Discovery experiment stage

Primary validation— the second part of discovery where a larger number of samples are used to verify or eliminate potential biomarkers. This type of experiment is normally focused on the best (statistical) biomarkers observed in the discovery or identified in the discovery experiment and may use different methodology to facilitate a larger number of samples to be used to increase the statistical confidence and eliminate weak or poor biomarkers from the discovery experiment. This is also the stage of the experiment where one starts using samples from related conditions, other geographical areas, etc. The goal of the validation experiment is to focus on the potential biomarkers that are most likely to answer the research question, and to reduce the number of possible biomarkers from the discovery phase before further time, effort, and money are invested in purification, identification, and characterization of the molecule (30-50 samples). 

Two rules have been kept in mind when thinking about samples for biomarker discovery and primary validation. These rules are designed to reduce the complexity of discovery experiments by reducing the number of artifact and false markers that are the result of poor sample collection and handling processes, which often contribute to problems in statistical data analysis. These artifact markers can also mask real biomarkers and cause them to be ignored or missed in the early stages of the project. 

As previously discussed, compound form differs markedly between early discovery and the late discovery/development interface. The early discovery compound is poorly characterized as to its crystalline form - it may be nonsolid, amorphous, or possibly crystalline but uncharacterized as to polymorphic form. The late discovery/development interface compound is crystalline as defined by phase-contract microscopy or powder X-ray diffraction, and its polymorphic and salt form is frequently characterized. This difference has profound implications for the design of a discovery solubility assay. The key question is this Is it better to design an early discovery solubility assay as a separate type of experiment, or is it better to try to automate a traditional thermodynamic solubility assay to handle the very large number of compounds likely to be encountered in early discovery Another way to state this question is this Does it make sense to run a thermodynamic solubility assay on poorly crystalline early discovery compounds This is the type of question about which reasonable people could disagree. However, this author does have a distinct opinion. It is much better to set up a distinctively different solubility assay in early discovery and to maintain a clear distinction between the assay type appropriate in early discovery and the assay type appropriate at the late discovery/ development interface. Two issues are relevant to this opinion One relates to the need for a solubility assay to reflect/predict early discovery stage oral absorption and the other relates to people/chemistry issues. 

In a more traditional pharmaceutical setting, this characterization would be done during preformulation studies. With the availability of automation and the ability to conduct most of these experiments with small quantities of material, more preformulation activities are being shifted earlier into drug discovery. Recently, Balbach and Korn37 reported a "100 mg approach" to pharmaceutical evaluation of early development compounds. Additional absorption, metabolism, distribution, elimination, and toxicity38 screens may also be conducted at this stage. 

Chapter 2 systematically defines some of the important PK parameters and guides the reader through the types of quantitative LC-MS experiments performed to elucidate the PK parameters necessary to move a drag through discovery, preclinical development, and clinical stages. Chapters 3, 4, and 5 respectively introduce the readers to quadmpole mass filters and liner ion traps, time-of-flight mass... 

The terror comes with the discovery of transience. Nothing is fixed, no form solid. Everything you can experience is "nothing but" electrical waves. You feel ultimately tricked. A victim of the great television producer. Distrust. The people around you are lifeless television robots. The world around you is a facade, a stage set. You are a helpless marionette, a plastic doll in a plastic world. 

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