Tunaidi Ansari May 3, 2008
Biodegradable Long-Circulating Polymeric Nanospheres
R. Gref, Y. Minamitake, Y. Peracchia, V. Trubetskoy, V. Torchilin, and R. Langer
There are six specific features that form an efficient carrier, which can continuously deliver drugs intravenously. These include: 1) encapsulated agents of relatively high weight fraction compared to the rest of the carrier system, 2) efficient incorporation of the agents from the first step to the final carrier, 3) competence in freeze-drying and subsequent thawing, 4)
biodegradability, 5) small size, and 6) ability to stay in the blood stream for relatively longer periods of time [1].
Gref et al developed degradable polymeric nanospheres that exhibit the previously listed features of an efficient carrier. Biodegradable materials known to be harmless in the human body, such as poly(lactic-co-glycolic acid) (PLGA) (shown in Figure 1), polycaprolactone (PCL), and their copolymers, were used as the core of these particles [2,4,6,7]. In order to form adequate coating, polyethylene glycol (PEG) was covalently bonded to the nanosphere core. This was accomplished by diblock copolymers, PEG-R, and R being one of the listed biodegradable materials. Since these diblock polymers
are amphiphilic, they were used to obtain a phase-separated structure.
The nanospheres were constructed by first dissolving PEG-R in an organic solvent. Subsequent vortexing and sonication formed an emulsion in an aqueous phase. This is because PEG is
hydrophilic and R is lipophilic [5]. Afterwards, evaporation of the solvent yielded a solidified nanosphere core. These were collected through centrifugation and then lyophilized for easy redispersion in aqueous solutions.
Quasi-elastic light scattering (QELS) verified that aggregation did not occur. Atomic force microscopy (AFM) was utilized to confirm the nanospheres’ spherical shape, reporting a mean diameter of 90 and 150 nm. X-ray photoelectron spectroscopy (XPS) established that PEG was concentrated mainly within 5 nm of the nanospheres’ outer layer, and only a nominal amount of PEG was detached.
In Figure 2, (A) and (B) show images of Figure 2: Images of nanospheres and related
plots.
Figure 1: Illustration of the chemical structure of PLGA [3].
nanospheres and respective lengths taken from the AFM, where (A) is PLGA and (B) is PEG-PLGA. (C) depicts the QELS results and displays diameters of the PEG-PLGA nanospheres. Finally, (D) displays the XPS analysis, where trace 1 corresponds to PLGA nanospheres, trace 2 corresponds to PEG-PLGA nanospheres, and trace 3 corresponds to PEG alone.
PEG-coated and non-coated particles were injected into mice in order to determine their respective effectiveness. As shown in Figure 3 (A), as the molecular weight of PEG increases, there is a corresponding increase in blood circulation time. This is due to the increased thickness of the PEG coating, which lessens the effect of opsonization. In
addition, Figure 3 (B) proves that within five minutes, 66% of the non-coated particles were removed by the liver, while fewer than 30% of the PEG-coated nanospheres met the same fate only after five hours. Gamma scintigraphy proved that non-coated nanospheres were present only in the liver and spleen, while large amounts of PEG-coated particles were discovered in the blood pool, mainly the heart and lungs [18].
Next, lidocaine, a model drug, was encapsulated into the PEG-coated nanospheres and achieved high drug loadings and entrapment efficiencies. Lidocaine was continually released in vitro for over 14 hours. It was established that the higher the particle drug content, the slower the release. This trend can be seen in Figure 4 (A).
This experiment by Gref et al presents a novel development of intravenously administered carriers with adequate blood circulation times. It is improves upon previously attempted projects such as albumin or galactose microspheres, which are frequently used in clinical studies for imaging [19,20,21]. However, the downfall of these microspheres is their rapid clearance from the blood within 20 seconds, which prevents subsequent usage in various imaging applications. Other innovations also challenged the rapid clearance rate frontier. Progress has been made on the micro-particle scale by
appending poloxamer or polysorbate onto nondegradable polystyrene or polymethylmethacrylate particles. Another alternative that has been performed was the formation of liposomes and other carriers containing glycolipids, albumin, or derivatives of PEG [22-30]. Gref et al hoped to make further advancements into this field on the nano-particle scale and attempted to solve the issues with previously performed experiments.
When forming their nanosphere cores, they relied on the principle of adjusting chemical
compositions and molecular weights of polymers in order to precisely control degradation times of the core and the release kinetics of the encapsulated agents [8]. Opsonization and macrophage detection present barriers to long term blood circulation [17]. As such, PEG was selected as a
Figure 3: Data on mice, which were injected with varying
molecular weights of PEG-PLGA.
Figure 2: Results pertaining to lidocaine drug release.
component of the nanosphere coating. Since the diblock copolymers that were used consisted of ethylene glycol groups bonded together and attached to repeated hydrophobic monomers, Gref et al were able to utilize the different solubilities of the PEG and R to acquire the phase-separated organization. This was used in the emulsion process of the nanosphere formulations.
The lidocaine-loaded nanospheres, which exhibit the inverse relationship between drug content and release speed, may be accounted for by drug crystallization within the nanospheres.
Referring to Figure 4 (B), calorimetric and x-ray diffraction have shown that with low content, the drug exists as a dispersion inside the core and with high content, phase separation causes a portion of lidocaine to be crystallized [14,15]. Thus, in order to control timing of drug-release, several factors are involved. These include: manipulation of the diblock polymers’ chemical composition, effective drug loading, and optimal nanosphere particle sizes [9].
Gref et al’s innovation can be applied in multiple ways. Further research and experiments will inevitably allow these biodegradable long-circulating polymeric nanospheres to be fruitful in areas such as drug delivery, medical imaging, and gene therapy. Specific applications include attaching a suitable protein to the surface of the nanospheres, which may possibly permit
endocytosis of DNA-containing particles, or even attaching antibodies to PEG, and thus forming highly specific immune defense systems [10,11,12].
Nevertheless, there may be areas of improvement regarding this particular study. Researching other types of particle coating most effective for their functions or delivery target may be even more useful in future applications. For example, amylose serves as an effective coating for carriers targeting the colon [13]. Alternatively, further investigation into the unloading properties of agent release may be useful for specific drug delivery and the timing of their resulting effects. This may be directly applied to patients who need to regulate insulin levels as a result of diabetes mellitus. Since diabetes mellitus is a chronic disease, administered medication needs immediate effect; altering timing and release of drugs from the nanospheres would therefore serve to better monitor and suppress the symptoms of the disease [16].
A very practical next step may encompass evaluating the different environmental factors, such as pH, temperature, and solvents, and their effects on the nanoparticle PEG coating. Regulating these factors is useful for determining the speed of degradability and subsequently the release of agents. Controlling the endurance of the protective layer will be a milestone achievement in studying the encapsulation properties of the nanospheres. Additionally, further research into the differences between the various materials used in the R segment of the PEG-R complex may help to fine tune the experimental results. Perhaps, even the integration of triblock copolymers onto the nanospheres can be performed and studied for comparison with the currently used diblock copolymers.
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