Abstract 992
Background
S100A4, is a small EF-hand calcium-binding protein originally identified in myoepithelial/mesenchymal-like cell lines of the breast (Barraclough et al, 1982, 1987). S100A4, like many S100 proteins (Gross et al, 2014) induces cell migration and invasion (Jenkinson et al, 2014) which are the principal steps in the formation of metastases. Although there are several ways to target S100 proteins (e.g. Dahlmann et al, 2012), only small chemicals can enter cancer cells relatively easily. Based on a screen of compounds from CRUK, we have synthesised a few derivative compounds that can inhibit the binding of S100A4 to non-muscle myosin IIA (Ismail et al, 2008). Our lead compound, US10113, showed a similar Kd to that of pentamidine at about 10-6M, and at 33µM it inhibited specifically S100A4-induced cell migration in cell lines (Preliminary Data (PD) 1B) and invasion/metastasis by 84% in our Drosophila model (Ismail et al, 2017) with little apparent toxicity (PD Figure 2A, C).
Methods
To improve the efficiency of our inhibitor US10113 we synthesised it de novo coupled to thalidomide. The target of thalidomide is cereblon (CRBN), a component of a cullin-RING ubiquitin ligase (CRL) complex (Ito et al, 2010), which promotes CRBN-dependent proteasomal degradation (Lu et al, 2014). This approach, termed PROTAC, is used to convert a chemical inhibitor into a reagent that selectively destroys its target protein (Winter et al, 2015). In our hands the coupled chemical inhibitor, RGC, is soluble in water and shows little toxic effects on cells. Its inhibitory effects on cell migration and invasion, and metastasis in our drosophila model have been evaluated.
Results
The RGC could inhibit cell migration and invasion in nM concentrations compared to the 10’s µM range for US10113, i.e. 10,000 improvement in efficiency with complete specificity. This is due to the efficient degradation of S100A4 by cells. Using our drosophila metastasis model, we have demonstrated that the RGC is much more efficient in reducing S100A4-enhanced cancer cell spreading than US10113 that only inhibits S100A4 function but not degrades S100A4 protein.
Conclusions
PROTAC is a novel approach to degrade metastasis-promoting proteins and its efficacy is much higher than ordinary inhibitors.
Editorial acknowledgement
Clinical trial identification
Legal entity responsible for the study
Professor Philip Rudland, Institute of Integrative Biology, University of Liverpool.
Funding
Cancer and Polio Research Fund.
Disclosure
All authors have declared no conflicts of interest.