Proteomic Analysis for Biomarker Discovery in Autoimmune Disease

一个从去年开始一直持续到现在的项目是由博士后莱蒂西亚·卡诺（Leticia Cano）发起的。它试图通过结合质谱法的蛋白质分析来识别自身免疫性疾病中的生物标志物。从组织和免疫复合物中分离的蛋白质被用作生物标志物的来源。已经与涉及干燥病、系统性红斑狼疮和炎症性肌炎的几个临床小组（G. Illei （NICDR）、F. Miller （NEIHS）、E. Rushing （AFIP）、Anil Chauhan （Progen Biologics）、Craig Blackstone （NINDS）和Howard Jaffe (NINDS)）建立了合作关系。我们还获得了OHSR批准，可以从合作人体组织网络（CHTN）和AFIP （rush）获取人体样本。从健康对照和疾病对照中获得的组织都经过了检查，以确保我们不仅仅是在识别炎症标志物。

A project beginning last year and continuing to date, was started by postdoctoral fellow Leticia Cano. It attempts to identify biomarkers in autoimmune disease using protein profiling in combination with mass spectrometry. Proteins isolated from tissue and immune complexes are used as a source of biomarkers. Collaborations have been established with several clinical groups involved in Sjogren's disease, systemic lupus erythematosus and inflammatory myositis (G. Illei (NICDR), F. Miller (NEIHS), E. Rushing (AFIP), Anil Chauhan (Progen Biologics), Craig Blackstone (NINDS), and Howard Jaffe (NINDS). We have also received OHSR approval to obtain human samples from the Cooperative Human Tissue Network (CHTN) and AFIP (Rushing). Tissues obtained from healthy controls and disease controls have been examined to ensure that we are not merely identifying markers of inflammation. New ways to separate and profile proteins prior to LC/MS/MS analysis are also being investigated, i.e., chromatofocusing, ion exchange, salt precipitations and affinity columns. As an example of the latter, we have isolated circulating immune complexes (CICs) from the plasma of patients with dermatomyositis (DM) and systemic lupus erythematosus (SLE) using a receptor-based affinity column from ProGen Biologics (Chauhan). We have found many proteins that were previously identified as candidate biomarkers for autoimmune diseases. The presence of these proteins in circulating immune complexes suggests that they are targeted by the immune system and that autoantibody assays for these proteins may be developed as biomarkers for systemic autoimmune diseases. We have begun using ELISAs to verify these findings and to show the utility of this approach in defining new biomarkers for immune-mediated diseases. A Beckman PF2D system for 2D-HPLC separation of intact proteins has been acquired and will be used for examining the spastic paraplegia samples (Blackstone). In a recent collaborations we identified a 225kDa marker belonging to myosin heavy chain IIA (MYHIIA) that reacts specifically with chronic lymphocytic leukemia antibody. The MYHIIA form structures on the surface of the HEp-2 cells which are undergoing stress. This suggests that chronic lymphocytic leukemia antibody may bind to surface markers that are the consequence of stress related cell events.