Global Proteomic Screening by MALDI Spectrometric Imaging of Protein-Bead Arrays

DESCRIPTION provided by applicant The advent of high density DNA microarrays almost years ago has been an essential step in the genomics revolution Major successes include genome wide gene expression profiling GEP which has led to a new understanding of cellular control pathways and powerful diagnostic prognostic tests such as for predicting breast cancer recurrence In principle a similar approach namely the utilization of high density protein microarrays could herald a revolution in proteomics particularly in the field of cancer research Potential applications include measuring protein protein interactions immune response profiling aimed at discovery of tumor autoantibody biomarkers and profiling enzyme substrate interactions involved in signal transduction e g kinase substrate interactions However despite continued progress high density protein microarrays have not yet lived up to their original promise Current limitations include low array density poor reproducibility high cost poor assay kinetics and difficulty in detecting a diversity of bait prey interactions including enzyme induced protein modifications In contrast mass spectrometry used in conventional proteomics provides many important capabilities including protein identification by sequencing and fingerprinting as well as molecular level identification of protein modifications e g phosphorylation yet does not have the parallelization and high throughput capacity of microarrays During Phase I we evaluated the feasibility of a new proteomic approach developed by AmberGen termed Bead based Global Proteomic Screening Bead GPSandquot This method combines the advantages of mass spectrometry MS and microarray technology by using correlated MALDI mass spectrometric imaging MALDI MSI and fluorescence imaging of photocleavable Mass Tag coded random bead arrays All of the specific aims were achieved including i Demonstration of Bead GPSandquot synchronized fluorescent and MALDI MS imaging ii Demonstration of high capacity coding of protein peptide bead libraries iii Development of improved photocleavable linkers iv Development of improved micro well substrates for the bead arrays v Demonstration of the feasibility to use Bead GPSandquot to screen large combinatorial synthesized one bead one compound OBOC bead libraries comprised of photocleavable peptides biomimetic peptides During Phase II we will further optimize major components of Bead GPSandquot including the construction of photocleavable bead libraries of peptides and peptoids formatting the libraries into high density random bead arrays and obtaining correlated fluorescence and MALDI MS images of the bead arrays to identify and decode bio molecular interactions of andquot preyandquot molecules with the bead library the andquot baitandquot Specific emphasis will be on validating Bead GPSandquot for two applications relevant for cancer research and clinical diagnosis The first application involves use of Bead GPSandquot to screen mid size parallel synthesized photocleavable peptide bead libraries and then larger combinatorial synthesized OBOC libraries for kinase substrate interactions The second application is the use of Bead GPSandquot to screen large combinatorial OBOC biomimetic peptide libraries peptoids against patient sera for the discovery of surrogate antigens that can be used as biomarkers in cancer immunoassays In order to accelerate the development and commercialization of Bead GPSandquot AmberGen will work a closely with Dr Ron Zuckermann Director of the Biological Nanostructure Facility at the Molecular Foundry located at Berkeley and a recognized pioneer in the field of biomimetic peptide synthesis Dr Cathy Costello Director Center for Biomedical Mass Spectrometry at Boston University Medical School and a leading expert in the field of MALDI and its biomedical applications Dr Gary Kruppa V P of Business Development at Bruker Daltonics Billerica MA a world leading provider of MALDI MS instrumentation and Dr Paul Billings the Chief Medical Officer at Life Technologies a leading company in marketing biotechnology products and services PUBLIC HEALTH RELEVANCE Although high density DNA microarrays introduced almost years ago have had a major impact in facilitating the genomic revolution a similar impact has not yet occurred in the field of proteomics despite the availability of commercial high density protein microarrays We will further develop in Phase II a new approach for proteomics termed Bead based Global Proteomic Screening Bead GPSTM which overcomes existing limits in proteomic technology by combining the advantages of MALDI mass spectrometric imaging and microarray technology Potential benefits of the new approach include the discovery of new biomarkers for cancer diagnostics and increased understanding of the causes of cancer