PowerPoint Presentation: Aptamers ( aptus - fit, and Greek meros - region) are Oligonucleic acid or Peptide molecules that bind to a specific target molecule. Aptamers are usually created by selecting them from a large random sequence pool. N atural aptamers also exist in riboswitches . Aptamers can be used for both basic research and clinical purposes as macromolecular drugs . PowerPoint Presentation: Aptamers can be combined with ribozymes to self-cleave in the presence of their target molecule. These compound molecules have additional research, industrial and clinical applications. PowerPoint Presentation: A ptamers can be classified as : DNA or RNA or XNA aptamers . They consist of (usually short) strands of oligonucleotides . Peptide aptamers . They consist of a short variable peptide domain, attached at both ends to a protein scaffold. PowerPoint Presentation: Nucleic Acid aptamers Nucleic acid aptamers are nucleic acid species that have been engineered through repeated rounds of in vitro selection Or SELEX ( systematic evolution of ligands by exponential enrichment ) to bind to various molecular targets such as small molecules, proteins, nucleic acids, and even cells, tissues and organisms. PowerPoint Presentation: Aptamers are useful in biotechnological and therapeutic applications as they offer molecular recognition properties that rival that of the commonly used biomolecule , antibodies. 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. PowerPoint Presentation: In 1990 , the lab independently developed the technique of selection: the Gold lab, using the term SELEX for their process of selecting RNA ligands against T4 DNA polymerase ; PowerPoint Presentation: Since the discovery of aptamers , many researchers have used aptamer selection as a means for application and discovery. In 2001, the process of in vitro selection was automated by the Ellington lab at the university of Texas at Austin , and at SomaLogic , Inc (Boulder, CO), reducing the duration of a selection experiment from six weeks to three days. PowerPoint Presentation: aptamers in the natural world had yet to be uncovered until 2002 when two groups led by Ronald Breaker and Evgeny Nudler discovered a nucleic acid-based genetic regulatory element called a riboswitch that possesses similar molecular recognition properties to the artificially made aptamers . PowerPoint Presentation: Both DNA and RNA aptamers show robust binding affinities for various targets.  DNA and RNA aptamers have been selected for the same target. These targets include lysozyme ,  thrombin ,  human immunodeficiency virus trans-acting responsive element (HIV TAR),  hemin ,  interferon γ ,  vascular endothelial growth factor (VEGF),  and dopamine .  PowerPoint Presentation: Both DNA and RNA aptamers show robust binding affinities for various targets.  DNA and RNA aptamers have been selected for the same target. These targets include lysozyme ,  thrombin ,  human immunodeficiency virus trans-acting responsive element (HIV TAR),  hemin ,  interferon γ ,  vascular endothelial growth factor (VEGF),  and dopamine .  PowerPoint Presentation: Considering that not all DNA analogs of RNA aptamers show functionality the correlation between DNA and RNA sequence and their structure and function requires further investigation. PowerPoint Presentation: In the case of lysozyme , HIV TAR, VEGF and dopamine the DNA aptamer is the analog of the RNA aptamer , with thymine replacing uracil . The hemin , thrombin, and interferon γ, DNA and RNA aptamers were selected through independent selections and have unique sequences. PowerPoint Presentation: Recent developments in aptamer -based therapeutics have been rewarded in the form of the first aptamer -based drug approved by the U.S. Food and Drug Administration (FDA) in treatment for age-related macular degeneration (AMD), called Macugen offered by OSI Pharmaceuticals . PowerPoint Presentation: In addition, Cambridge, MA - based Archemix ( http://www.archemix.com ) is leading the development of aptamers as a new class of directed therapeutics for the prevention and treatment of chronic and acute diseases. ARC1779, its lead proprietary candidate, is a potent, selective, first-in-class antagonist of von Willebrand Factor ( vWF ). ARC1779 is being evaluated in patients diagnosed with acute coronary syndrome (ACS) who are undergoing percutaneous coronary intervention (PCI). Phase I testing for ARC1779 was initiated in December 2006, and a Phase 2 study in ACS is planned to begin by the end of 2007. PowerPoint Presentation: Non-modified aptamers are cleared rapidly from the bloodstream, with a half-life of minutes to hours, mainly due to nuclease degradation and clearance from the body by the kidneys, a result of the aptamer's inherently low molecular weight. Unmodified aptamer applications currently focus on treating transient conditions such as blood clotting, or treating organs such as the eye where local delivery is possible. PowerPoint Presentation: This rapid clearance can be an advantage in applications such as in vivo diagnostic imaging. An example is a tenascin -binding aptamer under development by Schering AG for cancer imaging. Several modifications, such as 2'-fluorine-substituted pyrimidines , polyethylene glycol (PEG) linkage, etc. (both of which are used in Macugen , an FDA-approved aptamer ) are available to scientists with which to increase the serum half-life of aptamers easily to the day or even week time scale. PowerPoint Presentation: Another approach to increase the nuclease resistance of aptamers is to use a Spiegelmer . In addition to the development of aptamer -based therapeutics, many researchers such as the Ellington lab and independently another company SomaLogic (Boulder, CO) have been developing diagnostic techniques for aptamer based plasma protein profiling called aptamer plasma proteomics . PowerPoint Presentation: Peptide aptamers Peptide aptamers are proteins that are designed to interfere with other protein interactions inside cells. They consist of a variable peptide loop attached at both ends to a protamersein scaffold. This double structural constraint greatly increases the binding affinity of the peptide aptamer to levels comparable to an antibody's ( nanomolar range). PowerPoint Presentation: The variable loop length is typically composed of ten to twenty 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. PowerPoint Presentation: Peptide aptamer selection can be made using different systems, but the most used is currently the yeast two-hybrid system . Selection of Ligand Regulated Peptide Aptamers ( LiRPAs ) has been demonstrated. By displaying 7 amino acid peptides from a novel scaffold protein based on the trimeric FKBP- rapamycin -FRB structure, interaction between the randomized peptide and target molecule can be controlled by the small molecule Rapamycin or non-immunosuppressive analogs. PowerPoint Presentation: Peptide aptamer can also be selected from combinatorial peptide libraries constructed by phage display and other surface display technologies such as mRNA display , ribosome display , bacterial display and yeast display . These experimental procedures are also known as biopannings . PowerPoint Presentation: Among peptides obtained from biopannings , mimotopes can be considered as a kind of peptide aptamers . All the peptides panned from combinatorial peptide libraries have been stored in a special database with the name MimoDB ,  which is freely available at http://immunet.cn/mimodb . PowerPoint Presentation: AptaBiD or Aptamer -Facilitated Biomarker Discovery is a technology for biomarker discovery.  AptaBiD is based on multi-round generation of an aptamer or a pool of aptamers for differential molecular targets on the cells which facilitates exponential detection of biomarkers. PowerPoint Presentation: It involves three major stages: ( i ) differential multi-round selection of aptamers for biomarker of target cells; (ii) aptamer -based isolation of biomarkers from target cells; and (iii) mass spectrometry identification of biomarkers. The important feature of the AptaBiD technology is that it produces synthetic affinity probes ( aptamers ) simultaneously with biomarker discovery. PowerPoint Presentation: In AptaBiD , aptamers are developed for cell surface biomarkers in their native state and conformation. In addition to facilitating biomarker identification, such aptamers can be directly used for cell isolation, cell visualization, and tracking cells in vivo . They can also be used to modulate activities of cell receptors and deliver different agents (e.g., siRNA and drugs) into the cells. PowerPoint Presentation: ^ Walsh, R.; M. DeRosa (2009). "Retention of function in the DNA homolog of the RNA dopamine aptamer .". Biochemical and Biophysical Research Communications 388 : 732–735. PMID 19699181 . Cho EJ, Lee JW, Ellington, AD (2009). " Applications of Aptamers as Sensors" . Annual Review of Analytical Chemistry 2 (1): 241–64. doi : 10.1146/annurev.anchem.1.031207.112851 . PMID 20636061 . http://arjournals.annualreviews.org/doi/abs/10.1146/annurev.anchem.1.031207.112851 . PowerPoint Presentation: Aptamers are artificial oligonucleotides (DNA or RNA) selected in vitro that bind a broad range of targets with high affinity and specificity; a sensitive yet simple method to utilize aptamers as recognition elements for the development of biosensors ( aptasensors ) is to transduce the signal electrochemically. PowerPoint Presentation: Aptamer molecules exist in nature in the form of genetic regulators called riboswitches [ 1 ] PowerPoint Presentation: In vitro SELEX has been widely used for the identification of a variety of targets, ranging from small molecules (metal ions, organic dyes, amino acids, or short peptides) to large proteins or complex targets (whole cells, viruses, virus-infected cells, or bacteria). PowerPoint Presentation: The ability of aptamers to selectively bind to different targets is based on their distinct three-dimensional structure, allowing them to form stable and specific complexes with different targets of complementary shape . PowerPoint Presentation: in view of target inhibition, aptamers are different from ribozymes and antisense oligonucleotides , which are used to prevent the translation of genetic information from mRNAs to proteins . The binding affinity of aptamers to their targets is very high, with typical dissociation constants in the picomolar to nanomolar range, depending on the nature of the targets. PowerPoint Presentation: Also, aptamers recognize their targets with extremely high specificity. For example, aptamers can discriminate among homologous proteins that contain only a few amino acid changes PowerPoint Presentation: The molecular recognition properties of aptamers , such as high affinity and specificity, are similar to antibodies, but the unique properties of aptamers set them apart from antibodies. Aptamers are produced by chemical synthesis rather than tedious biological expression. This allows researchers to quickly and reproducibly synthesize any DNA or RNA sequence with little or no batch-to-batch variation. As synthetic molecules, aptamers readily support site-specific modifications toward a specific purpose. PowerPoint Presentation: For research, aptamers can be easily labeled with florescent dyes, biotin or radionuclides . For clinical purposes, aptamers can be conjugated to nanoparticles ,drug molecules, enzymes viruses or small interfering RNAs ( siRNAs ) PowerPoint Presentation: Unlike antibodies, aptamers are very stable across a wide range of temperature or storage conditions. Even thermally-denatured aptamers can return to their original conformation without losing binding affinity by one cycle of heating and cooling, whereas antibodies are temperature sensitive and denaturation is usually irreversible. PowerPoint Presentation: In addition, chemical modifications, such as 2′-fluoro and 2′- O -methyl substitutions, can enhance their biochemical stability against nuclease degradation [ 4 , 15 ]. Furthermore, their small size allows for rapid penetration into tissues and organs, with low toxicity and low immunogenicity, which may facilitate long-term therapeutic efficacy and safety. PowerPoint Presentation: These unique biochemical properties make aptamers highly suitable for the detection, diagnosis and treatment of disease.