Ruthenium complex binding to DNA G-quadruplexes

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

Photoactive ruthenium polypyridyl complexes are versatile and tunable for a range of applications in the life sciences because of their interactions with nucleic acids. The photoactivity can be harnessed as a diagnostic 'light switch' or towards DNA damage. In Reading we have become world leaders in understanding these interactions using a combination of a range of spectroscopic techniques combined with X-ray crystallography. Recently we examined the specific question of synthesising new complexes with high G-quadruplex/duplex preference, making use of our existing structural information. This is a very active field since the demonstration that G-quadruplexes can be recognised by specific antibodies in living cells, and furthermore, that G-quadruplex forming sequences play a key role in promoter regions of the human genome. The interaction of G-quadruplexes and our ruthenium complexes is determined by the rigidity and inertness to substitution of the complexes, and as we have recently shown, by the substituents on the dppz ligand. We have defined a structural basis for the clear lambda enantiomer preference and now seek to define the role of the extended dppz chromophore in the G-quadruplex interaction. We will carry out a range of synthetic studies to optimise further our ligand design, aiming to optimise both the binding constant and the luminescence properties. The photooxidising TAP complexes are useful tools, since quenching of the luminescence indicates binding adjacent to a G-quadruplex, whereas binding to loop regions containing only adenine and thymine residues gives enhancement of luminescence. We already have some X-ray data showing enantiomeric binding to loops in the human telomeric quadruplex and aim to develop full high resolution structural models. With our partners we will study the inhibition of G-quadruplex replication by these complexes and the effect on the binding of the BG4 antibody in a range of cell lines.

The fundamental nature of the proposed research has clear benefits to the academic community, with insights into the binding of enantiomers of ruthenium complexes to G-quadruplexes. Behind this academic impact is a range of scientific techniques that require a high level of skill and coordination from the interdisciplinary team to optimise the techniques and to develop a coherent interpretation of the results. The PDRA and technician will enhance their skills within a variety of techniques, and be able to use these skills in subsequent employment. Several members of the Cardin group who have developed key skills in crystallography as students and PDRAs within the group have gone on to obtain positions at central facilities such as Diamond Light Source, ILL and NPL, as well as industries such as GSK and Evotek. Several of the technicians have gone on to BSc or MSc level studies in chemistry, environmental chemistry and pharmacy, or to more senior analytical roles, and one is in permanent employment in Diamond Light Source as a Laboratory Instrument Technician.
The detection and specific targeting of nucleic acid assemblies such as the G-quadruplex is relevant to the design of new fluorophores and therapeutically active species. Ruthenium complexes have been considered promising candidates for photodynamic therapy for several years. The proposed research will enhance our understanding of the complex nature behind DNA binding, especially of the different enantiomers and extended chromophores. If pharmaceutical companies are to develop these metal complexes into drugs for treatment of disease, then a full understanding of selective or specific binding is essential, along with knowledge of how derivatisation can tune the binding properties.

The awareness of the public about DNA structure is often limited to that found in biology textbooks, most commonly the double helical nature of DNA. Some may be aware of the damage that chemicals can do to human health, but the underlying mechanisms and structural changes to DNA that occur are not as well known. The team in Reading have an expertise in understanding and visualising the structure of DNA and putting these structures into a biological context. Using our expertise to show how DNA can adopt multiple structures, and that these structures can be damaged to different degrees and in different ways, will help to educate school-age students and the scientifically interested lay public to the importance of understanding how damage can occur and why it is important to protect our DNA from this damage, for example by wearing suntan lotion, or refraining from smoking. As a part of this project, an extensive programme of outreach activities will be undertaken (as detailed in the Pathways to Impact), the aim of which is to educate students and those interested in science about our work and how nucleic acids are relevant in everyday life.