Non-invasive delivery of stealth, brain-penetrating nanoparticles across the blood − brain barrier using MRI-guided focused ultrasound☆
Graphical abstract
Introduction
Many therapeutic agents have potential for the treatment of central nervous system (CNS) disorders; however, few are able to cross the blood–brain barrier (BBB) and/or penetrate within CNS tissue. The BBB is essential for the maintenance of the CNS environment and regulates the traffic of most molecules to and from the brain [1], [2]. Unfortunately, the BBB also limits systemically administered drugs from reaching the brain in therapeutically relevant concentrations [3], [4]; thus, drug dose and efficacy are often limited by systemic side effects [5]. In the case of some CNS disorders, such as glioblastoma, Alzheimer's, Parkinson's, cerebral palsy, epilepsy and stroke, the BBB can be impaired and “leaky” [6], [7], [8]; however, this impairment is often heterogeneous and diseased cells are often found in normal brain parenchyma in regions supplied by healthy blood vessels with normal BBB function [9]. Other CNS disorders, including lysosomal storage diseases [10], depression [11], and recurrent migraines [12] present even higher obstacles to effective drug delivery into the brain.
Transcranial MRI-guided focused ultrasound (MRgFUS) can non-invasively permeabilize the BBB in a safe, reversible fashion. Image guidance adds the ability to target specific regions with a high degree of accuracy [13], [14], [15]. Using this strategy, standard microbubble (MB) contrast agents [16], [17], [18] are first injected intravenously. In the region receiving FUS, the MBs are activated, producing a variety of mechanical, chemical and thermal effects [19], [20], [21], [22], [23], [24] that lead to a transient disruption of tight junction complexes and induction of active transport of agents into the brain parenchyma [25], [26], [27]. Small animal studies have demonstrated that the BBB or blood–tumor barrier permeability is increased by MRgFUS [28], improving delivery of therapeutic agents, such as Herceptin and Doxil, antibody- and liposomal-based delivery systems respectively, as well as treatment outcomes in experimental models of diseases such as glioma and Alzheimer's [29], [30], [31], [32]. Additionally, MRgFUS-mediated BBB disruption in non-human primates was shown to be safe and accurate, while producing no significant brain tissue damage or compromised visual function in highly eloquent occipital cortex regions [15].
Once therapeutics cross the BBB, they next encounter a complex microenvironment within the brain extracellular space (ECS), which significantly limits and controls their movement. The ECS is anisotropic with mixed electrostatically charged and hydrophobic regions comprising 15–20% of the total normal brain volume [33]. Although it was previously thought that a therapeutic nanoparticle must be smaller than 64 nm to penetrate within the ECS [34], it was recently shown that much larger particles, up to 114 nm in size, can penetrate within normal brain parenchyma, if densely coated with low molecular weight poly(ethylene glycol) (PEG) [35]. PEG sterically stabilizes nanoparticles and minimizes protein absorption, increasing nanoparticle circulation time. However, PEGylated stealth particles also show decreased interactions with cells, limiting cell uptake or passage across an intact BBB. MRgFUS has yet to be combined with a drug delivery platform that can provide sustained release of a therapeutic and can overcome the tissue-penetration barrier within the brain microenvironment once crossing a permeabilized BBB.
The ideal method to deliver drugs to the CNS would include (i) a stealth circulating nanoparticle that can avoid rapid clearance by the reticuloendothelial system (RES), (ii) a non-invasive approach to bypass the BBB, (iii) the ability of the nanoparticle to penetrate within the brain parenchyma and (iv) provide sustained release of a therapeutic agent at the sites of disease. We hypothesized that coupling MRgFUS-mediated BBB opening with circulating, brain penetrating nanoparticles (BPNs) would achieve significant accumulation and spread of BPNs in select regions of the brain with minimal side-effects. We designed and characterized the behavior of densely PEGylated NPs within brain tissue and demonstrated that 60 nm BPNs can penetrate the rat brain parenchyma when delivered with MRgFUS. This is the first study to show, in combination with MRgFUS and MBs, the successful trans-BBB delivery of a biodegradable polymeric nanoparticle that is capable of penetrating within the brain microenvironment. This approach represents a promising strategy to overcome the significant hurdles for drug delivery to the brain and improve therapeutic efficacy for many CNS diseases.
Section snippets
Nanoparticle preparation and characterization
Model nanoparticles were prepared as previously described [35]. Briefly, 40- to 200-nm red fluorescent COOH-modified polystyrene (PS) particles (Molecular Probes) were covalently modified with methoxy (MeO)-PEG-amine (NH2) (5 kDa MW; Creative PEG Works) by carboxyl amine reaction. An excess of MeO-PEG-NH2 was added to the PS particle suspension and mixed to dissolve the PEG. N-hydroxysulfosuccinimide (Sigma) was added to the reaction tube and 200 mM borate buffer, pH 8.2, was added to a 4-fold
Transport of BPN in normal rat brain ECS
To ensure that large, therapeutically relevant nanoparticles could penetrate within the rodent brain, we first determined the effect of particle size and surface chemistry on transport rates of modified particles in the rat brain tissue. The hydrodynamic diameters of the particles are listed in Table 1. The 100-nm and 200-nm COOH-modified particles displayed low transport rates in freshly excised viable rat brain slices, as measured by arithmetic ensemble mean squared displacements (< MSD >) (
Discussion
We report here the first use of MRgFUS and MBs with a biodegradable polymeric nanoparticle platform that can penetrate within the brain microenvironment. MRgFUS can be used to safely deliver 60 nm PS-PEG NPs and 75-nm PLGA-PEG across the BBB in vivo. Densely PEG-coated PS NPs (BPN) circulate and accumulate in the brain in regions where the BBB is disrupted by FUS and MBs. The 60-nm PS-PEG NPs can rapidly diffuse and penetrate within the brain parenchyma 10-fold slower than their effective
Acknowledgments
Supported by NIH RO1 CA164789.
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Cited by (0)
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Densely PEG-coated PS nanoparticles (NP) injected systemically can disperse in the brain following delivery with microbubbles (MB) and non-invasive MR-guided focused ultrasound (FUS).
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Authors contributed equally to this work.