LB3
A biochemical and chemical genetic analysis of Aurora and Polo-like kinase inhibitors
Paul Scutt2, Dominic Sloane1, Patrick Eyers1
1University of Sheffield, UK; 2University of Manchester, UK
The Aurora and Polo-like kinases are central components of mitotic signalling pathways and recent evidence suggests that substantial cross-talk exists between Aurora A and Plk1. In addition to their validation as novel anti-cancer agents, small molecule kinase inhibitors are increasingly important tools to help dissect clinically-relevant protein phosphorylation networks. However, one major problem associated with kinase inhibitors is their promiscuity towards off-target members of the kinome, which makes interpretation of data obtained from complex cellular systems extremely challenging. Additionally, the emergence of inhibitor-resistance in patients makes it clear that an understanding of resistance mechanisms is important for optimising drug design.
In this study, we have exploited structural knowledge of the binding modes of VX-680, an Aurora kinase inhibitor, and BI 2536, a Polo-like kinase inhibitor, to design and evaluate drug-resistant kinase mutants for Aurora A, Aurora B, Plk1 and Plk4. Using inducible stable human cell lines, we authenticate mitotic targets for both compounds, and validate Aurora B as a critical anti-proliferative target for VX-680. We establish that cells expressing VX-680 resistant Aurora A are still sensitive to the effects of the Aurora inhibitor MLN8054, proving that these unrelated compounds utilise distinct interaction mechanisms to block Aurora A activity in cells. In addition, this chemical genetic approach allows us to prove that Aurora A phosphorylation on Thr 288 is controlled by a Plk1-mediated pathway in human mitotic cells, suggesting that drug discovery approaches targetting Plk1 might have the added benefit of preventing the activation of Aurora A in human cancer cells.
