Characterization of novel HuR inhibitors and their effect on macrophages and cancer cells

Inflammation represents a pivotal, fine-tuned response in mammalian organisms. Not surprisingly, an uncontrolled and misregulated inflammation is major cause for important diseases, such as autoimmune disorders or cancer. Regulation of the inflammatory response occurs mainly through cytokines, which are strongly controlled at post-transcriptional levels by RNA binding proteins (RBPs). Among these, the Human Antigen R (HuR or ELAVL1), a member of the ELAVL family, embodies the multifaceted role of RBPs, participating in diverse processes including organismal development, cell growth, inflammation, and others, regulating roughly 7% of the full transcriptome. Consequently, small alterations in HuR expression or function can lead to severe consequences and numerous associated diseases. Importantly, HuR has been identified as a main regulator of the innate immune response, therefore its inhibition can have beneficial anti-inflammatory potential. The research for HuR inhibitors represents a challenging area in the drug discovery field, due to its pleiotropic functions and its structural complexity. Although the extensive efforts spent for this goal, no HuR inhibitor has succeeded so far in clinic. In 2015 our group identified through a High-throughput Screening a natural compound, DHTS as a novel HuR inhibitor. With a functional oriented approach, based on the chemical structure of DHTS-I, we synthesized a novel class of small molecules called Tanshinone Mimics (TMs), with a new molecular structure not previously described, aiming at increasing potency, specificity and solubility. In this thesis, I extensively characterized some of these molecules biochemically, in cellular models and in vivo. TMs are showing promising inhibitory activity against HuR modulating the immune response in murine and human macrophages. In human macrophages, TMs seem to activate caspase 3/7 apoptotic response, as well as to alter M1/M2 differentiation markers expression. Interestingly, TMs also affected some cancer traits in triple negative breast cancer cells. Studies on drug cell entrance suggest the TMs to have a quick metabolism within the cells. Characterization of TMs activity in vivo, appears in line with drug cell entrance studies and suggest the need for a frequent administration. However, TMs HuR specificity is in doubt due to some general toxicity observed in HuR negative cells. Finally, in a parallel approach, with a DEL-Open screen we have identified another promising HuR inhibitor, named WUXI7, which appears to be more stable over time inside cells. In a similar perspective to TMs, WUXI derivates have been synthesized and are currently under characterization.