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The dreams of targeted drug delivery Drug delivery Drug (pharmaceutical agent, API) chemical substances applied to body where they are metabolized and are expected to have beneficial effect (in treatment, mitigation or prevention of disease or facilitating repair of injury etc.) 2 Conventional drug delivery Problem: lack of selectivity and missing control of the release Targeting efficiency: 0.01% Non cotrolled delivery process. OK for low cost 3 safe drugs and short term treatment. Targeted drug delivery and controlled release – ideal course 4 Way to tissue/cells – carrier – targeting Goal tissue/cells, way of the transport into Goal tissue or cells Carrier Biocompatibility? How carrier recognises the target Kind of carrier Biodegradability? Targeting Tools of transport 5 Way to tissue/cells – obstacles After the application into the bloodstream carrier is exposed to quite a number of pitfalls: Phagocytosis: the first step of phagocytosis is particles opsonization – binding of blood serum proteins on the surface of particles the elimination (clearance) of so stained particles is very quick (liver 80-90%, spleen 5-8%, bone marrow 1-2%) opsonization depends on: size, shape, surface charge and hydrophilic/hydrophobic character of surface – small, uncharged and hydrophilic particles are eliminated slowly protection of carriers: hydrophilization of surface – stealth carriers are used (PEG, dextrans, liposoms, hydrophilic polymeric micelles) positive: up to 100x lower clearance negative: in some cases changes in interaction with receptors 6 Way to tissue/cells – obstacles Trapping by cell receptors: typical mainly for carriers having on the surface biopolymers or other biological active substances (liver, spleen) hyaluronan interacts with HARE receptors in liver, very effective clearance protection of carriers: probably saturation of liver receptors by hyaluronan fragments Elimination from bloodstream by kidney: carriers having diameter ≤ 5 nm are removed into urine by filtration protection of carriers: carrier diameter ≥ 5 nm Recommended carrier diameter (hydrodynamic diameter) 5 (6) – 100 (200) nm 7 Way to tissue/cells – obstacles Organ/tissue barriers: majority of organs/tissues protect their internal environment by various barriers (exception – liver parenchym and several other tissues) barriers could be overcome directly by small lipophilic (steroid hormones) or hydrophilic (glucose) substances if their concentration is high enough (diffusion) blood-brain barrier (BBB) is one of the most effective barriers for overcoming BBB – intranasal carrier application is used, more effective is combination of carrier with penetrating peptides 8 Way to tissue – EPR effect inflammed tissue – capillary fenestration: gaps among endothelial cells (coats of capillaries) using these gaps, macromolecules, carriers and immune cells can penetrate into the tissue it is possible to use this effect for carriers transported into pathological tissue EPR effect could be amplified by the application of inflammation eliciting/supporting substances (TNFα) in solid tumours EPR effect is endorsed by minimal lymphatic system – extension of acting time of cytostatics in tumour despite the EPR effect, only roughly slightly less than 1% of applied cytostaticum reaches tumour EPR effect = enhanced permeability and retention effect 9 Way to tissue/cells – carrier – targeting Goal tissue/cells, way of the transport into Goal tissue or cells Carrier Biocompatibility? How carrier recognises the target Kind of carriers Biodegradability? Targeting Tools of transport 10 Carriers Carriers with effect on the whole system liposomes, polymersomes polymeric micelles microbubbles dendrimers synthetic/peptidic nanotubes pro-drugs Carriers with local effect nano-fibres and micro-fibres 11 Carriers Liposomes, polymersomes: liposomes: most often spheroidal vesicles having a hydrophilic core surrounded by lipidic bilayer similar to the cell membrane which separates the inner part from its surroundings liposomes are made namely from natural or synthetic phospholipids or their derivatives (self – assembling system) they are used namely for the transport of hydrophilic substances, hydrophobic could be transported embeded into lipidic bilayer 12 Carriers Liposomes, polymersomes: polymersomes: amphiphilic diblock copolymers is used for the formation of monolayers at the oil-water interface. After evaporation polymer bilayers are formed 13 Carriers 14 Carriers Polymeric micelles: polymers with one part being hydrophilic and second part being hydrophobic (amphiphilic polymers) form in water polymeric micelles, where hydrophobic parts form a core and hydrophilic shell hydrophobic drugs can be solubilized in the hydrophobic core while the hydrophilic shell stabilizes carriers in water solution block copolymer or grafted polymers could be used for the formation of polymeric micelles 15 Carriers Microbubbles: microbubbles are small (1-5 microns in diameter), lipid or albumin shelled gas bubbles which are strong reflectors of sound due to the influence of ultrasound waves, collapse of the bubbles occurs which leads to: 1. generation of high temperature in the core of bubbles 2. shock waves are generated able to mechanically destroyed surrounding tissue (support for better drug penetration into tissue) 3. fragments of bubble wall carrying active substances are, in principal, micro-projectiles (shrapnel) able to transport drugs into surrounding tissue 16 Carriers Dendrimers: polymeric substance prepared by chemical synthesis they are effective namely in nucleic acid (genes) and special substances delivery active substances are incorporated/encapsulated into the structure where they are stabilized by ionic interaction or by van der Waals forces size in tens of nanometres some of them show the problem with biocompatibility 17 Way to tissue/cells – carrier – targeting Goal tissue/cells, way of the transport into Goal tissue or cells Carrier Biocompatibility? How carrier recognises the target Kind of carrier Biodegradability? Targeting Tools of transport 18 Carrier targeting via specific structures on the cell surface for the „real“ targeting of drug, each carrier has to be furnished with specific molecules able to recognise and bind to typical structures on the cell surface Most common molecules are: ligands for receptors antibody against surface antigens Drug lectins peptides aptamers 19 Carrier targeting via specific structures on the cell surface cell receptors specific for certain molecules (hyaluronan – CD 44, mannose – mannose receptor on macrophages, galactose –galactose receptor on hepatocytes) monoclonal antibodies against cell surface antigens 20 Carrier targeting via specific structures on the cell surface Lectins: groups of glycoproteins that bind specifically and reversibly to sugar moiety can interact with glycoconjugates present on the cell surface. The unique carbohydrate specificities of lectins can facilitate carrier targeting. Peptides: RGDXY peptide for interaction with integrins (specific integrin of cancer cells αVβ3) binding sites on the surface of some cells pro specific peptide sequences (NPNWGPR at melanoma) Aptamers: synthetic oligonucleotides that bind to a specific targets such as small molecules, proteins, nucleic acids, cells and carriers they reveal high specificity, affinity and thermal stability and low 21 immunogenicity Non-specific carrier internalization Phagocytosis: non-specific internalization of largest particles (2 – 3 μm), macrophages, dendritic cells Pinocytosis: mechanism by which cells ingest extracellular fluid and its contents (solubilized molecules, not particles) Macropinocytosis: nonspecific internalization of smaller particles ~ 1 μm Cell penetrating peptides: CPP (next page) 22 Non-specific carrier internalization - CPP Cell penetrating peptides: group of peptides derived from peptides of viruses peptides having as far as 30 amino acids their primary structure is rich in arginine non-immunogenic, low cytotoxicity penetrates through cell membrane of different cells, do not invade their structure and stability they can transfer across cell membrane small molecules, biopolymers and nanoparticles (carriers) Penetratin: RQIKIWFQNRRMKWKK, Tat peptide: YGRKKRRQRRR, short CPP peptide VPMLK, 23 Biocompatibility and biodegradability Biodegradability enzymatically or hydrolytically fully degradable material used for carrier preparation fragments after degradation are metabolized or excluded from the organism Biocompatibility it is the ability of material not to negatively influences processes running in the tissue (not to initiate pathological processes) it is specific interaction between this tissue and that material (and its standard impurities) and its exact form. It is very difficult to say that this material is biocompatible without specification of tissue or cells 24 Controlled drug release from carrier External impulses λNIR Redox Internal impulses pH in cells compartments: • physiological conditions about 7.3-7.4 • cancer tissue about 6.5 • endosome about 5.5 • lysosome 4.6-5,5 Carrier contains enzyme Carrier contains substrate for enzyme 25 „Classical carriers“ and theranostics „Classical carrier“ Theranostics carrier is targeted into selected tissue/cells carrier is able to release drug by controlled manner drug release is controlled by internal and external impulses carrier is targeted into selected tissue/cells carrier is able to release drug by controlled manner drug release is controlled mainly by external impulses carrier is furnished by the system for localization of its position in the body and allowing to quantify amount of theranostics in focused tissue 26 Theranostics For someone a diagnostic agent really informs about illness development for others it is system giving us the information about carrier position and its quantity in the body/tissue magnetic nanoparticles could be used mainly for localization and quantification of carriers in tissue fluorescent substrate for caspase 3 could be used primarily for quantification of apoptosis in the tumour tissue 27 Theranostics 28 Carrier from Contipro C6+NR Water C6em C18+NR C18em Oil Carrier from Contipro Thank you for your attention NF-CZ07-ICP-1-040-2014: Formation of research surrounding for young researchers in the field of advanced materials for catalysis and bioapplications 31