Nanoparticles for Drug Delivery

Transcription

1 Introduction Systemic Drug Delivery Examples : Methods of Drug Delivery Advantages of ral Delivery Inhalation Transdermal Implantation Injection University of Illinois, Urbana-Champagne Examples of Examples of iposomal Amphoterin, sold by Gilead (Ambisome) and Enzon (Abelcet) ~$200 Million in Ambisome sales in Applications: Non-resistant cancers Gilead Sciences Amphotericin treats fungal and parasite infection Most commonly used in patients with depressed white blood cell count (cancer and chemotherapy patients, HIV-infected patients, elderly patients). iposomal formulation is preferred because of decreased side effects and prolonged drug exposure (due to slow release) In hepatic metastases model, the reduction in number of metastases was greater with Dox-loaded nanospheres than free dox. (Why hepatic model?) No special affinity for tumor tissue detected. Most nanospheres located within Kupffer cells. See proposed mechanism of action Note that this approach reduces side effects and toxicity! Small molecules are distributed throughout the body. Vauthier, et al. Adv. Drug Del Rev v55: (2003)

2 Examples of Applications: Intracellular Infections Introduction Systemic Drug Delivery Macrophages infected with Salmonella incubated with ampicillin-loaded PACA nanospheres Antibiotic-loaded nanospheres Resistance of many microorganisms to antibiotics is often related to low uptake of antibiotics or reduced activity in acidic ph of lysosomes. 1. Ampicillin-loaded nanospheres for isteria treatment. Dramatic improvement over free drug; bacterial counts in liver reduced at least 20-fold. 2. Ampicillin-loaded nanospheres for Salmonella treatment. Drug alone required 32 mg per mouse; with nanoparticle, only 0.8 mg ensured survival. Vauthier, et al. Adv. Drug Del Rev v55: (2003) Examples : Drug Carrier Systems Generati on First > 1 µm Second < 1 µm Third < 1 µm Size Definition Examples Able to release a drug at the target site but needing a particular type of administration Carriers that can be given by a general route able to transport a drug to the target site Carriers able to recognize a specific target Microspheres and microcapsules for chemoembolization iposomes, nanoparticles, polymer drug carriers Monoclonal antibodies; second-generation carriers with targeted antibodies or other ligands Metal-based nanoparticles ipid-based nanoparticles Polymer-based nanoparticles Biological nanoparticles from Polymeric Biomaterials (2002), 2nd Ed., Edited by S. Dumitriu Metal Nanoparticles for drug delivery Metal Nanoparticles

3 Metal Nanoparticles ipid-based nanoparticles for Drug Delivery DPE DSPC DTAP Note: direct tissue injection ipid-based nanoparticles for Drug Delivery ipid-based nanoparticles for Drug Delivery Deliverables: -Small molecules (Amphotericin B, Daunorubicin, Doxorubicin all approved and marketed drug formulations) Gilead, Alza -Viruses and bacteria (as vaccines) in development -Nucleic acids in development Many formulation methods Mixing lipids together in organic solution. 2. Remove solvent by evaporation 3. Hydration with aqueous solvent containing drug to form multilamellar vesicles 4. Sonication or extrusion are common methods to reduce the size of the liposomes Avanti Polar ipids ipid-based nanoparticles for Drug Delivery Polymer-based nanoparticles for Drug Delivery Drug properties and iposome association Hydrophilic Retained in aqueous interior Slowly released over **may be difficult to get several hours-several days high loading Hydrophobic Inserted into hydrophobic Excellent retention interior of the liposome bilayer **can disrupt liposome at high concentrations Intermediate Rapidly partition between Rapid release from liposomes lipid bilayer and aqueous but ph manipulation or phase formation of molecular complexes can result in good retention Poly(alkylcyanoacrylates) used extensively for tissue adhesives for skin wounds and surgical glue (late 1960 s) Application as drug nanoparticulate carriers (1980s) For detailed review, see: Vauthier, et al. Adv. Drug Del Rev v55: (2003)

4 Polymer-based nanoparticles for Drug Delivery Polymer-based nanoparticles for Drug Delivery Vauthier, (2003) et al. Adv. Drug Del Rev v55: Vauthier, et al. Adv. Drug Del Rev v55: (2003) Polymer-based nanoparticles for Drug Delivery Polymer-based nanoparticles for Drug Delivery DEGRADATIN F NANPARTICES Hydrolysis of ester bond; degradation products (alkylalcohol and poly(cyanoacrylic acid) are eliminated by kidney filtration. Vauthier, (2003) et al. Adv. Drug Del Rev v55: Vauthier, et al. Adv. Drug Del Rev v55: (2003) Biological nanoparticles --Viruses Biological nanoparticles --Viruses Stratagene, Inc. From an excellent review by Thomas et al. (2003) Nature v4:346

5 Biological nanoparticles --Viruses Introduction Systemic Drug Delivery Examples : Thomas et al. (2003) Nature v4:346 Stability of Colloids in Physiological Environments Stability of Colloids in Physiological Environments 1. Attractive van der Waals forces and random Brownian motion cause particle flocculation 2. Stabilization of colloids by electrostatic stabilization (DV theory): -- Charged particles have a counter-ion layer. The charged surface and counter-ion layer is called the electrostatic double layer. For homogenous colloid suspensions, this electrostatic double layer acts as a repulsive force between particles. --The sum of the van der Waals force and the double layer repulsion force gives the DV interaction potential: Aggregation of Colloids A -higher ionic strengths collapse this boundary layer water 150 mm salt M. Nikolaides -at high salt concentration, no stable region -physiologic salt concentration ~150 nm -- in intravenous delivery, this can be a major cause of toxicity Stability of Colloids in Physiological Environments Steric Stabilization 3. Stabilization of colloids by steric stabilization: -- Add polymers to the surface of particles to prevent the particles from coming in close proximity to each other. At these distances, there is not enough attractive force for flocculation to occur. --Note that this phenomena is solvent dependent (affects the structure and interaction of the surface polymers) Steric Stabilization Dispersion Stabilization Choi, et al. J Disp Sci Tech (2003) v24:415

6 Steric Stabilization Steric Stabilization Choi, et al. J Disp Sci Tech (2003) v24:415 gris et al. (1999) Gene Therapy 6:595 (1999). iposome Stability iposome Stability ptimization Chemical stability Reducing oxidation addition of antioxidants, storage at low temperatures and ph 6.5 Removal of water spray drying or lyophilization (but both have to occur under controlled and optimized conditions) Physical stability Electrostatic stabilization Steric stabilization Cream or hydrogel incorporation Mechanisms of Removal from Circulation Introduction Systemic Drug Delivery Examples : Fast removal from circulation -binding to cells, membranes, or plasma proteins -uptake by phagocytes (macrophages) -trapping in capillary bed (lungs) Renal clearance -size restriction for kidney glomerulus is ~30-35 kda for polymers (~20-30 nm) Extravasation -depends on the permeability of blood vessels -capillaries are thought to be more permissive to extravasation -note: for cancer applications this works to our advantage: EPR! from Polymeric Biomaterials (2002), 2nd Ed., Edited by S. Dumitriu

7 Drug Carrier Systems: Influence of Physicochemical Properties Drug Carrier Systems: Influence of Physicochemical Properties Molecular weight -macromolecules smaller than renal threshold are rapidly eliminated -for larger, non-degradable molecules, excretion is useful to decrease toxicity. Kupffer Cells Charge -positively-charged macromolecules will interact with cells and membranes (remember the negatively charged proteoglycans?) -negatively charged macromolecules are picked up by macrophages such as Kupffer cells, that contain polyanion scavenger receptors on their surface. PEGylation -may increase circulation by reducing non-specific protein binding. from Polymeric Biomaterials (2002), 2nd Ed., Edited by S. Dumitriu Drug Carrier Systems: Influence of Physicochemical Properties Biodistribution of PEGylated Polymer-based Nanoparticles 1000 nm 6000 nm 500 nm 200 nm 10,000 nm + -charged - -charged neutral PEGylated Non- PEGylated gris et al. (1999) Gene Therapy 6:595 (1999). Non-PEGylated liposomes Biodistribution of ipid-based Nanoparticles PEGylation Introduction Systemic Drug Delivery Why is there a dose-dependence? PEGylated liposomes Examples Allen, TM and Stuart, DD. iposome Pharmacokinetics In iposomes Ed. Janoff, AS (2000)

8 Non-specific targeting: EPR Effect Non-specific Targeting Tumors generally can t grow beyond 2 mm in size without becoming angiogenic (attracting new capillaries) because difficulty in obtaining oxygen and nutrients. Tumors produce angiogenic factors to form new capillary structures. Tumors also need to recruit macromolecules from the blood stream to form a new extracellular matrix. Permeability-enhancing factors such as VEGF (vascular endothelial growth factor) are secreted to increase the permeability of the tumor blood vessels. This effect is called the enhanced permeability and retention effect (EPR) from Polymeric Biomaterials (2002), 2nd Ed., Edited by S. Dumitriu Targeting igands Targeting Small Molecules Galactose/Glucose/Mannose Folate Peptides RGD Proteins Transferrin Antibodies Ds Choi, et al. J Disp Sci Tech (2003) v24:415 Targeting Introduction Systemic Drug Delivery Examples : Choi, et al. J Disp Sci Tech (2003) v24:415

9 Examples of Examples of DXI DXI Proposed Mechanism of Action Formulation 1. Doxorubicin-containing core 2. ipid Bilayer membrane 3. PEG-coated surface 4. <100 nm Examples of Examples of DXI Pharmacokinetics DXI Toxicity Rats Dogs Mild white blood cell depression Skin toxicity (unique to Doxil) with full recovery. (ong distribution time?) Cardiac toxicity insignificant up to 1500 mg/m 2 ; Much lower toxicity than doxorubicin (lower peak plasma level; decrease availability to cardiac muscle) Hair loss rare; only seen in ~6% of patients Mucositis ulceration of oral mucosa. Dose-limiting toxicity Gabizon, et al. Pharm Res v10:703 (1993) Examples of DXI Efficacy Doxil (n = Topotecan (n = 118) 119) Examples of DXI Future improvements? Time to progression 18.4 wks 18.3 wks RR CR 4* 5* PR 16* 12* TTP Plat Res 12.3 wks 6.5 wks TTP Plat Sens 28.4 wks 28.8 wks SPlat Res 33.4 wks 37.3 wks SPlat Sens 86.1 wks** 63.3 wks *Expressed as percentage. **p =.01. TTP Plat Res = time-to-progression platinum resistant; TTP Plat Sens = time-to-progression platinum sensitive; S = overall survival.

10 H IDEA SYSTEMIC DEIVERY VECTR CYCDEXTRIN PYMER Non-toxic vehicle Non-immunogenic Condensation of DNA Intracellular delivery Targeting ligand Stabilization of particles H2N H H H H H H H H H H H H H H H H H S H H H N N H H S (CH2)6 H S H H H NH2 H H 2 H S + H H NH + H H H H X H H H Ave. MW 5.8 kda, polydispersity index Mw/Mn = 1.12 Degree of polymerization ~ 5 (10 charges/chain) verified by end group analysis and light scattering H NH2 Gonzalez et al. Bioconjugate Chemistry (1999) v10: PARTICE ASSEMBY TRANSFECTIN CMPARISN DNA (a) 1.00E E+07 BHK-21 CH-K1 1.00E+06 Relative ight Units 1.00E E E E E E+00 PEI (b) Background Poly--ysine Superfect (Dendrimer) ipofectamine CD-polymer nm Gonzalez et al. Bioconjugate Chemistry (1999) v10: TXICITY CMPARISN T BHK-21 CES INCUSIN CMPEX FRMATIN Polymer PEI IC 50 =23µM Polycation Superfect IC 50 =11 µm ipid, ipofectamine IC 50 =6.4 µm Cyclodextrin polymer IC 50 =320 µm + IC 50 values reported as charge concentrations in the presence of DNA Association Constant is

11 NEW METHD F SURFACE MDIFICATIN TEM IMAGES F PARTICES Pre-formed Particle Stabilized Particle Targeted, Stabilized Particle A B A. Polyplexes in water B. PEGylated polyplexes in water C. Polyplexes in 50 mm NaCl D. PEGylated polyplexes C D in 50 mm NaCl Space bar is 100 nm except in C where it is 1000 nm Pun and Davis Bioconjugate Chemistry (2002 ) v13:630 Pun and Davis Bioconjugate Chemistry (2002 ) v13:630 Targeted Particles Biodistribution Nucleic acid delivery Nanoparticle delivery Pun, et al. Canc Biol Ther (in press) Pun, et al. Canc Biol Ther (in press) Tumor uptake Nucleic acid delivery Nanoparticle delivery Pun, et al. Canc Biol Ther (in press)