Fig. 1. General structure of liposome vaccines. The liposome
vaccines consist of a two-layer structure with added cholesterol
phospholipids, as well as pH-sensitive materials and (polyethylene
glycol) PEG modifications for versatility. The ways in which liposomes
carry antigens include encapsulation, absorption or admixed. Lipid A is
an adjuvant added in a liposome vaccine which is a lipid layer.
Preparation of liposome carriers. The properties of
liposomes are closely related to the preparation liquid. In the past,
liposome preparation techniques had similar general steps: drying from
volatile solvents, dispersion, purification, and synthesis in aqueous
medium, and finally liposomes were obtained [20].
Thin film hydration is the earliest liposome preparation technology,
which mainly includes dissolving phospholipids in organic solvents,
removing phospholipids by evaporation, and finally pouring them into
aqueous buffer solution, stirring to make the dried lipid membranes
hydrate successfully [21]. However, this method will produce many
heterogeneous paleosecular liposomes. The reverse phase evaporation
method is to reisolate the phospholipid molecules in the organic phase,
and then adding aqueous phase buffer to a mixed solution to decompress
distillation [22]. The two-phase system was ultrasonic treated to
transparent and uniform single-phase dispersion, and it was observed
that the solution state did not change for at least 30 minutes, then the
organic solvent was removed by rotary evaporation. The liposomes
prepared by reversed-phase evaporation method have unique advantages in
encapsulating water-soluble substances, but may have drug residue
[23]. The solvent injection method does not need to be helpful in
ultrasonic waves, simply dissolve lipids in the organic phase which is
generally diethyl ether or ethanol, and then injects the lipid solution
into the aqueous solution, the particle diameter is obtained by
narrowing a smaller-distribution [24]. With using ethanol, it can
remain stable at higher temperature, but it is difficult to remove all
the solvent, and the possibility of
biological deactivation is high. Compared with ethanol, ether injection
can completely remove the solvent and form liposomes with high
entrapment efficiency, but due to the low boiling point of ether, it
will lead to uneven aggregation and dissolution at high temperature
[25]. The bubble method is a method without the use of organic
solvents, in which inert gas bubbles are introduced into the lipid
mixture to prepare liposomes with mild preparation conditions and high
entrapment efficiency [26]. The uniqueness of supercritical fluids
is that there is no difference in liquid and gaseous. The researchers
found that the properties of liposomes obtained by supercritical fluids
were better than those prepared by conventional solvents [27].
Supercritical carbon dioxide has become a substitute for organic
solvents because of its low critical temperature and pressure, similar
solubility to non-polar solvents, low application cost, and non-toxicity
[28].
Conventional preparation techniques generally lack the precise
regulation of liposome particle size on macro levels, batch
reproducibility and quality repetitive control. Microfluidics refers to
the precise control of fluid behaviors in micro-channels or designing
different microfluidic channels to realize the mixed reaction of
different liquids. Delivery carriers with small particle size
distribution, high repeatability between batches and significantly
improved drug loading rate can be prepared by microfluidic technology
[29]. Microfluidic techniques can overcome the disadvantages of
conventional methods, a highly
stabilizing liposomes that can accurately control physical parameters
can be obtained by designing flow rate, flow rate ratio, mixing speed,
and lipid concentration. In order to be able to scale production, the
preparation steps should be as simple as possible. For human health,
liposomes should avoid using hazardous chemicals or solvents when
prepared. The residual solution is potentially toxic to human body and
causes great environmental pollution, so it should be eliminated in the
preparation process. Therefore, when preparing liposomes, we should
consider comprehensively and choose the most suitable preparation method
to achieve the most beneficial results.
Relationship between the properties and immunogenicity of
liposome carrier. Since Allison first reported in 1974, liposomes
mainly exert immunity through the ”pool effect” and immune stimulation,
and gradually become the research object of vaccines as adjuvants [30,
31]. A mass of liposomes containing antigens are ingested by
macrophages through natural channels, they can be stored in macrophages
for slow release, thus introducing antigens to APC, promoting downstream
cells to secrete a series of cytokines, and enabling the body to
maintain high-valence antibodies (Fig. 2). Although vaccine carriers
based on other materials, such as metal nanoparticles, inorganic (such
as silica) nanoparticles, and polymer nanoparticles [32, 33] have
also shown advantages over traditional preparations in achieving
delivery and adjuvant functions, most vaccine vectors are still in the
preclinical development stage, and their feasibility needs to be further
explored. Also, based on the studies
of existing tumor-specific antigen delivery, we can tell liposome
carriers show better properties than non-liposomes [34].
Several key parameters mentioned above, such as surface charge, physical
size, and degree of PEG modification, play a very important role in
regulating the biological behavior of liposome vaccine vectors. The
interaction of various factors finally plays a comprehensive role in the
immunostimulatory activity of the whole carrier. In addition, the loaded
way of antigen is also very important. Mixing the carrier with the
antigen and using mild adsorption may be more beneficial to maintain the
relative activity of the antigen, but the surface adsorption also means
a higher antigen release rate, resulting in low efficiency of antigen
uptake. Encapsulating antigens in liposome carriers can achieve
sustained release of antigens or drugs, resulting in a more effective
cross-presentation process and a better lysosome escape [35].