Biophysical and biological properties of quadruplex oligodeoxyribonucleotides

Single-stranded guanosine-rich oligodeoxyribonucle-otides (GROs) have a propensity to form quadru-plex structures that are stabilized by G-quartets. In addition to intense speculation about the role of G-quartet formation in vivo, there is considerable interest in the therapeutic potential of quadruplex oligonucleotides as aptamers or non-antisense anti-proliferative agents. We previously have described several GROs that inhibit proliferation and induce apoptosis in cancer cell lines. The activity of these GROs was related to their ability to bind to a speci®c cellular protein (GRO-binding protein, which has been tentatively identi®ed as nucleolin). In this report, we describe the physical properties and biological activity of a group of 12 quadruplex oligonucleotides whose structures have been characterized previously. This group includes the thrombin-binding aptamer, an anti-HIV oligonucle-otide, and several quadruplexes derived from telo-mere sequences. Thermal denaturation and circular dichroism (CD) spectropolarimetry were utilized to investigate the stability, reversibility and ion dependence of G-quartet formation. The ability of each oligonucleotide to inhibit the proliferation of cancer cells and to compete for binding to the GRO-binding protein was also examined. Our results con®rm that G-quartet formation is essential for bio-logical activity of GROs and show that, in some cases, quadruplex structures formed in the pres-ence of potassium ions are signi®cantly more active than those formed in the presence of sodium ions. However, not all quadruplex structures exhibit anti-proliferative effects, and the most accurate factor in predicting biological activity was the ability to bind to the GRO-binding protein. Our data also indicate that the CD spectra of quadruplex oligonucleotides may be more complex than previously thought