Alan Clarke - Overview

The principal interest of my laboratory lies in understanding the pathogenesis of disease, with specific focus on genes involved in the very early stages of divergence from normality. The genes that control these key early events are often regarded as gatekeepers to the disease process and represent new potential targets for therapeutic intervention. Although we are investigating a number of different molecules, including those involved in signal transduction and those implicated in the prion diseases, most effort is placed on understanding the genetic control of the very early stages of cancer. The approach we have adopted relies heavily upon the use of transgenic models, as we believe that, despite the many advantages and uses of in vitro experimentation, gene function can only be truly understood in an in vivo context. Although we make great use of existing models, we are also aware of considerable difficulties in their use and a parallel goal has therefore been to enhance and refine these systems. The classical transgenic route has been to simply add sequences and characterise any alteration in phenotype. However, although this approach has proven very powerful it is also relatively clumsy, with results often confounded due to a lack of control over the site of transgene integration. This route also does not allow genetic material to be removed, so blocking the generation of models of loss of gene function. These difficulties were largely overcome through the use of gene targeting to specifically alter a chosen sequence and permit the generation of knockout strains. More recently, we have used Cre-lox technology to render targeted genes conditional (switchable). Some years ago we used knockout technology to generate a model null for the p53 gene, at the time considered to be the ultimate gatekeeper for the prevention of neoplasia. We used this strain to characterise an in vivo role for p53 in eliciting cellular suicide. Thus, in the presence of functional p53, cells within a variety of tissue types commit suicide following exposure to potential mutagens. In the absence of p53 this mechanism fails to operate and cells persist, presumably carrying a level of DNA damage. It is these cells which have been hypothesised to be the founders of cancer. P53 is not the sole focus of such cancer studies, and we have successfully identified a number of other genes which play a similar role in clearing DNA damage by invoking cell suicide. These include the mismatch repair genes Msh2 and Pms2 and the Methyl Binding Domain protein Mbd4. We are currently engaged in a series of studies with models bearing mutations in all of these genes. In pursuing this central hypothesis, we have already revealed a remarkable complexity in the reliance of different cell types upon defence mechanisms such as the ability to commit suicide. We believe that by unravelling these dependencies we will ultimately understand how and why a cell enters the pathway to become cancerous and that this knowledge will be key to the generation of radically new therapies.