Finding membrane proteins in cutaneous T-cell lymphomas

DESCRIPTION (provided by applicant): Cutaneous T-cell lymphoma (CTCL) affects 5000 cases in the United States every year, 20% of which will present with a poor prognosis and a mean survival of 3 years. Despite the impact of biological response modifiers, such as interferon alpha, there still remains up to 25 to 50% of patients who present with advanced disease and who progress to rapidly fatal disease. We propose to directly compare the relative abundance of cell surface proteins between benign and malignant T-cells using the recently developed ICAT TM technology. In the first step we will isolate membrane proteins from malignant cells from two patients representative of the Sezary population and from benign cells from a pool of normal controls. In the second step, the proteins from the malignant cells will be labeled with an isotopically heavy (d8) and the proteins from normal cells with an isotopically light (dO) coded affinity tag (ICATTM), respectively. The proteins of both samples will then be combined and proteolyzed by trypsin. In the third step, this peptide mixture will be separated by multidimensional chromatography, and the fractions analyzed by automated tandem mass spectrometry (MS/MS). The raw data will be searched against protein and DNA databases and the differential peptide representation will assess using application specific software tools. These proteins will become candidate marker antigens, which have potential as targets for diagnostic or therapeutic monoclonal antibodies.