Epigenetic regulation enables cells to modulate gene expression, with the modification of the histone N-termini representing a central mechanism of regulating access to associated DNA coding regions. One of the responsible enzyme families is the arginine methyltransferases, which dimethylate key arginines of histone and non-histone proteins.
Increased levels of protein arginine methyltransferase 5 (PRMT5) are found in cancers as diverse as melanoma, mantle cell lymphoma and oesophageal cancer and have been connected to hyperproliferation and increased translation. PRMT5 methylates the tumour suppressor p53, which determines a cell’s fate after DNA damage, and can modulate the p53 response. Under hypoxic conditions, PRMT5 is involved in activation of hypoxia induced factor (HIF-1α) in cancerous cells, providing the cells time to adapt and survive. However, currently there are no specific inhibitors of PRMT5 available for therapy.
To identify small molecule inhibitors of PRMT5 we utilised a robust non-radiometric assay of peptide substrate methylation based on Transcreener EPIGENTM technology. We screened a diverse library of 350,000 small lead-like molecules and known pharmacologically active compounds in 1536-well microplates. Hits were re-synthesised and confirmed by surface plasmon resonance, radiometric methyltransferase activity, and microscale thermophoresis. We identified several chemical scaffolds of low micromolar activity, which proved to be highly selective when tested in a panel of 25 protein and DNA methyl transferases and 12 kinases, inhibition of MLL4 being the only significant off-target activity of one scaffold.
Our modifications of the original screening hits by medicinal chemistry have produced compounds with 100-fold greater inhibition of PRMT5 methylation and improved physicochemical properties. Preliminary results indicate that these compounds display on-target activity in cell lines and reduce proliferation. These results confirm the potential of PRMT5 inhibitors as a new class of therapy for treating several forms of cancer.