Support is requested to develop and validate a high-throughput screening assay to identify small molecule inhibitors of the integrin a4-paxillin interaction. Inhibition of a4 integrin function by antibodies, such as Tysabri (Natalizumab) is of proven therapeutic value in diseases such as Multiple Sclerosis and may be beneficial in other autoimmune and inflammatory diseases. Nevertheless, mechanism-based toxicities involving developmental abnormalities of the heart and placenta and of hematopoiesis may accompany complete blockade of a4 integrin function. The applicants' studies using biochemistry, cell biology, and genetically-modified mice show that blocking a4 integrin signaling by inhibiting the binding of paxillin to the a4 cytoplasmic domain can inhibit inflammation, yet avert some mechanism- based toxicity. Furthermore, a low throughput screen has identified small molecule inhibitors of the interaction, thus establishing the feasibility of this approach. He proposes to adapt the existing ELISA to a high throughput mode by modifying the a4 integrin ligand for in vivo biotinylation and purifying paxillin's two paralogues, HIC-5 and leupaxin, for comparative studies. The assay will be adapted to chemiluminescence-based detection and a 384 well format, variation and stability will be established and a small pilot screen will be conducted. Secondary screens will eliminate false positives and toxic compounds and will position the applicant to screen up to 500,000 compounds provided by the Molecular Libraries Screening Centers Network (MLSCN) through collaboration with Dr. Cosford, director of the Center for Chemical Genomics at the Burnham Institute. These studies may lead to new treatments of inflammatory diseases such as Multiple Sclerosis. Furthermore, integrins other than a4 are important therapeutic targets in diseases such as heart attack, stroke, and cancer; these studies will advance a novel approach to anti-integrin therapy, blockade of integrin signaling. Health Relevance: Inhibition of a4 integrin function is of proven therapeutic value, but mechanism-based toxicities may limit therapy. The applicant has shown that blocking a4 integrin-paxillin interaction can block inflammation with less toxicity. The applicant proposes to develop screens that may lead to small molecule inhibitors of the a4-paxillin interaction and thus serve as new treatments of inflammatory diseases such as Multiple Sclerosis ?
| Kummer, Christiane; Petrich, Brian G; Rose, David M et al. (2010) A small molecule that inhibits the interaction of paxillin and alpha 4 integrin inhibits accumulation of mononuclear leukocytes at a site of inflammation. J Biol Chem 285:9462-9 |
