Nanotechnology for Drug and Gene Delivery
Smaller size means easier access to the back of the eye.
UDAY B. KOMPELLA, PhD • STEPHANIE YOULIOS, BS
Therapy and management of retinal diseases, including choroidal neovascularization, geographic atrophy, diabetic retinopathy, retinitis pigmentosa, and retinoblastoma, are challenging tasks. The challenge arises due to the variations in disease presentation, the absence of suitable surrogate markers, the need for chronic therapy, and biological ocular barriers that limit the delivery of therapeutic agents to the target site. This article addresses how the emerging field of nanotechnology can advance therapy and management of retinal diseases.
Nanotechnology entails the design of materials of nanosize dimensions with unique properties and applications. Nanomedicines, the therapeutic products of nanotechnology, provide unique opportunities to treat diseases of the back of the eye, especially by improving drug delivery, gene delivery, and imaging. This article summarizes nanotechnology fundamentals and the application of nanosystems for treating diseases of the eye, previously elaborated elsewhere.1
Uday B. Kompella, PhD, serves on the faculty of the University of Colorado Anschutz Medical Campus in Aurora. Stephanie Youlios, BS, is a research assistant at EyeTrans Technologies in New York, NY. Dr. Kompella is cofounder of EyeTrans and reports financial interest in that regard. Dr. Kompella can be reached via e-mail at Uday.Kompella@ucdenver.edu.
FUNDAMENTALS OF NANOSYSTEMS
Delivery systems designed through nanotechnology or nanosystems have at least one dimension in the range of 1 to 1,000 nm, with unique properties imparted to the particles by design. Potential advantages of nanosystems include the following.
• Enhanced permeation and retention in leaky vascular regions vs normal tissue, thereby improving drug targeting
• Enhanced cellular and organelle delivery of therapeutic agents relative to larger delivery systems
• Prolonged circulation similar to antibodies
• Small nanosystems being potentially more suitable for intracellular and organelle-targeted delivery of therapeutic agents than larger delivery systems, because mammalian cells are a few microns in diameter, and intracellular organelles are much smaller
• Prolonged suspension in ophthalmic formulations, as well as fluid compartments of the body, including vitreous, due to their lower settling velocity
• Incorporation of targeting ligands (functionalized nanosystems) to facilitate tissue-, cell-, and organelle-selective delivery of the associated therapeutic agent
• Design of chemical moieties in nanosystems to allow stimuli (eg, light, enzyme, pH, etc.)–responsive release of the associated therapeutic agent
• Bioadhesion due to the design of nanomaterials that can interact with mucous layers
• Sustained delivery of the associated therapeutic agent
TYPES OF NANOPARTICLES EMPLOYED FOR RETINAL DISEASES
With the growing number of innovations, the prefix nano- has been affixed to a variety of terms, bringing to the fore nanoparticles, nanosuspensions, nanoliposomes, nanotubes, etc., which are collectively referred to as nanosystems or nanoparticles in this article.
Nanosystems are prepared using the pure drug, primarily or along with a significant amount of carrier, including more commonly used materials, such as polymers, lipids, amino acids/peptides/proteins, and carbohydrates.
Additionally, other nontraditional ingredients, including gold, silver, iron oxide, pure carbon-based structures, etc., are also employed in designing nanosystems for therapeutic purposes. These nanosystem products can be solid, liquid, or semisolid dosage forms. Like any other pharmaceutical product, the use of materials should be such that the benefits outweigh any risks posed by the materials.
The selection of a nanosystem should weigh a number of factors, including the disease environment, target tissue and cell properties, route of administration, pathways and kinetics of disposition, safety, and scalability. Additionally, intellectual property should be secured through patents for innovative technologies addressing unmet medical needs, thus facilitating funding for drug product development.
Retinal drug therapies are at the forefront of drug delivery in human medicine, with a variety of innovative therapeutic agents and drug delivery systems making inroads in the eye ahead of other parts of the body.
Examples include the approval of the first antisense oligonucleotide, fomavirsen (Vitravene, Isis Pharmaceuticals, Carlsbad, CA); the first aptamer, pegaptanib sodium (Macugen, Bausch + Lomb, Rochester, NY); and the first injectable, miniature, nondegradable fluocinolone acetonide implant (Iluvien, Alimera Sciences, Alpharetta, GA), for treating retinal diseases. With this legacy, it is anticipated that new nanosystems will make inroads for treating retinal diseases.
Nanosystems are not foreign to eye products. Molecular medicines in solutions, including small-molecule drugs in ophthalmic solutions, as well as macromolecule drug products such as fomavirsen, pegaptanib, ranibizumab (Lucentis, Genentech, South San Francisco, CA), bevacizumab (Avastin, Genentech), and aflibercept (Eylea, Regeneron, Tarrytown, NY), all include the therapeutic agent at the smallest nanodimension possible. Additionally, some of the ophthalmic emulsions and suspensions on the market have particles/droplets at a nanodimension.
Nanoparticles modified on their surfaces with ligands capable of recognizing cell surface receptors, referred to as functionalized nanoparticles, are a new type of nanoparticle that can localize to target cells. By selecting a target receptor that can internalize into cells, functionalized nanoparticles are engineered to enhance intracellular and potentially organelle delivery of the associated therapeutic agent. For instance, nanoparticles functionalized with transferrin enhance the uptake by ocular cells, as well as transport across tissues, compared with nanoparticles that are nonfunctionalized.2
Commonly assessed/developed products of nanomedicine include pure drug suspensions, emulsions, gels, micelles, and liposomes with a nanodimension. All of these systems have been assessed for retinal delivery. Diverse types of drug molecules, including small molecules, proteins, and genes, have been assessed in nanoparticles for enhanced, sustained, or targeted drug delivery to the back of the eye. Products that utilize nanosystems for their unique advantages to address unmet needs are more likely to succeed as new products in the clinic.
DISPOSITION OF THE NANOSYSTEMS FROM THE BACK OF THE EYE
Disposition or removal of a nanosystem from the site of administration depends on the physiological and pathological surroundings of the system, as well as the properties of the nanoparticle and materials composing the system. Size, charge, recognition by innate clearance mechanisms/cells/proteins, and rate of biodegradation are some critical parameters that can influence nanoparticle disposition, although other properties may also affect disposition.
For instance, as the size of a macromolecule increases, its removal from the vitreous is reduced, at least to a point.3 Similarly, relative to a small drug molecule in solution, particulate systems, including nanoparticles, prolong the residence time of the drug in the vitreous, as well as other parts of the back of the eye.3,4 Given the negative charge of vitreal matrix components, it is expected that positively charged nanosystems may bind to vitreous matrix and be retained for longer periods relative to other types of particles.5
If a nanosystem is administered by routes other than the intravitreal route, the half-life will differ. Current evidence indicates that small nanoparticles of 20 nm in diameter are cleared more rapidly from the periocular space than the suprachoroidal space, for instance.6-8 Binding of proteins and other biological materials to the particles and changes in the properties of nanoparticles at the site of administration may alter particle disposition.9
The different materials used in preparing nanoparticles may erode or degrade at different rates, resulting in particle disintegration and removal of the smaller components thus formed. Understanding the disposition of a nanosystem is one of the critical factors in determining the frequency of dosing, as well as selecting the most appropriate route of administration for drug therapy. Additionally, understanding the disintegration and disposition of the nanosystem components and byproducts would be beneficial in establishing the safety of nanosystems.
APPLICATIONS OF NANOTECHNOLOGY FOR RETINAL DISEASE
Nanoparticle-based delivery benefits therapeutic agents of a diverse nature, including small molecules, proteins, and nucleic acids. Among small molecules, those that are poorly soluble and/or poorly permeable will benefit the most from enhanced delivery with nanoparticles. Proteins, especially those that act on remote or intracellular targets, may benefit from nanoparticles if the materials and methods used in nanoparticle preparation do not perturb protein structure and conformation. Nanoparticles can also protect nucleic acids from degradation, while enhancing their intracellular delivery. Some examples of small molecule, protein, and nucleic acid delivery using nanoparticles are described below.
Enhanced Cell Entry and Efficacy for Poorly Permeable Molecules
Among particles ranging in size from 20 to 2,000 nm, retinal pigment epithelial cells take up the 20-nm particles to the greatest extent.10 Thus, if a therapeutic agent acts inside the cell and has poor permeability or requires prolonged intracellular residence, the design of a small nanoparticle would be suitable.
Indeed, nanoparticles based on the biodegradable polymer poly(lactide-co-glycolide) enhanced the intracellular delivery of an antisense oligonucleotide targeting VEGF in RPE cells.11 These nanoparticles are more effective than the oligonucleotide alone in suppressing VEGF secretion.
The potential reasons for the benefit of nanoparticles in this case include enhanced uptake of a hydrophilic macromolecule, as well as protection of the oligonucleotide from degradation prior to cell entry.
Enhanced Solubility, Cell Entry, and Persistence
Dendrimers are branched polymer structures that are typically water-soluble and that allow drug entrapment or chemical conjugation within their architecture.12 Using gatifloxacin complexation with dendrimers, Durairaj et al demonstrated that gatifloxacin’s solubility, corneal epithelial cell entry, and conjunctival persistence and trans-scleral transport were improved.13
In this approach, gatifloxacin interacts via ionic, hydrogen bond, and hydrophobic interactions with g6 dendritic polyguanidilyated translocator, a polymer that is very soluble in water. As a result, the solubility of gatifloxacin is increased, allowing for the preparation of an ophthalmic solution with greater strength. Such an approach could potentially reduce dosing frequency and/or allow for greater drug delivery to the back of the eye.
The majority of medications are administered remotely for effects on distant targets in the body. Thus, although the safety of repeated intravitreal injections is improving, macromolecules currently administered by this local route can potentially be administered by a conventional intravenous route to minimize the growing number invasions of the vitreous and the retinal surroundings in aging patients.
At the same time, we cannot exclude that intravitreal injections with improved safety may become as routine as intravenous injections. Use of functionalized nanoparticles capable of recognizing disease sites can selectively localize any associated therapeutic agent at the target site, following intravenous administration.
Nanoparticles functionalized on the surface with RGD peptide, transferrin, or their combination and administered intravenously better target CNV regions of the eye and enhance the efficacy of an anti-VEGF plasmid, compared to plain nanoparticles.15
Thus, targeted therapy for retinal diseases is feasible following intravenous dosing. Verteporfin (Visudyne, Bausch + Lomb), a therapeutic nanoliposome approved for use with photodynamic therapy in age-related macular degeneration, is another example of an intravenously administered therapeutic product. However, verteporfin is not a functionalized nanosystem, and its functionalization can potentially improve its target site association and accumulation.
Protein molecules are large and complex in their structure, with conformation that controls their activity and lability in the presence of ubiquitous proteolytic enzymes in the body. Because protein therapeutics, such as pegaptanib, ranibizumab, bevacizumab, and aflibercept, are shifting the ophthalmic pharmaceutical market largely toward the back of the eye and because these molecules improve vision in potentially blinding diseases, there is a significant focus on improving the delivery of protein drugs to the retina.
One key unmet need is sustained protein delivery to reduce the frequency of macromolecule dosing from once every month or two. One approach to sustaining the delivery of protein drugs is to use nanosystems that are capable of sustaining protein drug release, while protecting the system from degradation.
In one approach, a protein drug coated on a nanoparticle is trapped in porous microspheres, using supercritical fluid technology.16 This technology minimizes or eliminates the exposure of the protein drug to organic solvents during polymeric nanoparticle preparation, thereby maintaining protein stability during manufacturing. An alternative approach is to use polymeric gels that are preformed or formed in situ to sustain protein drug delivery.17
In addition to the aforementioned approaches, a variety of nanomaterials, including devices that are nanofabricated,18,19 could potentially be used for enhancing, targeting, or sustaining drug delivery to the back of the eye. In addition to therapeutic agent delivery, nanoparticles can also be used as diagnostic agents in either staging the disease prior to dosing or assessing the outcomes of drug therapy.
In summary, nanoparticles and other products of nanotechnology are made of a variety of materials that are biocompatible to enhance, target, or sustain drug and gene delivery. Although nanomedicines are administered through various routes, each nanomedicine is tailored for the target disease, drug, and site of administration.
While many therapeutic molecules may benefit from nanoparticles, nanoparticle technologies are readily applicable to molecules that are poorly soluble and/or permeable to improve their delivery. As nanoparticle technologies evolve for ophthalmic use, the safety, scalability, and core value of these delivery systems will be better established. RP
1. Kompella UB, Amrite AC, Ravi RP, Durazo SA. Nanomedicines for back of the eye drug delivery, gene delivery, and imaging. Prog Retin Eye Res. 2013;36:172-198.
2. Kompella UB, Trivedi R, Tyagi P, Vooturi SK. Novel amphiphilic peptide-polymer conjugates for micelle based drug delivery to the eye. Poster presented at: Conference of the American Association of Pharmaceutical Sciences; San Antonio, TX; November 10-14, 2013.
3. Durairaj C, Shah JC, Senapati S, Kompella UB. Prediction of vitreal half-life based on drug physicochemical properties: quantitative structure-pharmacokinetic relationships (QSPKR). Pharm Res. 2009;26:1236-1260.
4. Andrew JS, Anglin EJ, Wu EC, et al. Sustained release of a monoclonal antibody from electrochemically prepared mesoporous silicon oxide. Adv Funct Mater. 2010;20:4168-4174.
5. Kim H, Robinson SB, Csaky KG. Investigating the movement of intravitreal human serum albumin nanoparticles in the vitreous and retina. Pharm Res. 2009;26:329-337.
6. Amrite AC, Edelhauser HF, Singh SR, Kompella UB. Effect of circulation on the disposition and ocular tissue distribution of 20 nm nanoparticles after periocular administration. Mol Vis. 2008;14:150-160.
7. Amrite AC, Kompella UB. Size-dependent disposition of nanoparticles and microparticles following subconjunctival administration. J Pharm Pharmacol. 2005;57:1555-1563.
8. Patel SR, Berezovsky DE, McCarey BE, Zarnitsyn V, Edelhauser HF, Prausnitz MR. Targeted administration into the suprachoroidal space using a microneedle for drug delivery to the posterior segment of the eye. Invest Ophthalmol Vis Sci. 2012;53:4433-4441.
9. Kim H, Robinson SB, Csaky KG. Investigating the movement of intravitreal human serum albumin nanoparticles in the vitreous and retina. Pharm Res. 2009;26:329-337.
10. Aukunuru JV, Kompella UB. In vitro delivery of nano- and micro-particles to human retinal pigment epithelial (ARPE-19) cells. Drug Deliv Technol. 2002;2:50-57.
11. Aukunuru JV, Ayalasomayajula SP, Kompella UB, Nanoparticle formulation enhances the delivery and activity of a vascular endothelial growth factor antisense oligonucleotide in human retinal pigment epithelial cells. J Pharm Pharmacol. 2003;55:1199-1206.
12. Kambhampati SP, Kannan RM. Dendrimer nanoparticles for ocular drug delivery. J Ocul Pharmacol Ther. 2013;29:151-165.
13. Durairaj C, Kadam RS, Chandler JW, Hutcherson SL, Kompella UB. Nanosized dendritic polyguanidilyated translocators for enhanced solubility, permeability, and delivery of gatifloxacin. Invest Ophthalmol Vis Sci. 2010;51:5804-5816.
14. Sampat KM, Garg SJ. Complications of intravitreal injections. Curr Opin Ophthalmol. 2010;21:178-183.
15. Singh SR, Grossniklaus HE, Kang SJ, Edelhauser HF, Ambati BK, Kompella UB. Intravenous transferrin, RGD peptide and dual-targeted nanoparticles enhance anti-VEGF intraceptor gene delivery to laser-induced CNV. Gene Ther. 2009;16:645-659.
16. Yandrapu SK, Upadhyay AK, Petrash JM, Kompella UB. Nanoparticles in porous microparticles prepared by supercritical infusion and pressure quench technology for sustained delivery of bevacizumab. Mol Pharm. 2013;10:4676-4686.
17. Tyagi P, Barros M, Stansbury JW, Kompella UB. Light-activated, in situ forming gel for sustained suprachoroidal delivery of bevacizumab. Mol Pharm. 2013;10:2858-2867.
18. Freitas RA Jr. The future of nanofabrication and molecular scale devices in nanomedicine. Stud Health Technol Inform. 2002;80:45-59.
19. Chen L, Henein G, Luciani V. Nanofabrication techniques for controlled drug-release devices. Nanomedicine (Lond). 2011;6:1-6.