Design, synthesis and biological evaluation of novel 2,4-thiazolidinedione derivatives able to target the human BAG3 protein

: The Bcl-2-associated athanogene 3 (BAG3) protein plays multiple roles in controlling cellular homeostasis, and it has been reported to be deregulated in many cancers, leading tumor cell apoptosis escape. BAG3 protein is then an emerging target for its oncogenic activities in both leukemia and solid cancers, such as medulloblastoma. In this work a series of forty-four compounds were designed and successfully synthesized by the modification and optimization of a previously reported 2,4-thiazolidinedione derivative 28. Using an efficient cloning and transfection in human embryonic kidney HEK-293T cells, BAG3 was collected and purified by chromatographic techniques such as IMAC and SEC, respectively. Subsequently, through Surface Plasmon Resonance (SPR) all the compounds were evaluated for their binding ability to BAG3, highlighting the compound FB49 as the one having the greatest affinity for the protein (Kd = 45 ± 6 μM) also against the reference compound 28. Further analysis carried out by Saturation Transfer Difference (STD) Nuclear Magnetic Resonance (NMR) spectroscopy further confirmed the highest affinity of FB49 for the protein. In vitro biological investigation showed that compound FB49 is endowed with an antiproliferative activity in the micromolar range in three human tumoral cell lines and more importantly is devoid of toxicity in human peripheral mononuclear cell deriving from healthy donors. Moreover, FB49 was able to block cell cycle in G1 phase and to induce apoptosis as well as autophagy in medulloblastoma HD-MB03 treated cells. In addition, FB49 demonstrated a synergistic effect when combined with a chemotherapy cocktail of Vincristine, Etoposide, Cisplatin, Cyclophosphamide (VECC). In conclusion we have demonstrated that FB49 is a new derivative able to bind human BAG3 with high affinity and could be used as BAG3 modulator in cancers correlated with overexpression of this protein.