Mass spectrometry - based proteomics currently lacks a robust technology for performing affinity separation in a multiplex format (10's to 1000's of analytes), which would be analogous to planar microarrays and suspension bead arrays on the fluorescence detection platforms. Achieving better integration of affinity separation with mass spectrometry will benefit both bottom-up and top-down proteomics and facilitate progress in basic research in systems biology, preclinical drug discovery, disease diagnostics and reagent development. Adeptrix is developing an analytical assay technology termed Affinity Bead-Assisted Mass Spectrometry for PTM profiling, or Affi-BAMS PTM. Based on our proprietary multiplexed sample separation and analytical workflow, this assay will enable targeted quantification of protein post-translational modifications (PTMs) in key cellular signalin pathways without using liquid chromatography. Affi-BAMS PTM achieves purification of analytes on single affinity beads in a bead suspension format followed by analyzing individual beads in a bead array format. It includes a key step of controlled elution of analytes onto a mass spec-compatible surface of a picotiter plate, which effectively converts an array of beads into a spatially related array of microspots analyzable by MS. The elution step ensures compatibility with all bead types used in the proteomic assays including materials known for poor compatibility with MS, e.g. polystyrene and agarose. Affi-BAMS PTM is designed for both Matrix-Assisted Laser Desorption Ionization (MALDI) and ElectroSpray Ionization (ESI) MS, the latter made possible on the recently introduced imaging ESI platforms: MALDESI, LAESI, DESI and flow probe. In Phase II we will produce a chromatography-free mass spectrometric assay for discovering substrates of protein phosphorylation in key protein sites within the Akt, MAPK, NF-?B and Jak/Stat kinase signaling pathways in response to the disease states and/or drug treatments in cell cultures and animal models. The subsequent technology development will include the assays for measuring protein phosphorylation within the DNA Damage, Cell Cycle Control, Apoptosis, Autophagy, Ser/Thr kinase and Tyr kinase signaling pathways, as well as detecting other types of PTMs, e.g. methylation, acetylation and ubiquitination. The analytical performance of Affi-BAMS PTM will be evaluated in the context of an ongoing drug repositioning study for novel therapeutics for kidney, pancreatic and prostate cancers.
We are developing a new analytical assay that will enable cost-effective monitoring of protein phosphorylation in a large number of protein sites within key cellular signaling pathways. This technology could be highly useful in drug discovery and drug repositioning research with potential applications also in disease diagnostics.
