Synthesis and Biological Evaluation of Novel 2-Aroyl Benzofuran-Based Hydroxamic Acids as Antimicrotubule Agents.

Because of synergism between tubulin and HDAC inhibitors, we used the pharmacophore fusion strategy to generate potential tubulin-HDAC dual inhibitors. Drug design was based on the introduction of a N-hydroxyacrylamide or a N-hydroxypropiolamide at the 5-position of the 2-aroylbenzo[b]furan skeleton, to produce compounds 6a-i and 11a-h, respectively. Among the synthesized compounds, derivatives 6a, 6c, 6e, 6g, 11a, and 11c showed excellent antiproliferative activity, with IC50 values at single- or double-digit nanomolar levels, against the A549, HT-29, and MCF-7 cells resistant towards the control compound combretastatin A-4 (CA-4). Compounds 11a and 6g were also 10-fold more active than CA-4 against the Hela cell line. When comparing the inhibition of tubulin polymerization versus the HDAC6 inhibitory activity, we found that 6a-g, 6i, 11a, 11c, and 11e, although very potent as inhibitors of tubulin assembly, did not have significant inhibitory activity against HDAC6.

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.

Molecular and functional profiling of chemotolerant cells unveils nucleoside metabolism-dependent vulnerabilities in medulloblastoma.

Chemotherapy resistance is considered one of the main causes of tumor relapse, still challenging researchers for the identification of the molecular mechanisms sustaining its emergence. Here, we setup and characterized chemotherapy-resistant models of Medulloblastoma (MB), one of the most lethal pediatric brain tumors, to uncover targetable vulnerabilities associated to their resistant phenotype. Integration of proteomic, transcriptomic and kinomic data revealed a significant deregulation of several pathways in resistant MB cells, converging to cell metabolism, RNA/protein homeostasis, and immune response, eventually impacting on patient outcome. Moreover, resistant MB cell response to a large library of compounds through a high-throughput screening (HTS), highlighted nucleoside metabolism as a relevant vulnerability of chemotolerant cells, with peculiar antimetabolites demonstrating increased efficacy against them and even synergism with conventional chemotherapeutics. Our results suggest that drug-resistant cells significantly rewire multiple cellular processes, allowing their adaptation to a chemotoxic environment, nevertheless exposing alternative actionable susceptibilities for their specific targeting.

New bioisosteric sulphur-containing choline kinase inhibitors with a tracked mode of action.

Since the identification of human choline kinase as a protein target against cancer progression, many compounds have been designed to inhibit its function and reduce the biosynthesis of phosphatidylcholine. Herein, we propose a series of bioisosteric inhibitors that are based on the introduction of sulphur and feature improved activity and lipophilic/hydrophilic balance. The evaluation of the inhibitory and of the antiproliferative properties of the PL (dithioethane) and FP (disulphide) libraries led to the identification of PL 48, PL 55 and PL 69 as the most active compounds of the series. Docking analysis using FLAP suggests that for hits to leads, binding mostly involves an interaction with the Mg2+ cofactor, or its destabilization. The most active compounds of the two series are capable of inducing apoptosis following the mitochondrial pathway and to significantly reduce the expression of anti-apoptotic proteins such as the Mcl-1. The fluorescence properties of the compounds of the PL library allowed the tracking of their mode of action, while PAINS (Pan Assays Interference Structures) filtration databases suggest the lack of any unspecific biological response.