4th Generation aromatase inhibitors for the treatment of breast cancer: design, synthesis, biochemical and molecular biology studies

Breast cancer is one of the most common forms of cancer worldwide with 11.6% of all cancer incidence in 2018. One in every eight women will be diagnosed with breast cancer during their lifetime and approximately 70% of all patients are oestrogen receptor (ER) positive depending upon oestrogen for their growth. Oestrogens are synthesised from androgens through three steps, the last of which is catalysed by aromatase enzyme (CYP19A1), accounting for third generation aromatase inhibitors being the mainstay in the treatment of ER-positive breast cancer. Despite the success of current aromatase inhibitors, acquired resistance occurs after prolonged therapy. Although the precise mechanisms of resistance are not known, lack of cross resistance among aromatase inhibitors drives the need for a newer generation of inhibitors to overcome this resistance alongside minimising toxicity and adverse effects. Novel inhibitors including 22 triazole-based compounds and 12 pyridine-based compounds were designed based on previously published parent compounds (6a, 6d, 36a and 36b) by our group, making use of the now available crystal structure of CYP19A1 (PDB 3S79), to make modifications at specific sites to explore the potential of dual binding of both the active site and the access channel. Modifications included adding long chain substituents e.g. but-2-ynyloxy and pent-2-ynyloxy at different positions. The designed compounds were synthesised through various synthetic pathways and were fully characterised to ensure the effectiveness of the methods and quality of the products including the most active compound 23c with IC50 value in the picomolar range (0.09 nM). Aromatase inhibition results paired with the molecular dynamics studies provided a clear structure activity relationship and favourable dual binding mode was verified. Also, 11 sulfamate-based compounds were designed by the incorporation of the sulfamate group into the aromatase inhibitors to explore the aromatase/sulfatase dual inhibition. Six of these compounds (47-51) were based on the triazole scaffold, however biological evaluation revealed no sulfatase inhibitory activity despite the potent aromatase inhibition. Modifications led to the synthesis of the other five compounds (56) to achieve a balanced aromatase/sulfatase inhibition. However, the synthetic scheme for these compounds was not optimal either owing to poor reactivity of starting material and/or questionable purity of the products. This requires more investigation by working with larger scale and/or modifications to the structural design to overcome the low yield produced and the extensive purification process. Proliferation assays and CYP selectivity profile studies for the most active compounds will be performed to select the best candidate for further development and investigations