A chemical genetic approach to identify new treatments for melanoma
Melanoma is the most deadly form of skin cancer which develops from the pigment producing cells called melanocytes. Unlike the majority of other cancers, the incidence rates of melanoma are still on the rise and the few treatment options currently available are being hindered by resistance. A recent chemical genetic screen carried out in Xenopus laevis embryos identified that the already FDA approved drug leflunomide used for rheumatoid arthritis patients also holds potential therapeutic value in treating melanoma. This thesis shows the results of a new successful chemical genetic and cell based viability screen of the NCI Diversity set II library. 13 potential novel targets for treating melanoma were identified. This thesis also further characterised the function of leflunomide and showed that leflunomide reduces the number of viable cells in both wild type and BRAFV600E mutant melanoma cell lines. Further experiments revealed leflunomide reduces cell proliferation and causes cells to arrest in G1 of the cell cycle. Cell death assays showed leflunomide to cause apoptosis coupled with a stable mitochondrial membrane potential at 25 and 50μM leflunomide. However at 100μM the number of apoptotic cells decreased and an increase in the number of viable cells with a hyperpolarised mitochondrial membrane potential was observed. To determine if leflunomide had the potential to be used in combination with other melanoma drugs, it was tested in combination with the MEK inhibitor selumetinib. This combination showed a synergistic effect in the majority of the cell lines tested. The M375 melanoma cell line produced strong synergy values across all of the combinations of leflunomide and selumetinib tested. This combination led to an enhanced decrease in tumour size when tested in vivo in a mouse xenograft model when compared to either drug alone. Key words; Melanoma, BRAFV600E, leflunomide, cell viability, selumetinib, synergy, combination index value, NCI Diversity Set II, Xenopus laevis.
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