Supramolecular Pharmacy 11. Nanoparticles for drug delivery 2 (polymeric and inorganic NPs) Ondřej Jurček 1 Nanoparticles (NPs) in drug delivery (nanocarriers) 2 • Oral, local, topical, and systemic nano-/microparticle delivery systems have been approved by FDA or EMA • Intravenously administered NPs are the most studied preclinically and clinically • Systemic delivery provides direct access to nearly all parts of the body • Organic (including polymeric micelles and vesicles, liposomes, dendrimers, polymers, and hydrogels) and inorganic (including gold and mesoporous silica NPs) • Porous NPs provide enhanced drug protection, controlled release, extended circulation, improved targeting to diseased tissues/cells • Inorganic NPs provide the same and stimuli responsive functions due to surface plasmon resonance (thermal heating or imaging) or magnetic responsiveness (MRI, magnetic targeting) Anselmo et al. Bioengineering & Translational Medicine 2016, 1, 10-29. Clinically relevant NPs and their applications 3Anselmo et al. Bioengineering & Translational Medicine 2016, 1, 10-29. Black: Approved applications, Red: Application in clinical trials (in 2016). • Nanoparticles have been developed to overcome the limitations of free therapeutics and navigate biological barriers — systemic, microenvironmental and cellular — that are heterogeneous across patient populations and diseases • Personalized delivery – precision medicine Nanoparticles in pharmacy 4Mitchell et al. Nature Reviews: Drug Discovery 2021, 20, 101–124. Polymeric nanoparticles 5Mitchell et al. Nature Reviews Drug Discovery 2021, 20, 101–124. • Can be formed of biocompatible natural or synthetic polymers • Therapeutics can be encapsulated within the NP core, entrapped in the polymer matrix, chemically conjugated to the polymer or bound to the NP surface • Nanocapsules (cavities surrounded by a polymeric membrane or shell) and nanospheres (solid matrix systems) • Modulating properties such as composition, stability, responsivity and surface charge, the loading efficacies and release kinetics of these therapeutics can be precisely controlled • NPs are divided further into shapes such as polymersomes, micelles and dendrimers Polymeric nanoparticles 6Mitchell et al. Nature Reviews Drug Discovery 2021, 20, 101–124. • Polymersomes are artificial vesicles, with membranes made using amphiphilic block copolymers. They are comparable to liposomes, but are reported to have improved stability and cargo-retention efficiency, making them effective vehicles for the delivery of therapeutics to the cytosol, e.g., made of PEG, poly(dimethylsiloxane) (PDMS) • Polymeric micelles are also typically responsive block copolymers, self-assemble to form nanospheres with a hydrophilic core and a hydrophobic coating: this serves to protect aqueous drug cargo and improve circulation times. Polymeric micelles can load various therapeutic types — from small molecules to proteins — and have been studied for the delivery of cancer therapeutics in clinical trials Polymeric nanoparticles 7Mitchell et al. Nature Reviews Drug Discovery 2021, 20, 101–124. • Dendrimers are hyperbranched polymers with complex three-dimensional architectures for which the mass, size, shape, and surface chemistry can be highly controlled. Active functional groups present on the exterior of dendrimers enable conjugation of biomolecules or contrast agents to the surface while drugs can be loaded in the interior. Dendrimers can hold many types of cargo but are the most commonly investigated for the delivery of nucleic acids and small molecules. Charged polymers such as poly(ethylenimine) (PEI) and poly(amidoamine) (PAMAM) are used. Polymeric nanoparticles 8 • Polymeric NPs have one important benefit, their scale-up and manufacturing are possible under Good Manufacturing Practices (GMP by WHO) • Other benefits are significant stability of polymeric nanoparticles in biological fluids along with the wide availability of various polymers, the opportunity to functionalize their surfaces and to modulate polymer degradation and the leakage of the entrapped compound(s) as a function of specific stimuli • Ideally the polymers must be biocompatible, biodegradable, and possess specific mechanical and physicochemical properties • The first polymers used to develop polymeric nanoparticles (PNs) were nonbiodegradable polymers, such as poly(methylmethacrylate) (PMMA), polyacrylamide, polystyrene, and polyacrylates. The nanosystems made up of these materials exhibited a rapid and efficient clearance, also, chronic toxicity and inflammatory reactions were observed • Degradation rate of biodegradable polymeric nanoparticles can be influenced by several parameters, including their physicochemical properties (size, structure, molecular weight) and external factors, such as pH and temperature Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. Polymers for nanoparticles 9 • Synthetic and non-synthetic biodegradable polymers • Synthetic biopolymers: include synthetic polymers such as poly(D,L-lactide) (PLA), poly(D,L-glycolide) (PLG), co-polymer poly(lactide-co-glycolide) (PLGA), polyalkylcyanoacrylates, poly-Ɛ-caprolactone, polyhydroxyalkanoates (PHAs), and poly(alkyl cyanoacrylates) (PACA) (some approved by FDA and EMA) • Non-synthetic natural biopolymers: • Animals – albumin nanoparticles (Abraxane®), gelatin, hyaluronic acid • Plants – cellulose, starch, soy protein, zein • Marine organisms – chitosan (cationic), alginate, carrageenan Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. Hyaluronic acid Chitosan Cellulose Albumin Preparation of polymeric NPs 10 • The most important: emulsification and solvent evaporation (small-moderate scale manufacturing: Polymer in organic solvent into water with surfactants and stabilizers – emulsification – evaporation of organic solvent (spray-drying) • Nanoprecipitation: water miscible organic solvent with polymer (drug) is slowly dropped into aqueous solution • Supercritical anti-solvent method: polymeric solutions sprayed into CO2 under high pressure • Salting-out: electrolytes such as magnesium chloride, calcium chloride and magnesium acetate or saccharides (non-electrolyte) added to a polymeric solution • The length of polymer plays an important role (effects also degradation), stabilizers.. • Prevent aggregation – electrostatic and steric stabilization (surfactants) • Surfactants: (i) anionic (negative charge); (ii) cationic (positive charge); (iii) zwitterionic or amphoteric (charge depends on the pH of the medium), and; (iv) non-ionic (no charge) • Vitamin E TPGS (vitamin E D-α-tocopheryl-polyethylene glycol 1000 succinate) enhances drug encapsulation, cellular uptake, therapeutic efficacy, and oral bioavailability of nanocarriers Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. Physico-chemical properties of polymeric NPs 11 • Size, shape, stability, drug-release profiles and surface characteristics • The European Nanomedicine Characterization Laboratory (EUNCL) and the US National Cancer Institute Nanotechnology Characterization Laboratory (NCI-NCL) have developed multiple standard operating procedures for nanomaterial assessment, establishing mean size and polydispersity index as the critical quality attributes (∼100 nm) • Interaction with a biomaterial could favor the formation of aggregates/particles of different mean sizes, leading to significant differences in cell uptake and distribution, toxic effects, and fate within the cell Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. Drug 12 • Encapsulating hydrophilic and hydrophobic molecules, such as salts, proteins, and high-molecular-weight DNA or antisense nucleic acids • Drugs encapsulated in PNPs are released by means of diffusion through the polymeric network, erosion of the matrix material, hydrostatic swelling, or by a combination of these mechanisms Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. Tumor microenvironment 13Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. • Large differences between tumor and healthy tissues, e.g., vascular abnormalities, oxygenation and perfusion levels, pH, and metabolic status • Solid tumors are characterized by a heterogeneous population of neoplastic cells supplied by an irregular and discontinuous endothelium with large gaps between the endothelial cells, and abnormally thick or thin basement membranes where pericites are loosely attached to endothelial cells • The abnormal and disorganized tumor vasculature results in inefficient blood flow inside the tumor mass, hypoxia, and low extracellular pH Enhanced permeability and retention (EPR) effect 14Xie et al. Curr. Cancer Drug Target 2019, 19(4), 257. • high–molecular weight nontargeted drugs and prodrugs accumulate in tissues that offer increased vascular permeability, such as in sites of inflammation or cancer Tumor microenvironment 15Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. • Hypoxia plays a crucial role in tumor growth and metastasis through the induction of molecular signaling which is responsible for genetic instability, inflammation, immunosuppression, epithelial–mesenchymal transition and altered metabolism • It also confers resistance against several kinds of treatment, such as radiation, chemo-, photodynamic and immunotherapies, which require oxygen for efficacy • Oxygen can diffuse for maximum 150 μm beyond the capillary wall, which implies that when a tumor reaches a certain size (∼2mm3), a state of cellular hypoxia begins • Angiogenesis is a cellular mechanism which is upregulated in tumoral microenvironments and creates new blood vessels to further assist tumor growth by supplying oxygen and nutrients • Hypoxia also results in metabolic acidosis caused by increased glycolysis (the Warburg effect); this lowers the extracellular pH to 6.0–7.0 • The elevated breakdown of glucose produces large amounts of lactic acid and significant amounts of free protons (H+) which are pumped into the extracellular milieu Tumor microenvironment 16Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. • The resulting pH gradients between intra- and extracellular compartments within the tumor tissue, as well as between the tumor mass and the general host tissue, are potential sources of variable and often inefficient partitioning and distribution of drugs • Exposure to chemotherapy may favor the selection of tumor-cell clones with acidic organelles, which are able to entrap the drugs, and if these organelles are part of the secretory pathway, then the drug will be transported out of the cell through exocytosis. All these factors in the tumor microenvironment contribute to multidrug resistance (MDR) phenomena Various drug targeting approaches 17Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. Nanoparticle passive targeting and PEGylation 18 • Passive targeting: Convection (large molecules travel through large pores) or passive diffusion (concentration gradient, small molecules) – enhanced permeability and retention effect (EPR) – 100-800 nm • Polymeric nanoparticles, micelles, liposomes, and dendrimers of 80–150 nm are retained in the solid tumor tissue, but smaller particles (<20–30 nm) can easily diffuse into other compartments • Total amount of protein adsorbed onto the surfaces of the nanoparticles significantly decreased when the MW of PEG was a maximum of 5 kDa • AntiPEG-antibodies - at the end of the last century, only 0.2% of the population had them, while since 2012 a dramatic increase in the number (up to 25%) has been observed - “accelerated blood clearance” (ABC) • Active targeting moieties: antibodies, glucose, transferrin, folate, transporters and integrin ligands Gagliardi et al. Frontiers in Pharmacology 2021, 12, 601626. Polymeric nanoparticles 19Mitchell et al. Nature Reviews Drug Discovery 2021, 20, 101–124. • Overall, polymeric NPs are ideal candidates for drug delivery because they are biodegradable, water soluble, biocompatible, biomimetic and stable during storage. Their surfaces can be easily modified for additional targeting — allowing them to deliver drugs, proteins and genetic material to targeted tissues, which makes them useful in cancer medicine, gene therapy and diagnostics. • However, disadvantages of polymeric NPs include an increased risk of particle aggregation and toxicity. Only a small number of polymeric nanomedicines are currently FDA approved and used in the clinic, but still polymeric nanocarriers are undergoing testing in numerous clinical trials Polymer-based nanoparticles approved by FDA 20Lombardo et al. Journal of Nanomaterials 2019, Article ID 3702518. Inorganic nanoparticles 21 • Inorganic materials such as gold, iron, and silica have been used to synthesize nanostructured materials for various drug delivery and imaging applications • Variety of sizes, structures and geometries, e.g., gold NPs can form nanospheres, nanorods, nanostars, nanoshells, and nanocages • Inorganic NPs have unique physical, electrical, magnetic and optical properties, due to the properties of the base material itself • For example, AuNPs possess free electrons at their surface that continually oscillate at a frequency dependent on their size and shape (surface plasmon resonance), giving them photothermal properties • AuNPs are also easily functionalized, granting them additional properties and delivery capabilities Mitchell et al. Nature Reviews Drug Discovery 2021, 20, 101–124. Inorganic nanoparticles 22 • Iron oxide is another commonly researched material for inorganic NP synthesis, and iron oxide NPs make up the majority of FDA- approved inorganic nanomedicines • Magnetic iron oxide NPs — composed of magnetite (Fe3O4) or maghemite (Fe2O3) — possess superparamagnetic properties at certain sizes and have shown success as contrast agents, drug delivery vehicles, and thermal-based therapeutics Mitchell et al. Nature Reviews Drug Discovery 2021, 20, 101–124. Inorganic nanoparticles 23 • Inorganic NPs include calcium phosphate and mesoporous silica NPs, which have both been used successfully for gene and drug delivery • Quantum dots — typically made of semiconducting materials such as silicon — are unique NPs used primarily in in vitro imaging applications, but they show promise for in vivo diagnostics • Due to their magnetic, radioactive or plasmonic properties, inorganic NPs are uniquely qualified for applications such as diagnostics, imaging and photothermal therapies. Most have good biocompatibility and stability, and fill niche applications that require properties unattainable by organic materials. However, they are limited in their clinical application by low solubility and toxicity concerns, especially in formulations using heavy metals Mitchell et al. Nature Reviews Drug Discovery 2021, 20, 101–124. Gold nanoparticles 24 • Noble metal NPs (Au, Pt, Ag) have surface plasmon resonance (SPR, nanosized photon confinement) Wikipedia Gold nanoparticles as drugs 25 • Imaging – light scattering × tumor ablation – photothermal properties (light absorption) • Properties dependent on size and shape of NPs • Traditionally, loading of AuNPs with a variety of therapeutic molecules through surface modification (affinity of AuNPs to thiols) • Limited toxicity to cells • The total extinction of light at the SPR is made up of both absorption and scattering. For the smaller axial diameter nanorods (~10 nm), absorption dominates, whereas for the larger axial diameter nanorods (>35 nm) scattering can dominate. • Aurimune, a tumor necrosis factor-bound AuNP, has completed key clinical phase 1 trials (no drug approved yet) for treatment of cancer Anselmo et al. Am. Assoc. Pharm. Sci. 2015, 17(5), 1041. • AuroLase® is being developed by Nanospectra, are silica-gold nanoshells coated with (poly)ethylene glycol (PEG) designed to thermally ablate solid tumors following stimulation with a nearinfrared energy source 26 • AuNPs capable of treating acne (Sebashells) are being developed by Sebacia Inc • ~150 nm silica-gold nanoshells, coated with PEG • treat acne by disrupting overactive sebaceous glands in the skin Gold nanoparticles as drugs Anselmo et al. Am. Assoc. Pharm. Sci. 2015, 17(5), 1041. Iron oxide nanoparticles 27 • Iron oxide nanoparticles (IONPs), either superparamagnetic iron oxides (SPIO) or ultrasmall superparamagnetic iron oxides (USPIO) • Properties: innate magnetic responsiveness facilitating targeting, imaging, localized heating (hyperthermia, tumor ablation); biocompatibility and biodegradability • IONPS are important contrast agents for noninvasive diagnostic imaging using magnetic resonance imaging (MRI) • These are the most clinically studied inorganic NPs, approved by FDA for imaging of different pathologies and for treatment of iron deficiency (anemia) • Ferumoxytol are USPIO nanoparticles (17-31 nm) coated with polyglucose sorbitol carboxymethylether – approved for treatment of anemia of patients with chronic kidney disease or studied as imaging agent of a variety of diseases and conditions, ranging from: multiple sclerosis to numerous cancers (e.g., prostate, bladder, breast, lung, ovarian, to name a few) to heart conditions to type 1 diabetes • Generally, IONPs could be used to: (i) enhance targeting to diseased tissues via magnetic field, (ii) deliver loaded drugs to this target site, and (iii) be activated externally to destroy surrounding tissue, effectively providing a targeted combination therapy. Anselmo et al. Am. Assoc. Pharm. Sci. 2015, 17(5), 1041. Ferumoxytol imaging 28Anselmo et al. Am. Assoc. Pharm. Sci. 2015, 17(5), 1041. Iron-based nanoparticles approved by FDA 29Anselmo et al. Am. Assoc. Pharm. Sci. 2015, 17(5), 1041. Silica-based NPs 30 • Or silicon dioxide (SiO2) are tunable materials where we can control the size, shape, and porosity + do surface modifications (conjugation of various stealth or targeting ligands to influence circulation, targeting, drug loading and release, and internalization by cells of intravenously administered NPs • Silica NPs have limited toxicity and favorable biodegradability • Synthesis, e.g., by Stober‘s or microemulsion method Selvarajan et al. Front. Chem. 2020, 8, Article 602 Synthesis and surface modification of silica NPs 31Selvarajan et al. Front. Chem. 2020, 8, Article 602 Silica-based NPs (SNPs) in drug delivery 32 • incorporation of hydrophilic or hydrophobic drugs • diagnostic applications, including imaging, gene transfection vectors for cellular uptake, and stimuli-responsive delivery (many SNPs are used in combination with other inorganic materials (e.g., gold as in AuroLase®)) • One of the main challenges facing SNPs is determining their long-term in vivo fate if they are not cleared by the kidneys Selvarajan et al. Front. Chem. 2020, 8, Article 602 In the next class… Photomedicine Thank you for your attention!