Progress in sequencing the human genome has led to a new goal - expressing and characterizing the human proteome. A promising area where proteomics can have a major impact is autoimmune disease. Proteome-wide screening against sera from patients with particular autoimmune diseases can enable the discovery of new autoantigens, the development of microarray-based diagnostic assays, and effective treatments for autoimmune disease. During Phase I, we developed a novel, low-cost, high throughput approach for proteomics based on Bead Sorted Libraries of In Vitro Expressed Proteins (BS-LIVE- PRO). These protein libraries can be expressed inexpensively in a single cell-free translation reaction using a Bead Sorted Library of In Vitro Expressible DNA (BS-LIVE-DNA) as a template. Additional novel technologies developed during Phase I which augment this approach include: i) solid-phase PCR to produce BS-LIVE-DNA from cDNA libraries, ii) a method (PC-PRINT) to rapidly phototransfer the protein on each bead in the BS-LIVE-PRO onto discrete spots on a microarray surface using proprietary photocleavable linkers, and iii) methods to decode the randomly arrayed spots generated by PC-PRINT using photo-transferable DNA or mass-tags (PC-CODE). We propose to extensively optimize and evaluate this new technology during Phase II with the aim of commercializing BS-LIVE-PRO. We will apply BS-LIVEPRO to autoimmune diseases in collaboration with Dr. Donald Bloch at the Massachusetts General Hospital, a leading expert on autoantigen discovery and primary biliary cirrhosis. Similar studies will be carried out on other vasculitis autoimmune diseases in association with Dr. Peter Merkel, Director of the Vasculitis Research Consortium. ? Proteome-wide screening against sera from patients with particular autoimmune diseases can lead to the discovery of new autoantigens, the development of diagnostic assays to identify autoimmune disease and, ultimately, improved treatments. We have developed a novel, low-cost, high throughput approach for proteomics based on Bead Sorted Libraries of In Vitro Expressed Proteins (BS-LIVE-PRO). We propose to extensively optimize and evaluate this new technology during Phase II with the aim of commercializing diagnostic assays for autoimmune diseases. ? ? ?
| Norman, Gary L; Yang, Chen-Yen; Ostendorff, Heather P et al. (2015) Anti-kelch-like 12 and anti-hexokinase 1: novel autoantibodies in primary biliary cirrhosis. Liver Int 35:642-51 |
| Lim, Mark; Rothschild, Kenneth J (2008) Photocleavage-based affinity purification and printing of cell-free expressed proteins: application to proteome microarrays. Anal Biochem 383:103-15 |
