Drug discovery - hit throughput screening, concept of rational drug design.pptx

hit throughput screening concent of drug design

1. PRINCIPLES OF DRUG DISCOVERY SHAMA M KILLE NU24PHPY12 HIGH THROUGHPUT SCREENING

2. TABLE OF CONTENTS INTRODUCTION AND SHORT HISTORY DEFINITION AND SCOPE OF HIGH THROUGHPUT SCREENING INSTRUMENTATION TECHNIQUE AND PROCEDURE IMPORTANCE AND APPLICATIONS OF HTS LIMITATIONS OF HTS REFERENCE

3. INTRODUCTION AND SHORT HISTORY High Throughput Screening (HTS) is a drug-discovery process widely used in the pharmaceutical industry. It leverages automation to quickly assay the biological or biochemical activity of a large number of drug-like compounds. It is a useful for discovering ligands for receptors, enzymes, ion-channels or other pharmacological targets, or pharmacologically profiling a cellular or biochemical pathway of interest. Typically, HTS assays are performed in "automation-friendly" microtiter plates with a 96, 384 or 1536 well format.

4. DEFINITION HTS is process by which large nos. of compounds are rapidly tested for their ability to modify the properties of a selected biological target. HIGH THROUGHPUT SCREENING (HTS) is identification of one or more positive candidates extracted from a pool of possible candidates based on specific criteria. It is a drug-discovery process widely used in the pharmaceutical industry. It allows automation to quickly assay the biological or biochemical activity of a large number of compounds.

5. Basically helps to identify a compound that can chemically modify a target. This compound is identified as a hit and may be generated to a lead. A Hit is any compound that is confirmed to have binding activity to the target and appears on High throughput screen. It gives the desired effect of the HTS experiment and is confirmed on re-testing. The Lead is the compound with therapeutic or pharmacological activity but suboptimal structure that still requires modification.

6. High-Throughput Screening For Drug Discovery: Why High-Throughput Screening need arises? Fact 1-recent understanding of disease mechanism has dramatically increased no. of protein targets for new drug treatment. Fact 2-new technologies have increased the no. of drugs that can be tested for activity at these targets.

7. GOALS OF HTS: Goal is to identify ‘hits’ or ‘leads’ affect target in desired manner active at fairly low concentrations ( more likely to show specificity) new structure It is a useful for discovering ligands for receptors, enzymes, ion-channels or other pharmacological targets, or pharmacologically profiling a cellular or biochemical pathway of interest. HTS= 50,000 –100,000 compounds can be screened daily.

8. The majority of drug targets are: a) G-Protein coupled receptors. b) Nuclear receptor. c) Ion channels. d) Enzymes.

9. INSTRUMENTATION MICROTITER PLATES (ASSAY PLATES) Plates/containers made of plastic, having spaced wells — up to 384, 1536 or 3456 wells. They would contain solvents (e.g. DMSO + test compounds) They would also contain proteins, cells, etc. to be analysed. Some might be kept empty or contain pure solvents to serve as controls.

10. TECHNIQUES AND PROCEDURE TYPES OF HTS: Functional and Non-functional. Functional: Study exactly how the compound interacts with target Non-functional: To find out if the compound interacts with target or not.

11. PROCEDURE

12. High-throughput screening in drug discovery is used to screen: Novel biological active compounds Natural products Combinatorial libraries (Ex: peptides; chemicals) Biological libraries DNA chips RNA chips Protein chips

13. High-throughput screening main lab ware is the microtiter plate. Modern microplates for high-throughput screening assays are performed in automation-friendly microtiter plates with a 96, 384, 1536 or 3456 well format. These wells contain experimentally useful matter, often an aqueous solution of dimethyl sulfoxide (DMSO). For most drug discovery labs, the library collection has grown from 400,000 to 1 million or more compounds. The standard paradigms used to screen these libraries have evolved to automated 384 wells or higher density single compound test formats.

14. Primary screen is designed to rapidly identify hits from compound libraries. The goals are to minimize the number of false positives and maximize the number of confirmed hits. Depending on the assay, hit rates typically range between 0.1 –5 per cent. This number also depends on the cutoff parameters set by the researchers, as well as the dynamic range of a given assay. Primary screens are run in multiplets of single compound concentrations. Hits are then retested, usually independently from the first assay. If a compound exhibits the same activity, it is coined as confirmed hit, which proceeds to secondary screens or lead optimization. The results from lead optimization are used to decide which substances will make it on to clinical trials.

15. In combination with bioinformatics, it allows potential drugs to be quickly and efficiently screened to find candidates that should be explored in more detail. Initial screening of these compounds for their binding ability is the job for high-throughput screening. The key to high-throughput screening is to develop a test, or assay, in which binding between a compound and a protein causes some visible change that can be automatically read by a sensor. Typically the change is emission of light by a fluorophore in the reaction mixture.

16. One way to make this occur is to attach the fluorophore to the target protein in such a way that its ability to fluoresce is diminished (quenched) when the protein binds to another molecule. A different system measures the difference in a particular property of light (polarization) emitted by bound versus unbound fluorophores. Bound fluorophores are more highly polarized and this can be detected by sensors.

17. Detection methods in HTS: Spectroscopy Mass spectroscopy Chromatography Calorimetry X-ray diffraction Microscopy Radioactive Methods

18. Use of robotics Robotics and automated systems are and impotent component of HTS. They optimize the process and save manpower. Robot arms can be used effectively to transfer microtiter plates to and fro the sampling, incubation and analysing spots.

19. IMPORTANCE AND APPLICATIONS OF HTS High-throughput technology can be also use in other areas besides drug development Genomics application DNA sequencing Protein analysis To screen for all kind of novel biological active compounds (libraries): • Natural products • Combinatorial Libraries (peptides, chemicals…) • Biological libraries

20. To screen Micro arrays such as: • DNA chips • RNA chips • Protein chips RECENT ADVANCEMENTS Use of living organisms in HTS to study drug action and identify lead molecules e.g. in Zebrafish and C aenorhabditis el egans . Ultra HTS where above 100,000 compounds are screened at a time; up to 300,000.

21. ADVANTAGES OF HTS High sensitivity of assay (single molecule detection) High speed of assay (automation) Minimization of assay (microtiter plate assay) Low background signal Clear message (best: Yes/No answer) Low complexity of assay (specific interaction) Reproducibility Fast data processing of results Acceptable costs.

22. LIMITATIONS OF HTS High cost Contamination of samples is possible. Low data quality Analysis of data and selection of relevant data from large moulds of data requires patience, professionalism, dedication and true expertise.

23. References: https://www.sinqerinstruments.com/resource/what-is-hiph-throughput-screening/ https:// southernresearch.org/news/nih-contract-hiah-throughput-screening-for-zika/ Szymański P, Markowicz M, Mikiciuk-Olasik E. Adaptation of high-throughput screening in drug discovery—toxicological screening tests. International journal of molecular sciences. 2011 Dec 29;13(1):427-52. Hagemeyer A, Strasser P, Volpe Jr AF, editors. High-Throughput Screening in Chemical Catalysis: Technologies, Strategies and Applications. John Wiley & Sons; 2006 Mar 6.