Live-Cell Assays of ER and Golgi Enzyme Activities

RESEARCH STRATEGY PROJECT SUMMARY ABSTRACTThis Administrative Supplement to our Small Business Innovation Research Phase II project will take advantage of new advances in liquid handling instrumentation to augment our efforts in developing high contentHCSand high throughput screeningHTSsystems for the discovery of novel therapeutics and or modulating systems for use in the treatment of neurodegenerative disorders including GaucherKrabbeFabryTay SachsSandhoff and other lysosomal storage diseases as well as Parkinsonandapos s diseasePDAlzheimerandapos s diseaseADamyotrophic lateral sclerosisALSTypediabetesLowe syndromeHuntingtonandapos s disease and their allied medical conditionsThe ability to accurately plate out equal numbers of live cells intowell orwell tissue culture plates for these HTS and HCS applications is paramount to obtaining consistent and reliable resultsRobust and reliable methods for the manipulation of human cell lines by passagingplating and labeling are required to enable consistentreproducible screens to be performedWith the help of this supplementwe intend to develop procedures and processes to maximize the level of consistency of cell plating and analysisto ensure that the assays which we are running on a routine basis remain consistent across multiple runs and to apply these methods in the screening of large libraries of compoundsThese procedures involve a variety of fully or semi automated stepsusing high quality commercially available liquid handling and dispensing technologiesThereforewe propose the purchase of a Molecular BioProducts ThermoFisher Versetteautomated liquid handling instrumentThe Versettehas replaced the ThermoElectron Multidrop as the state of the art cell dispensing technology at Thermo Fisher research labs and at HCS screening facilities worldwideThe Versette will be integrated with our existing ThermoFisher Countess II FL Cell Counter and CellInsightCXHigh Content ScreeningHCSPlatforms to provide absolute cell countsmeasure viabilitydetermine whether cells to be plated from suspension consist of single cellsand perform the HCS screening and analysis of potential drug candidatesThe ultimate goal and the overall impact of this project is to provide an understanding of the dynamic processes that occur in intracellular enzyme and protein trafficking defects in human disease and to obtain information that will provide a pathway to the discovery of more efficacious treatment optionsThe use of these systems for screening libraries of potential drug candidates for their ability to modulate enzyme functiontrafficking and intracellular localization will identify new small molecule therapeutic leads based on these parametersThese systems will be used in our CRO activities as well as for a variety of significant cell biologybiochemicaldiagnostic and medical applications with our collaborators in both academia and industry NARRATIVE The success of this project provides enormous commercial opportunities for medical intervention of a number of neurological diseases such as the lysosomal storage disorders including GaucherLALDSandhoffTaySachs and Krabbe diseases as well as in related neurological conditions including Parkinson s DiseaseAlzheimerandapos s diseaseADamyotrophic lateral sclerosisALSTypediabetesLowe syndrome and Huntingtonandapos s disease through discovery of new drugs for use in modulating key metabolic enzyme activities and intracellular protein traffickingThese assays will also enable clinicians to determine the efficacy of current and emerging therapies as well as add a new set of clinical diagnostic assays for patient assessmentIn additionit will lead to commercial and licensable products in these areas