Dynamic mechanisms of FGFR activation in cancer by kinase mutations

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The way in which cells divide, proliferate and, in turn, die and become 'recycled' must be very carefully regulated in a highly programmed manner. Both development and maturation, as well as normal functioning of the adult organism, need to follow well-defined paths, and responses to environmental influences such as temperature, availability of food etc. must occur in a predictable manner. These responses require very fine control of complex cellular processes at the level of individual molecules. When this fine control breaks down, diseases such as cancer, degenerative disorders (e.g. Alzheimer's disease) and inflammatory conditions can result. Understanding these cellular and molecular processes in detail is important both to understand normal growth and development, and to provide us with insights into how serious diseases can be treated.Fibroblast growth factors (FGFs) are protein 'hormones' produced by certain cells to stimulate the growth of other cells involved in important processes such as the development of an embryo, the growth of new blood vessels and the repair and healing of wounds. FGF molecules bind to the outer parts of FGF receptors (FGFRs), which are proteins that span across the cell's protective outer membrane, and cause FGFR molecules to pair up. The parts of the receptor proteins that are inside the cell, known as kinase domains, are then close enough to activate one another through addition of phosphate 'chemical labels' that induce a change in the shape of the kinase domains from an inactive to an active conformation, causing the kinase domains to activate other proteins in the cell in a 'signalling cascade' that tells the cell to start dividing and proliferating. In turn, this process results in the formation of new tissues. The role of FGFs and FGFRs in formation of new blood vessels is also significant in cancer, where tumour cells often artificially elevate FGFR signalling within and between themselves as a way of securing a supply of nutrients and oxygen for further growth. Starving cancers of their new blood supply by inhibiting FGFR signalling is a promising avenue for treatment, and drug companies are currently developing new medicines that inhibit the activity of FGFRs.Although we understand some of the mechanisms by which the kinase domain of FGFR is activated from static 'snapshots' of the protein by X-ray crystallography, we still lack knowledge of how the flexibility of the kinase protein contributes to this role. Most proteins are not rigid, but need to flex to change their shape, or parts of their shape, in subtle ways to allow them to perform their functions in the cell. We will use an innovative combination of experimental methods including nuclear magnetic resonance spectroscopy (NMR), surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC), together with advanced computational methods, to understand the role of flexibility of the protein in the transition between inactive and active conformations. NMR is a particularly powerful method for investigating flexibility in protein function at the level of individual atoms or groups of atoms, and here we will combine experimental information from NMR with cutting-edge computational modelling of kinase motion to describe these movements in much more detail than has been previously achieved.By understanding the protein motions that govern FGFR kinase activity, we can understand better how FGFRs function in normal tissues and how they can malfunction in certain diseases such as cancers and developmental disorders. For example, mutated forms of FGFRs are found in many cancers. These contain amino acid changes that short-circuit the normal activation process and result in a kinase that is permanently switched 'on'. Our work will lead to enhanced understanding of how to design drugs that specifically inhibit these mutant forms of FGFR, leading ultimately to better treatments for cancers and developmental disorders.