targeting
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A nanomedical device should perform multiple functions, so a multifunctional nanoparticle system can be constructed from the inner core outward. The core can contain a drug, while outer layers provide targeting ligands, such as antibodies, peptides, or aptamers directed against receptors overexpressed on diseased cells. Aptamers have advantages over antibodies like uniform activity between batches. Peptides can also serve as targeting ligands if they recognize receptors. Other ligands like folate or transferrin can bind their cognate receptors, but compete with circulating levels; antibodies are more specific but expensive. A variety of conjugation strategies can be used to attach targeting ligands to nanoparticles.
![LNA
• A locked nucleic acid (LNA), often
referred to as inaccessible RNA, is a
modified RNA nucleotide. The
ribose moiety is modified with an
extra bridge connecting the 2'
oxygen and 4' carbon. LNA
nucleotides can be mixed with DNA
or RNA residues in the
oligonucleotide whenever desired.
Such oligomers are commercially
available. The locked ribose
conformation enhances base
stacking and backbone pre-organization.
This significantly
increases the hybridization
properties (melting temperature)
of oligonucleotides.[1]](https://image.slidesharecdn.com/3targeting-141110133449-conversion-gate01/85/targeting-47-320.jpg)
targeting
- 1. 1 Surface Modification of Nanoparticles for Biomedical Applications 1
- 2. Multifunctionalization • A nanomedical device should perform several functions alltogether
- 4. • Consequently a multi-component nanomedical system can be constructed in reverse order of controlling events, namely from the inside out. The outer components are the first to be used. The inner components are the last.
- 5. Possible steps It is possible but rare for a single molecule to perform two or more functions
- 7. lipids, polimers (biocompatible-biodegradable materials) Also water is available (liposomes)
- 8. The drug can be inserted into the core
- 14. Targeting molecules A. antibodies B. Peptides C. Aptamers D. Other ligands
- 15. Ligand should be selected to target cell membrane surface molecules that 1)are physiologically overexpressed on healthy target organs or cells ( e.g. Transferrin receptor on blood brain barrier) or 2) are oversexpressed as a consequence of a pathology (e.g. tumour markers)
- 16. Antibodies • Antibodies directed against tissue-specific antigens.
- 17. Examples of antibodies against: Receptors: Vascular endothelial growth factor (VEGF); folate (highly expressed in tumours); Transferrin, opiod peptides (Brain), Apolipoproteins( Brain) , Human epidermal growth factor (EGF) αvβ3 Integrin Matrix metalloproteinases
- 20. APTAMERS
- 21. Aptamers are oligonucleotides that bind a specific target molecule. Aptamers are usually created by selecting them from a large random sequence pool, but natural aptamers also exist in riboswitches. Aptamers can be used for both basic research and clinical purposes as macromolecular drugs.
- 22. aptamers offer advantages over antibodies as they can be engineered completely in a test tube, are readily produced by chemical synthesis, possess desirable storage properties, and elicit little or no immunogenicity in therapeutic applications.
- 24. Aptamer target protein or molecule Application PSMA Prostate cancer diagnosis and therapy WT1 Understanding Wilm's tumor pathogenesis 4,4′-methylenedianiline Detecting DNA-damaging compounds VEGF Inhibiting angiogenesis RET Inhibition of pro-growth signaling HER-3 Reducing drug resistance in HER-2+ cancers TCF-1 Colon cancer growth inhibition Tenascin-C Glioblastoma (brain cancer) detection MUC1 Breast, pancreatic, ovarian cancers; targeting demonstrated PDGF/PDGFR Improving transport to tumors and targeting brain cancers NF-κB Targeting a transcription factor implicated in many diseases Raf-1 Inhibiting pro-growth signaling αvβ3 integrin Targeting tumor-associated vasculature Human keratinocyte growth factor Inhibiting pro-growth signali
- 25. Properties of aptamers versus antibodies Aptamers Binding affinity nanomolar to picomolar Selection is a chemical process carried out in vitro and can therefore target any protein Can select for ligands under a variety of conditions for in vitro diagnostics Uniform activity regardless of batch synthesis PK parameters can be changed on demand Investigator determines target site of protein Wide variety of chemical modifications to molecule for diverse functions of molecule Return to original conformation after temperature insult Unlimited shelf-life No evidence of immunogenicity Antibodies Binding affinity nanomolar to picomolar Selection requires a biological system, thus it is difficult to raise antibodies to toxins (not tolerated by animal) or non-immunogenic targets. Limited to physiologic conditions for diagnostics Screening monoclonal antibodies time consuming and expensive Activity of antibodies vary from batch to batch Difficult to modify PK parameters Immune system determines target site of protein Temperature sensitive and undergo irreversible denaturation Limited shelf-life Significant immunogenicity
- 26. PEPTIDES • Peptide sequences recognized by receptors responsible of binding can be identified and synthesized. • Examples are peptide sequences derived from ApoE apolipoprotein that are recognized by LDL receptor on cell membranes
- 27. Peptides aptamers • Peptide aptamers consist of a variable peptide loop attached at both ends to a protein scaffold. This double structural constraint greatly increases the binding affinity of the peptide aptamer to levels comparable to an antibody's (nanomolar range).The variable loop length is typically comprised of 10 to 20 amino acids, and the scaffold may be any protein which has good solubility and compacity properties. Currently, the bacterial protein Thioredoxin-A is the most used scaffold protein, the variable loop being inserted within the reducing active site, which is a -Cys-Gly-Pro-Cys- loop in the wild protein, the two Cysteines lateral chains being able to form a disulfide bridge.Peptide aptamer selection can be made using different systems, but the most used is currently the yeast two-hybrid system.
- 28. In vivo phage display Bacteriophage is a virus that infects and replicates within a bacterium Phage display technology is based on the ability to express foreign (poly)peptides as fusions to capsid proteins on the surface of bacteriophage A phage random peptide library displays as many as 1011 different peptides
- 29. In vivo phage display Tissue or vascular targeting ligand•Specific organs or tumors•Tumor blood vessels •Ischemic or inflammatory lesions
- 30. Peptides vs. antibodies • Smaller size; better tissue penetration • Less possibility of immunogenicity • Less possibility of liver and bone marrow toxicity • Easier processing and lower production cost • Small molecule peptide mimmeticsavailable • Fast blood-pool clearance; less background • Weaker affinity to antigen (epitope)
- 31. OTHER LIGANDS • Natural ligands for receptors can be employed to functionalize NP surface . Examples: Folate …..binds to folate receptor ApoE ……. binds to LDL receptor Trasferrin…. binds to Tf receptor
- 32. • Problems : competition from circulating Folate, ApoE(lipoproteins), Transferrin
- 33. Antibodies vs physiological ligands
- 34. Antibodies are mores specific than natural ligands Much more expensive Approach: 1- to eliminate physiological competitor in blood, 2- to inject NP functionalized with the ligand
- 43. biotin nanoparticle nanoparticle biotin streptavidin
- 44. streptavidin antibody nanoparticle biotin antibody 100 nm 10 nm
- 47. LNA • A locked nucleic acid (LNA), often referred to as inaccessible RNA, is a modified RNA nucleotide. The ribose moiety is modified with an extra bridge connecting the 2' oxygen and 4' carbon. LNA nucleotides can be mixed with DNA or RNA residues in the oligonucleotide whenever desired. Such oligomers are commercially available. The locked ribose conformation enhances base stacking and backbone pre-organization. This significantly increases the hybridization properties (melting temperature) of oligonucleotides.[1]
- 53. cystein