Title

Mechanism of action of the cell migration inhibitor quinocarmycin analog DX-52-1

Date of Completion

January 2008

Keywords

Chemistry, Biochemistry|Chemistry, Organic|Chemistry, Pharmaceutical

Degree

Ph.D.

Abstract

Cell migration is central to a number of normal and disease processes, including embryonic development, tissue repair, immune function, angiogenesis and cancer cell invasion and metastasis. Small organic molecules that inhibit cell migration have potential as both research probes and therapeutic agents. Utilizing a high-throughput cell migration assay for the identification of small-molecule inhibitors or accelerators of cell motility, we screened over 10,000 small molecules and identified 14 compounds that have reproducible antimigratory activity down to nanomolar or low micomolar concentrations. One of these compounds, quinocarmycin analog DX-52-1, a semisynthetic derivative of the natural product quinocarmycin (also known as quinocarcin), inhibits the migration of Madin-Darby canine kidney epithelial cells with nanomolar potency. While it had been assumed previously that the main target of DX-521 is DNA, we identified two protein targets of DX-52-1 and confirmed their relevance to cell migration. The primary target of DX-52-1 is the membrane-cytoskeleton linker protein radixin. A secondary target of DX-52-1 with functions in cell motility is galectin3, multifunctional protein whose best-known function is to bind carbohydrates. In addition, we have identified another tetrahydroisoquinoline with antimigratory activity that has far greater selectivity for galectin-3 over radixin, exhibiting little binding to radixin, both in vitro and in cells. Neither small molecule targets the carbohydrate-binding site of galectin-3, and our results imply that functions of galectin-3 independent of carbohydrate binding are required for cell migration. Exploring the signal transduction pathway that regulates radixin, we found that radixin is a substrate for G protein-coupled receptor kinase 2 (GRK2). Phosphorylation of a C-terminal threonine residue of radixin and related proteins by GRK2 results in localization of these proteins to the plasma membrane and formation of membrane protrusions required for cell migration. This work sheds new light on the functions of radixin and galectin-3 in epithelial cell migration. ^