Development of immunoassay to measure a novel TNF-like protein
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BIOPOWERTECH, 4734 BLUEGRASS PKY, TUSCALOOSA, AL, 35406
AbstractDESCRIPTION (provided by applicant): The tumor necrosis factor (TNF) family cytokines (ligands) mediate diverse immune responses via the activation of their cognate receptors expressed on the cell surface. Dysregulation and genetic defects of these cytokin es are known to be involved in various immune diseases. Adversary effects of increased TNF expression in inflammatory diseases can be readily evidenced by the success of TNF inhibitors to treat diseases including rheumatoid arthritis and inflammatory bowel disease. We have reported the discovery of TL1A, a TNF-like cytokine. TL1A mediates cellular effects via its cognate receptor DR3 (death receptor 3). TL1A induces both apoptosis and inflammatory responses in vivo and in vitro. Tissue localization of TL1A has been reported in inflammatory bowel diseases and atherosclerotic plaques thus indicating that TL1A might play adversary roles in these diseases. TL1A is known to be mainly present as a soluble form but so far, there have been no reports on the presence of soluble TL1A in the body. This was largely due to the lack of a sensitive and specific assay to detect soluble TL1A. This prompted us to report a prototype immunoassay to demonstrate the feasibility of measuring soluble TL1A released in human cell cult ure supernatant. However, as TL1A is known to bind to another soluble protein DcR3 that is known to be present in a relatively high level in human serum, the development of an assay to recognize soluble TL1A in the serum, either unbound or bound to DcR3, h as become an important issue. Therefore, we propose herein to develop a rapid ELISA and characterize it so that the assay can detect a total TL1A expression even in the presence of excess level of DcR3 in the serum. The assay will be also validated in acco rdance with the FDA regulatory guidelines so that it can be suitable for commercialization as well as future clinical use. In addition, we propose to carry out an exploratory study to screen a large number of coded archived clinical sera of different diagn oses in order to investigate whether there are certain inflammatory diseases with elevated soluble TL1A. Undoubtedly, the successful completion of phase-I feasibility study will allow us to rapidly commercialize ELISA, and also to demonstrate the feasibili ty of ELISA to detect soluble TL1A in human clinical sera. As TL1A will likely mediate various pro-inflammatory and pro-apoptotic responses in the body, the commercial value of a well characterized ELISA is undoubtedly high. Moreover the discovery of eleva ted soluble TL1A in certain inflammatory diseases will provide us with the rationale for the development of TL1A as a biomarker of the diseases in the future phase study. A family of proteins in the body can cause inflammatory reactions that advers ely affect the quality of life. In particular, a protein called TNF is known to be responsible for pathogenesis of inflammatory diseases, and therapeutic drugs designed to inhibit TNF have been quite successful to alleviate inflammatory symptoms including rheumatoid arthritis and inflammatory bowel diseases. We have recently discovered another TNF-like protein named TL1A that appeared to be responsible for both aberrant cell death and inflammatory reactions in the body. TL1A is believed to be circulating as a soluble protein in the body but there have been no reports on the detection of TL1A levels in patients with inflammatory diseases. Therefore, we propose to develop a sensitive and specific immunological assay to quantify the level of soluble TL1A in hum an clinical sera samples. The positive outcome of our project is two-fold. First, we will rapidly commercialize the assay so the assay can be used by other researchers to investigate the role of TL1A in a variety of inflammatory conditions. Second, we will find out whether there is indeed elevated TL1A expression in certain inflammatory diseases. These results will aid us to further investigate the role of TL
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