulations. The efficiency of the equipment and formulation printability outcomes were evaluated. Improved time and cost efficiency by 91.66% and 94.99%, respectively, has been confirmed using the modified SLA apparatus to deliver high quality, highly printable outputs, thus evidencing that such modifications offer a robust and reliable tool to optimize the throughput and efficiency of vat photopolymerisation techniques in formulation development processes, which can, in turn, support future clinical applications.

Literature review of medication administration problems in paediatrics by parent/caregiver and the role of health literacy.

OBJECTIVE: To identify studies that highlighted medication administration problems experienced by parents and children, which also looked at health literacy aspect using a validated tool to assess for literacy. STUDY DESIGN: Ten electronic databases were systematically searched and supplemented by hand searching through reference lists using the following search terms: (1) paediatric, (2) medication error including dosing error, medication administration error, medication safety and medication optimisation and (3) health literacy. RESULTS: Of the (1230) records screened, 14 studies were eligible for inclusion. Three analytical themes emerged from the synthesis. The review highlighted that frequencies and magnitudes of dosing errors vary by the measurement tools used, the dose prescribed and by the administration instruction provided. Parent's sociodemographic, such as health literacy and language, is a key factor to be considered when designing an intervention aimed at averting medication administration errors at home. The review summarised some potential strategies that could help in reducing medication administration errors among children at home. Among these recommendations is to show the prescribed dose to the parents or young people along with the verbal instructions, as well as to match the prescribed dose with the measuring tool dispensed, to provide an explicit dose intervals and pictographic dosing instructions. CONCLUSION: The findings suggest that in order to optimise medication use by parents, further work is needed to address the nature of these issues at home. Counselling, medication administration instructions and measurement tools are some of the areas in addition to the sociodemographic characteristics of parents and young people that need to be considered when designing any future potential intervention aimed at reducing medication errors among children and young people at home.

Does the Formulation of Oral Solid Dosage Forms Affect Acceptance and Adherence in Older Patients? A Mixed Methods Systematic Review.

OBJECTIVES: Age-related changes mean that the older population can encounter barriers toward taking medication orally. Further work is needed to identify the characteristics of oral solid dosage forms that will improve patient acceptance and adherence. The aim of this systematic review was to identify if and how formulation aspects of oral solid dosage forms affect acceptance and adherence in older people. DESIGN: Mixed methods systematic review using a data-based convergent synthesis design. SETTING AND PARTICIPANTS: Articles were selected if they included participants aged 60 years and older, or included health care professionals, social care professionals, and informal carers of patients aged 60 years and older. METHODS: A systematic search of the following databases was undertaken: Web of Science, MEDLINE, Scopus, and The Cochrane Databases. The search of databases was supplemented by a search of gray literature, and reference lists of included papers were manually searched. RESULTS: A total of 16 studies were included in the final synthesis. Three themes were generated from the thematic analysis: (1) dimensions, (2) palatability, and (3) appearance. The dimensions and palatability are often modified to improve swallowability by breaking tablets in half or taste masking with food. Polypharmacy can lead to patients using the appearance to identify tablets; however, this can lead to confusion when products appear similar. No study was identified that explored formulation characteristics across all 3 categories directly in the older population. CONCLUSION AND IMPLICATIONS: Manufacturers should take into account practical problems older people may encounter when considering the dimensions, palatability, and appearance of the final drug product. These characteristics should be optimized to aid visual identification and swallowability. Medical providers and pharmacists have an important role in ensuring that these patient-centric drug products are prescribed and dispensed appropriately so that patients receive the most suitable formulation.

Anti-RSV Peptide-Loaded Liposomes for the Inhibition of Respiratory Syncytial Virus.

Although respiratory syncytial virus (RSV) is one of the leading causes of acute respiratory tract infection in infants and adults, effective treatment options remain limited. To circumvent this issue, there is a novel approach, namely, the development of multifunctional liposomes for the delivery of anti RSV-peptides. While most of the peptides that are used for loading with the particulate delivery systems are the penetrating peptides, an alternative approach is the development of liposome-peptide systems, which are loaded with an RSV fusion peptide (RF-482), which has been designed to inhibit the RSV fusion and block infection. The results of this work have revealed that the liposomes themselves can serve as potential RSV inhibitors, whilst the anti-RSV-peptide with liposomes can significantly increase the RSV inhibition when compared with the anti-RSV peptide alone.

Activated carbon as a carrier for amorphous drug delivery: Effect of drug characteristics and carrier wettability.

Recent research on porous silica materials as drug carriers for amorphous and controlled drug delivery has shown promising results. However, due to contradictory literature reports on toxicity and high costs of production, it is important to explore alternative safe and inexpensive porous carriers. In this study, the potential of activated carbon (AC) as an amorphous drug carrier was investigated using paracetamol (PA) and ibuprofen (IBU) as model drugs. The solution impregnation method was used for drug loading, with loading efficiency determined by UV spectroscopy and drug release kinetics studied using USP II dissolution apparatus. The physical state of the drug in the complex was characterised using differential scanning calorimetry and X-ray diffractions techniques, whilst sites of drug adsorption were studied using Fourier transform infrared spectroscopy and N2 adsorption techniques. In addition, the cytotoxicity of AC on human colon carcinoma (Caco-2) cells was assessed using the MTT assay. Results presented here reveal that, for PA/AC and IBU/AC complexes, the saturation solubility of the drug in the loading solvent appears to have an effect on the drug loading efficiency and the physical state of the drug loaded, whilst drug release kinetics were affected by the wettability of the activated carbon particles. Furthermore, activated carbon microparticles exhibited very low cytotoxicity on Caco-2 cells at the concentrations tested (10-800µg/mL). This study, therefore, supports the potential of activated carbon as a carrier for amorphous drug delivery.

Environmental Scanning Electron Microscope Imaging of Vesicle Systems.

The structural characteristics of liposomes have been widely investigated and there is certainly a strong understanding of their morphological characteristics. Imaging of these systems, using techniques such as freeze-fracturing methods, transmission electron microscopy, and cryo-electron imaging, has allowed us to appreciate their bilayer structures and factors which can influence this. However, there are few methods which all us to study these systems in their natural hydrated state; commonly the liposomes are visualized after drying, staining, and/or fixation of the vesicles. Environmental Scanning Electron Microscopy (ESEM) offers the ability to image a liposome in its hydrated state without the need for prior sample preparation. Within our studies we were the first to use ESEM to study liposomes and niosomes and we have been able to dynamically follow the hydration of lipid films and changes in liposome suspensions as water condenses on to, or evaporates from, the sample in real time. This provides insight into the resistance of liposomes to coalescence during dehydration, thereby providing an alternative assay of liposome formulation and stability.

Microfluidics based manufacture of liposomes simultaneously entrapping hydrophilic and lipophilic drugs.

Despite the substantial body of research investigating the use of liposomes, niosomes and other bilayer vesicles for drug delivery, the translation of these systems into licensed products remains limited. Indeed, recent shortages in the supply of liposomal products demonstrate the need for new scalable production methods for liposomes. Therefore, the aim of our research has been to consider the application of microfluidics in the manufacture of liposomes containing either or both a water soluble and a lipid soluble drug to promote co-delivery of drugs. For the first time, we demonstrate the entrapment of a hydrophilic and a lipophilic drug (metformin and glipizide respectively) both individually, and in combination, using a scalable microfluidics manufacturing system. In terms of the operating parameters, the choice of solvents, lipid concentration and aqueous:solvent ratio all impact on liposome size with vesicle diameter ranging from ∼90 to 300nm. In terms of drug loading, microfluidics production promoted high loading within ∼100nm vesicles for both the water soluble drug (20-25% of initial amount added) and the bilayer embedded drug (40-42% of initial amount added) with co-loading of the drugs making no impact on entrapment efficacy. However, co-loading of glipizide and metformin within the same liposome formulation did impact on the drug release profiles; in both instances the presence of both drugs in the one formulation promoted faster (up to 2 fold) release compared to liposomes containing a single drug alone. Overall, these results demonstrate the application of microfluidics to prepare liposomal systems incorporating either or both an aqueous soluble drug and a bilayer loaded drug.

Developing solid particulate vaccine adjuvants: surface bound antigen favours a humoural response, whereas entrapped antigen shows a tendency for cell mediated immunity.

This present study compares the efficacy of microsphere formulations, and their method of antigen presentation, for the delivery of the TB sub-unit vaccine antigen, Ag85B-ESAT-6. Microspheres based on poly(lactide-coglycolide) (PLGA) and chitosan incorporating dimethyldioctadecylammonium bromide (DDA) were prepared by either the w/o/w double emulsion method (entrapped antigen) or the o/w single emulsion method (surface bound antigen), and characterised for their physico-chemical properties and their ability to promote an immune response to Ag85B-ESAT-6. The method of preparation, and hence method of antigen association, had a pronounced effect on the type of immune response achieved from the microsphere formulations, with surface bound antigen favouring a humoural response, whereas entrapped antigen favoured a cellular response.

The role of lipid geometry in designing liposomes for the solubilisation of poorly water soluble drugs.

Liposomes are well recognised for their ability to improve the delivery of a range of drugs. More commonly they are applied for the delivery of water-soluble drugs, but given their structural attributes, they can also be employed as solubilising agents for low solubility drugs as well as drug targeting agents. To further explore the potential of liposomes as solubilising agents, we have investigated the role of bilayer packaging in promoting drug solubilisation in liposome bilayers. The effect of alkyl chain length and symmetry was investigated to consider if using 'mis-matched' phospholipids could create 'voids' within the bilayers, and enhance bilayer loading capacity. Lipid packing was investigated using Langmuir studies, which demonstrated that increasing the alkyl chain length enhanced lipid packing, with condensed monolayers forming, whilst asymmetric lipids formed less condensed monolayers. However, this more open packing did not translate into improved drug loading, with the longer chain, condensed bilayers formed from long-chain, saturated lipids offering higher drug loading capacity. These studies demonstrate that liposomes formulated from longer chain, saturated lipids offer enhanced solubilisation capacity. However the molecular size, rather than lipophilicity, of the drug to be incorporated was also a key factor dominating bilayer incorporation efficiency.

Manipulation of the surface pegylation in combination with reduced vesicle size of cationic liposomal adjuvants modifies their clearance kinetics from the injection site, and the rate and type of T cell response.

The mechanism behind the immunostimulatory effect of the cationic liposomal vaccine adjuvant dimethyldioctadecylammonium and trehalose 6,6'-dibehenate (DDA:TDB) has been linked to the ability of these cationic vesicles to promote a depot after administration, with the liposomal adjuvant and the antigen both being retained at the injection site. This can be attributed to their cationic nature, since reduction in vesicle size does not influence their distribution profile yet neutral or anionic liposomes have more rapid clearance rates. Therefore the aim of this study was to investigate the impact of a combination of reduced vesicle size and surface pegylation on the biodistribution and adjuvanticity of the formulations, in a bid to further manipulate the pharmacokinetic profiles of these adjuvants. From the biodistribution studies, it was found that with small unilamellar vesicles (SUVs), 10% PEGylation of the formulation could influence liposome retention at the injection site after 4 days, whilst higher levels (25mol%) of PEG blocked the formation of a depot and promote clearance to the draining lymph nodes. Interestingly, whilst the use of 10% PEG in the small unilamellar vesicles did not block the formation of a depot at the site of injection, it did result in earlier antibody response rates and switch the type of T cell responses from a Th1 to a Th2 bias suggesting that the presence of PEG in the formulation not only control the biodistribution of the vaccine, but also results in different types of interactions with innate immune cells.

The impact of ageing on the barriers to drug delivery.

Generally, we like to see ageing as a process that is happening to people older than ourselves. However the process of ageing impacts on a wide range of functions within the human body. Whilst many of the outcomes of ageing can now be delayed or reduced, age-related changes in cellular, molecular and physiological functionality of tissues and organs can also influence how drugs enter, distribute and are eliminated from the body. Therefore, the changing profile of barriers to drug delivery should be considered if we are to develop more age-appropriate medicines. Changes in the drug dissolution and absorption in older patients may require the formulation of oral delivery systems that offer enhanced retention at absorption sites to improve drug delivery. Alternatively, liquid and fast-melt dosage systems may address the need of patients who have difficulties in swallowing medication. Ageing-induced changes in the lung can also result in slower drug absorption, which is further compounded by disease factors, common in an ageing population, that reduce lung capacity. In terms of barriers to drug delivery to the eye, the main consideration is the tear film, which like other barriers to drug delivery, changes with normal ageing and can impact on the bioavailability of drugs delivery using eye drops and suspensions. In contrast, whilst the skin as a barrier changes with age, no significant difference in absorption of drugs from transdermal drug delivery is observed in different age groups. However, due to the age-related pharmacokinetic and pharmacodynamic changes, dose adaptation should still be considered for drug delivery across the skin. Overall it is clear that the increasing age demographic of most populations, presents new (or should that be older) barriers to effective drug delivery.

Pegylation of DDA:TDB liposomal adjuvants reduces the vaccine depot effect and alters the Th1/Th2 immune responses.

The adjuvant efficacy of cationic liposomes composed of dimethyldioctadecylammonium bromide and trehalose dibehenate (DDA:TDB) is well established. Whilst the mechanism behind its immunostimulatory action is not fully understood, the ability of the formulation to promote a 'depot effect' is a consideration. The depot effect has been suggested to be primarily due to their cationic nature which results in electrostatic adsorption of the antigen and aggregation of the vesicles at the site of injection. The aim of the study was to further test this hypothesis by investigating whether sterically stabilising DDA:TDB with polyethylene glycol (PEG) reduces aggregation, and subsequently influences the formation of a depot at the site of injection. Results reported demonstrate that high (25%) levels of PEG was able to significantly inhibit the formation of a liposome depot and also severely limit the retention of antigen at the site, resulting in a faster drainage of the liposomes from the site of injection. This change in biodistribution profile was reflected in the immunisation response, where lower levels of IgG2b antibody and IFN-γ and higher level of IL-5 cytokine were found. Furthermore entrapping antigen within DDA:TDB liposomes did not improve antigen retention at the injection site compared surface adsorbed antigen.

The application of monolayer studies in the understanding of liposomal formulations.

The study of surfactant monolayers is certainly not a new technique, but the application of monolayer studies to elucidate controlling factors in liposome design remains an underutilised resource. Using a Langmuir-Blodgett trough, pure and mixed lipid monolayers can be investigated, both for their interactions within the monolayer, and for interfacial interactions with drugs in the aqueous sub-phase. Despite these monolayers effectively being only half a bilayer, with a flat rather than curved structure, information from these studies can be effectively translated into liposomal systems. Here we outline the background, general protocols and application of Langmuir studies with a focus on their application in liposomal systems. A range of case studies are discussed which show how the system can be used to support its application in the development of liposome drug delivery. Examples include investigations into the effect of cholesterol within the liposome bilayer, understanding effective lipid packaging within the bilayer to promote water soluble and poorly soluble drug retention, the effect of alkyl chain length on lipid packaging, and drug-monolayer electrostatic interactions that promote bilayer repackaging.

Formulation design considerations for oral vaccines.

Whilst oral vaccination is a potentially preferred route in terms of patient adherence and mass vaccination, the ability to formulate effective oral vaccines remains a challenge. The primary barrier to oral vaccination is effective delivery of the vaccine through the GI tract owing to the many obstacles it presents, including low pH, enzyme degradation and bile-salt solubilization, which can result in breakdown/deactivation of a vaccine. For effective immune responses after oral administration, particulates need to be taken up bythe M cells however, these are few in number. To enhance M-cell uptake, particle characteristics can be optimized with particle size, surface charge, targeting groups and bioadhesive properties all being considerations. Yet improved uptake may not translate into enhanced immune responses and formulating particulates with inherent adjuvant properties can offer advantages. Within this article, we establish the options available for consideration when building effective oral particulate vaccines.

Solubilisation of drugs within liposomal bilayers: alternatives to cholesterol as a membrane stabilising agent.

OBJECTIVES: The aim of this work was to investigate the effect of cholesterol on the bilayer loading of drugs and their subsequent release and to investigate fatty alcohols as an alternative bilayer stabiliser to cholesterol. METHODS: The loading and release rates of four low solubility drugs (diazepam, ibuprofen, midazolam and propofol) incorporated within the bilayer of multilamellar liposomes which contained a range of cholesterol (0-33 mol/mol%) or a fatty alcohol (tetradecanol, hexadecanol and octadecanol) were investigated. The molecular packing of these various systems was also investigated in Langmuir monolayer studies. KEY FINDINGS: Loading and release of drugs within the liposome bilayer was shown to be influenced by their cholesterol content: increasing cholesterol content was shown to reduce drug incorporation and inclusion of cholesterol in the bilayer changed the release profile of propofol from zero-order, for phosphatidyl choline only liposomes, to a first-order model when 11 to 33 total molar % of cholesterol was present in the formulation. At higher bilayer concentrations substitution of cholesterol with tetradecanol was shown to have less of a detrimental impact on bilayer drug loading. However, the presence of cholesterol within the liposome bilayer was shown to reduce drug release compared with fatty alcohols. Monolayer studies undertaken showed that effective mean area per molecule for a 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC):cholesterol mixture deviated by 9% from the predicted area compared with 5% with a similar DSPC:tetradecanol mixture. This evidence, combined with cholesterol being a much more bulky structure, indicated that the condensing influence of tetradecanol was less compared with cholesterol, thus supporting the reduced impact of tetradecanol on drug loading and drug retention. CONCLUSIONS: Liposomes can be effectively formulated using fatty alcohols as an alternative bilayer stabiliser to cholesterol. The general similarities in the characteristics of liposomes containing fatty alcohols or cholesterol suggest a common behavioural influence for both compounds within the bilayer.

Increased potential of a cationic liposome-based delivery system: enhancing stability and sustained immunological activity in pre-clinical development.

The combination of dimethyl dioctadecyl ammonium bromide (DDA) and the synthetic cord factor trehalose dibehenate (TDB) with Ag85B-ESAT-6 (H1 fusion protein) has been found to promote strong protective immune responses against Mycobacterium tuberculosis. The development of a vaccine formulation that is able to facilitate the requirements of sterility, stability and generation of a vaccine product with acceptable composition, shelf-life and safety profile may necessitate selected alterations in vaccine formulation. This study describes the implementation of a sterilisation protocol and the use of selected lyoprotective agents in order to fulfil these requirements. Concomitantly, close analysis of any alteration in physico-chemical characteristics and parameters of immunogenicity have been examined for this promising DDA liposome-based tuberculosis vaccine. The study addresses the extensive guidelines on parameters for non-clinical assessment, suitable for liposomal vaccines and other vaccine delivery systems issued by the World Health Organisation (WHO) and the European Medicines Agency (EMEA). Physical and chemical stability was observed following alteration in formulations to include novel cryoprotectants and radiation sterilisation. Immunogenicity was maintained following these alterations and even improved by modification with lysine as the cryoprotective agent for sterilised formulations. Taken together, these results outline the successful alteration to a liposomal vaccine, representing improved formulations by rational modification, whilst maintaining biological activity.

Environmental scanning electron microscope imaging of vesicle systems.

The structural characteristics of liposomes have been widely investigated and there is certainly a strong understanding of their morphological characteristics. Imaging of these systems, using techniques such as freeze-fracturing methods, transmission electron microscopy, and cryo-electron imaging, has allowed us to appreciate their bilayer structures and factors that influence this. However, there are a few methods that study these systems in their natural hydrated state; commonly, the liposomes are visualized after drying, staining and/or fixation of the vesicles. Environmental scanning electron microscopy (ESEM) offers the ability to image a liposome in its hydrated state without the need for prior sample preparation. We were the first to use ESEM to study the liposomes and niosomes, and have been able to dynamically follow the hydration of lipid films and changes in liposome suspensions as water condenses onto, or evaporates from, the sample in real-time. This provides an insight into the resistance of liposomes to coalescence during dehydration, thereby providing an alternative assay for liposome formulation and stability.

Liposomes act as stronger sub-unit vaccine adjuvants when compared to microspheres.

The ability of liposomes and microspheres to enhance the efficacy of a sub-unit antigen was investigated. Microspheres were optimised by testing a range of surfactants employed in the external aqueous phase of a water-in-oil-in-water (w/o/w) double emulsion solvent evaporation process for the preparation of microspheres--composed of poly(D,L-lactide-co-glycolide) and the immunological adjuvant dimethyl dioctadecyl ammonium bromide (DDA)--and then investigated with regard to the physico-chemical and immunological characteristics of the particles produced. The results demonstrate that this parameter can affect the physico-chemical characteristics of these systems and subsequently, has a substantial bearing on the level of immune response achieved, both humoral and cell mediated, when employed for the delivery of the sub-unit tuberculosis vaccine antigen Ag85B-ESAT-6. Moreover, the microsphere preparations investigated failed to initiate immune responses at the levels achieved with an adjuvant DDA-based liposome formulation (DDA-TDB), further substantiating the superior ability of liposomes as vaccine delivery systems.