Molecular Docking and Biophysical Studies for Antiproliferative Assessment of Synthetic Pyrazolopyrimidinones Tethered with Hydrazide-Hydrazones.

Chemotherapy represents the most applied approach to cancer treatment. Owing to the frequent onset of chemoresistance and tumor relapses, there is an urgent need to discover novel and more effective anticancer drugs. In the search for therapeutic alternatives to treat the cancer disease, a series of hybrid pyrazolo[3,4-d]pyrimidin-4(5H)-ones tethered with hydrazide-hydrazones, 5a–h, was synthesized from condensation reaction of pyrazolopyrimidinone-hydrazide 4 with a series of arylaldehydes in ethanol, in acid catalysis. In vitro assessment of antiproliferative effects against MCF-7 breast cancer cells, unveiled that 5a, 5e, 5g, and 5h were the most effective compounds of the series and exerted their cytotoxic activity through apoptosis induction and G0/G1 phase cell-cycle arrest. To explore their mechanism at a molecular level, 5a, 5e, 5g, and 5h were evaluated for their binding interactions with two well-known anticancer targets, namely the epidermal growth factor receptor (EGFR) and the G-quadruplex DNA structures. Molecular docking simulations highlighted high binding affinity of 5a, 5e, 5g, and 5h towards EGFR. Circular dichroism (CD) experiments suggested 5a as a stabilizer agent of the G-quadruplex from the Kirsten ras (KRAS) oncogene promoter. In the light of these findings, we propose the pyrazolo-pyrimidinone scaffold bearing a hydrazide-hydrazone moiety as a lead skeleton for designing novel anticancer compound

A potentiated cooperation of carbonic anhydrase IX and histone deacetylase inhibitors against cancer

The emergence of tumour recurrence and resistance limits the survival rate for most tumour-bearing patients. Only, combination therapies targeting pathways involved in the induction and in the maintenance of cancer growth and progression might potentially result in an enhanced therapeutic efficacy. Herein, we provided a prospective combination treatment that includes suberoylanilide hydroxamic acid (SAHA), a well-known inhibitor of histone deacetylases (HDACs), and SLC-0111, a novel inhibitor of carbonic anhydrase (CA) IX. We proved that HDAC inhibition with SAHA in combination with SLC-0111 affects cell viability and colony forming capability to greater extent than either treatment alone of breast, colorectal and melanoma cancer cells. At the molecular level, this therapeutic regimen resulted in a synergistically increase of histone H4 and p53 acetylation in all tested cell lines. Overall, our findings showed that SAHA and SLC-0111 can be regarded as very attractive combination providing a potential therapeutic strategy against different cancer models

Advances in Azorella glabra Wedd. Extract research: In vitro antioxidant activity, antiproliferative effects on acute myeloid leukemia cells and bioactive compound characterization

Azorella glabra Wedd. (AG) is traditionally used to treat gonorrhea or kidney's problems. The antioxidant, antidiabetic, anticholinesterase and in vitro antitumor activities of AG extracts were recently reported. The aim of this work was to investigate anti-leukemic properties of AG chloroform fraction (AG CHCl3) and of its ten sub-fractions (I-X) and to identify their possible bioactive compounds. We determined their in vitro antioxidant activity using 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), nitric oxide (NO) and superoxide anion (SO) assays, and their phytochemical profile by spectrophotometric and LC-MS/MS techniques. I-X action on two acute myeloid leukemia (AML) cell lines viability, apoptosis and cell cycle were evaluated by MTS, western blotting and cytofluorimetric assays. Different polyphenol, flavonoid and terpenoid amount, and antioxidant activity were found among all samples. Most of I-X induced a dose/time dependent reduction of cell viability higher than parent extract. IV and VI sub-fractions showed highest cytotoxic activity and, of note, a negligible reduction of healthy cell viability. They activated intrinsic apoptotic pathway, induced a G0/G1 block in leukemic cells and, interestingly, led to apoptosis in patient AML cells. These activities could be due to mulinic acid or azorellane terpenoids and their derivatives, tentatively identified in both IV and VI. In conclusion, our data suggest AG plant as a source of potential anti-AML agents

The new low-toxic histone deacetylase inhibitor S-(2) induces apoptosis in various acute myeloid leukemia cells

Histone deacetylase inhibitors (HDACi) induce tumour cell cycle arrest and/or apoptosis, and some of them are currently used in cancer therapy. Recently, we described a series of powerful HDACi characterized by a 1,4-benzodiazepine (BDZ) ring hybridized with a linear alkyl chain bearing a hydroxamate function as Zn(++)-chelating group. Here, we explored the anti-leukaemic properties of three novel hybrids, namely the chiral compounds (S)-2 and (R)-2, and their non-chiral analogue 4, which were first comparatively tested in promyelocytic NB4 cells. (S)-2 and partially 4– but not (R)-2 – caused G0/G1 cell-cycle arrest by up-regulating cyclin G2 and p21 expression and down-regulating cyclin D2 expression, and also apoptosis as assessed by cell morphology and cytofluorimetric assay, histone H2AX phosphorylation and PARP cleavage. Notably, these events were partly prevented by an anti-oxidant. Moreover, novel HDACi prompted p53 and α-tubulin acetylation and, consistently, inhibited HDAC1 and 6 activity. The rank order of potency was (S)-2 > 4 > (R)-2, reflecting that of other biological assays and addressing (S)-2 as the most effective compound capable of triggering apoptosis in various acute myeloid leukaemia (AML) cell lines and blasts from patients with different AML subtypes. Importantly, (S)-2 was safe in mice (up to 150 mg/kg/week) as determined by liver, spleen, kidney and bone marrow histopathology; and displayed negligible affinity for peripheral/central BDZ-receptors. Overall, the BDZ-hydroxamate (S)-2 showed to be a low-toxic HDACi with powerful anti-proliferative and pro-apototic activities towards different cultured and primary AML cells, and therefore of clinical interest to support conventional anti-leukaemic therapy

Star-shaped Magnetic-plasmonic Au@Fe3O4 nano-heterostructures for photothermal therapy

Here, we synthesize a Au@Fe3O4 core@shell system with a highly uniform unprecedented star-like shell morphology with combined plasmonic and magnetic properties. An advanced electron microscopy characterization allows assessing the multifaceted nature of the Au core and its role in the growth of the peculiar epitaxial star-like shell with excellent crystallinity and homogeneity. Magnetometry and magneto-optical spectroscopy revealed a pure magnetite shell, with a superior saturation magnetization compared to similar Au@Fe3O4 heterostructures reported in the literature, which is ascribed to the star-like morphology, as well as to the large thickness of the shell. Of note, Au@Fe3O4 nanostar-loaded cancer cells displayed magneto-mechanical stress under a low frequency external alternating magnetic field (few tens of Hz). On the other hand, such a uniform, homogeneous, and thick magnetite shell enables the shift of the plasmonic resonance of the Au core to 640 nm, which is the largest red shift achievable in Au@Fe3O4 homogeneous core@shell systems, prompting application in photothermal therapy and optical imaging in the first biologically transparent window. Preliminary experiments performing irradiation of a stable water suspension of the nanostar and Au@Fe3O4-loaded cancer cell culture suspension at 658 nm confirmed their optical response and their suitability for photothermal therapy. The outstanding features of the prepared system can be thus potentially exploited as a multifunctional platform for magnetic-plasmonic applications

Two novel precursors of the hiv-1 protease inhibitor darunavir target the UPR/proteasome system in human hepatocellular carcinoma cell line HepG2

Background: Several pre-clinical and clinical reports suggest that HIV-1 protease inhibitors, in addition to the antiretroviral properties, possess pleiotropic pharmacological effects including anti-cancer action. Therefore, we investigated the pro-apoptotic activity in tumor cells of two molecules, RDD-19 and RDD-142, which are hydroxyethylamine derivatives’ precursors of darunavir and several HIV-1 protease inhibitors. Methods: Three hepatoma cell lines and one non-pathological cell line were treated with RDD-19 and RDD-142, and cell viability was assessed. The expression levels of several markers for ER stress, autophagy, cellular ubiquitination, and Akt activation were quantified in HepG2 cells treated with RDD-19 and RDD-142 to evaluate apoptotic and non-apoptotic cell death. Results: RDD-19 and RDD-142 showed a greater dose-dependent cytotoxicity towards the hepatic tumor cell line HepG2 compared to the non-pathological hepatic cell line IHH. Both molecules caused two types of cell death, a caspase-dependent apoptosis, which was ascertained by a series of biochemical and morphological assays, and a caspase-independent death that was characterized by the induction of ER stress and autophagy. The strong increase of ubiquitinated proteins inside the cells suggested that the target of these molecules could be the proteasome and in silico molecular docking analysis that was used to support the plausibility of this hypothesis. Furthermore, cells treated with the two compounds displayed decreased levels of p-AKT, which interferes with cell survival and proliferation. Conclusions: These findings demonstrate that two compounds, RDD-19 and RDD-142, have pleiotropic effects and that they may represent promising anticancer candidates

u-PAR expression in cancer associated fibroblast: new acquisitions in multiple myeloma progression

BACKGROUND: Multiple Myeloma (MM) is a B-cell malignancy in which clonal plasma cells progressively expand within the bone marrow (BM) as effect of complex interactions with extracellular matrix and a number of microenvironmental cells. Among these, cancer-associated fibroblasts (CAF) mediate crucial reciprocal signals with MM cells and are associated to aggressive disease and poor prognosis. A large body of evidence emphasizes the role of the urokinase plasminogen activator (u-PA) and its receptor u-PAR in potentiating the invasion capacity of tumor plasma cells, but little is known about their role in the biology of MM CAF. In this study, we investigated the u-PA/u-PAR axis in MM-associated fibroblasts and explore additional mechanisms of tumor/stroma interplay in MM progression. METHODS: CAF were purified from total BM stromal fraction of 64 patients including monoclonal gammopathy of undetermined significance, asymptomatic and symptomatic MM, as well as MM in post-treatment remission. Flow cytometry, Real Time PCR and immunofluorescence were performed to investigate the u-PA/u-PAR system in relation to the level of activation of CAF at different stages of the disease. Moreover, proliferation and invasion assays coupled with silencing experiments were used to prove, at functional level, the function of u-PAR in CAF. RESULTS: We found higher activation level, along with increased expression of pro-invasive molecules, including u-PA, u-PAR and metalloproteinases, in CAF from patients with symptomatic MM compared to the others stages of the disease. Consistently, CAF from active MM as well as U266 cell line under the influence of medium conditioned by active MM CAF, display higher proliferative rate and invasion potential, which were significantly restrained by u-PAR gene expression inhibition. CONCLUSIONS: Our data suggest that the stimulation of u-PA/u-PAR system contributes to the activated phenotype and function of CAF during MM progression, providing a biological rationale for future targeted therapies against MM