Abstract 3681: Helping Eve overcome ADAM: Characterization of the G-quadruplex in the ADAM15 core promoter

Abstract

Abstract Breast cancer is the most common form of cancer in women, whose lethality is only surpassed by lung cancer. Through the last quarter-century there have been significant advances in detection and treatment options, such as Herceptin for HER2/neu positive cancers. While the development of targeted biological agents has greatly improved patient outcome, up to 70% of patients remain de novo refractory, relapse on therapy, or develop resistance. There is a pressing and eminent need for novel therapies to synergize with, or resensitize to, standard treatments. Enter ADAM15, with known zymogen, secretase, and disintegrin activies. This catalytically active member of the ADAM family is normally expressed in early embryonic development and is aberrantly expressed in various cancers, including breast. ADAM15 promotes extracellular shedding of E-cadherin, a soluble ligand for the HER2/neu receptor, leading to activation, increased motility, and proliferation. Seven independent microarray studies have shown that ADAM15 and HER2/neu are simultaneously upregulated in several stages of breast cancer, where their overexpression correlates with more aggressive and invasive disease. We have examined a unique string of GC-rich DNA within the critical core promoter of ADAM15. This region of DNA consists of seven tandem runs of three or more consecutive guanines. Under superhelical stress produced by the transcriptional complex, this region can relax from duplex DNA to form an intrastrand secondary structure known as a G-quadruplex. These globular entities generally serve as silencing elements for gene transcription. Furthermore, the topology of each G-quadruplex is as unique as a fingerprint. This allows for specific therapeutic targeting, offering a unique opportunity for treatment and selectivity over normal cells thus providing a potentially wide therapeutic window. Circular dichroism (CD) studies have confirmed the formation of a mixed parallel/anti-parallel G-quadruplex. Further CD analysis of the minimal G-quadruplex forming fragments has shown the capability of forming four unique thermally stable species. Electrophoretic mobility shift assay indicates the strongest intramolecular G-quadruplex formation in the 5′- and 3′-end runs of guanine, with minor intermolecular formations in the 3′-mid sequence, and a relatively unstable 5′-mid formation. To specifically determine which guanines are involved in the formation of the G-quadruplex and predict biologically relevant three-dimensional structures, DMS footprinting was performed on the full-length sequence and each of the four dissected regions. The pursuant data was used to build a molecular model of the structure. Full characterization of the G-quadruplex species formed will allow for specific drug targeting and stabilization, and the further development of novel, targeted therapeutics. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3681.