The viral main protease is one of the most attractive targets among all key enzymes involved in the life cycle of SARS-CoV-2. Considering its mechanism of action, both the catalytic and dimerization regions could represent crucial sites for modulating its activity. Dual-binding the SARS-CoV-2 main protease inhibitors could arrest the replication process of the virus by simultaneously preventing dimerization and proteolytic activity. To this aim, in the present work, we identified two series' of small molecules with a significant affinity for SARS-CoV-2 M-PRO, by a hybrid virtual screening protocol, combining ligand- and structure-based approaches with multivariate statistical analysis. The Biotarget Predictor Tool was used to filter a large in-house structural database and select a set of benzo[b]thiophene and benzo[b]furan derivatives. ADME properties were investigated, and induced fit docking studies were performed to confirm the DRUDIT prediction. Principal component analysis and docking protocol at the SARS-CoV-2 M-PRO dimerization site enable the identification of compounds 1b,c,i,l and 2i,l as promising drug molecules, showing favorable dual binding site affinity on SARS-CoV-2 M-PRO.

A series of biologically unexplored substituted 1,3,4-subtituted-pyrrolo[3,2-c]quinoline derivatives (PQs) was evaluated against a panel of about 60 tumor cells (NCI). Based on the preliminary antiproliferative data, the optimizations efforts permitted us to design and synthesize a new series of derivatives allowing us to individuate a promising hit (4g). The insertion of a 4-benzo[d] [1,3]dioxol-5-yl moiety on increased and extended the activity towards five panel tumor cell lines such as leukemia, CNS, melanoma, renal and breast cancer, reaching IG50 in the low ?M range. Replacement of this latter with a 4-(OH-di-Cl-Ph) group (4i) or introduction a Cl-propyl chain in position 1 (5), selectively addressed the activity against the entire leukemia sub-panel (CCRF-CEM, K-562, MOLT-4, RPMI-8226, SR). Preliminary biological assays on MCF-7 such as cell cycle, clonogenic assay, ROS content test alongside a comparison of viability between MCF-7 and non-tumorigenic MCF-10 were investigated. Among the main anticancer targets involved in breast cancer, HSP90 and ER receptors were selected for in silico studies. Docking analysis revealed a valuable affinity for HSP90 providing structural insights on the binding mode, and useful features for optimization.

A set of variously substituted 1-arylpyrazol-3-one derivatives, including the di-ortho-aryl substituted ones, was synthesized as new potential anticancer compounds. To fulfill this aim, herein a regiospecific synthesis was proposed utilizing a new revisited one pot procedure, starting from commercial anilines and easily accessible 2,5-dimethyl-furan-3-one. In the course of the sequential ordered steps, in some cases, a nitro group displacement by chlorine took place to a minor extent. The in vitro screening against the full panel of similar to 60 human cancer cell lines (NCI) showed a moderate, but promising selective antiproliferative activity against the UO31 renal tumor cell line, only in compounds with the introduction on the phenyl moiety of a -CF3 or two CI groups.

Herein we describe a combined experimental and in silico study of the interaction of a series of pyrazolo[1,2-a]benzo[1,2,3,4]tetrazin-3-one derivatives (PBTs) with parallel G-quadruplex (GQ) DNA aimed at correlating their previously reported anticancer activities and the stabilizing effects observed by us on c-myc oncogene promoter GQ structure. Circular dichroism (CD) melting experiments were performed to characterize the effect of the studied PBTs on the GQ thermal stability. CD measurements indicate that two out of the eight compounds under investigation induced a slight stabilizing effect (2-4 °C) on GQ depending on the nature and position of the substituents. Molecular docking results allowed us to verify the modes of interaction of the ligands with the GQ and estimate the binding affinities. The highest binding affinity was observed for ligands with the experimental melting temperatures (Tms). However, both stabilizing and destabilizing ligands showed similar scores, whilst Molecular Dynamics (MD) simulations, performed across a wide range of temperatures on the GQ in water solution, either unliganded or complexed with two model PBT ligands with the opposite effect on the Tms, consistently confirmed their stabilizing or destabilizing ability ascertained by CD. Clues about a relation between the reported anticancer activity of some PBTs and their ability to stabilize the GQ structure of c-myc emerged from our study. Furthermore, Molecular Dynamics simulations at high temperatures are herein proposed for the first time as a means to verify the stabilizing or destabilizing effect of ligands on the GQ, also disclosing predictive potential in GQ-targeting drug discovery.

As extension of previous results reached on the pyrazolo[1,2-a]benzo[1,2,3,4]tetrazin-3-one derivatives (PBTs) endowed with anticancer potential, we focused our research interest to new emerging and challenging biological targets involved in carcinogenic process. More precisely to gene promoter related to the oncoprotein c-myc as well as Tel22. C-mycis overexpressed in a large number of cancer and related binders are potential anticancer drugs because they target a promoter quadruplex present upstream of the proto-oncogene myc,down-regulating its expression. Indeed, one of the possible mechanisms underlying the antiproliferative activity of anticancer drugs is precisely based on the induction of quadruplex structure in c-myc, with consequent blockage of oncogene expression. Analogously, telomeric DNA binders inhibit the ability, found in most human cancer cells,to indefinitely replicate becoming immortal, as they prevent the hybridization of the telomerase RNA template onto the primer. Therefore, the discovery of new molecular entity able to tune the binding or the stabilization of the G4 quadruplex represents a current therapeutically approach for anticancer diseases [1]. A preliminary NCI screening on variously substituted pyrazolo[1,2-a]benzo[1,2,3,4]tetrazin-3-one derivatives (PBTs) against a panel of 60 tumor cell lines, evidenced a promising anti-proliferative activity in the ?M and sub-?M range. Depending on the position and nature of the substituents, several derivatives targeted selectively specific tumor cells or in other cases more than ones. These attractive results, for this tricycle scaffold, biologically unexplored so far, prompted us to move to new optimized structures followed by evaluation on apoptosis induction and cell cycle perturbation [2]. Once assessed their anticancer potential, on PBTs further deeper studies became necessary, in order to get insights on their mechanism of action. Thus, more specific biological targets, such as duplex DNA and topoisomerase(II) catalytic cycle, were approached, including in silico support [3]. Here, extending the research on other recently recognized anticancer targets, we planned to evaluate the effects of PBTs on c-myc and Tel22 by mean of circular dichroism (CD) and related melting experiments. In support, molecular dynamics (MD) and docking techniques permitted us to gain additional insights on the observed effects. The outcomes will be shown with more details at poster session. [1] [Stephen Neidle, Nature Reviews Chemistry1, 2017, 0041]. [2] F. Mingoia, C. Di Sano, F. Di Blasi, M. Fazzari, A. Martorana, A.M. Almerico, A. Lauria Eur. J. Med. Chem. 2013, 64, 345-356. [3] A. Lauria, F. Mingoia, A. Garcia-Arcaez, A. Martorana, L. Dalla Via, Chem. Biol. DrugDes. 2018, 91, 463-477

Aryl-pyrazolones are known in scientific literature for their wide range of biological activity [1]. The occurrence of the recent approval by the U.S. FDA (Food and Drug Administration, 2017) of an aryl-pyrazol-5-one substrate such as Edaravone (Fig.1), as a novel neuroprotective agent for the treatment of ALS (Amyotrophic Lateral Sclerosis) [2], attracted us to extend research investigations on the isomeric aryl-pyrazol-3-one motif. The use of commercially available substituted reagents coupled with the easy synthetic route, optimized by us during the years, allow to obtain a wide range of suitably substituted derivatives either for biological evaluation and for exploitation as synthons. The presence of the tautomeric equilibrium, in this class of compounds, promted us to clarify some regio-selective insights, useful for developing new pyrazolone containing scaffolds. Fig.1: EdaravoneTautomeric equilibrium of 1-aryl-pyrazol-3-one Thus, we initially investigated the reactivity of the 1-(2-nitroaryl)-pyrazol-3-one moiety through nucleophilic substitution reactions carried out with different selected electrophiles and under different experimental conditions. Once assessed the different behavior of the electrophiles (R-CH2-Br) against -NH/OH tautomers, we introduced a third nucleophilic competitor on the 1-aryl moiety (R1=NH2). For this purpose, R-PhCOCl reactants were selected as electrophiles. In this way, an overview of the regioselective reactivity was obtained along with a series of new substituted(2-benzamido-aryl)-pyrazol-3-one derivatives to be biologically explored. Leave one line blank [1]El Sayed M. T., et al.Bio Med Chem Let2018, 28, 952-957. [2]Jaiswal M. K.Med Res Rev2019,39, 733-748.

Recenti studi di reattività condotti su derivati 1-aril-pirazol-3-oni, sia come intermedi chiave che come molecole bioattive, hanno permesso di evidenziare alcuni aspetti di regioselettività con alcuni elettrofili, in funzione delle condizioni sperimentali utilizzate1. A partire da tali basi, gli studi sono stati estesi alla progettazione e sintesi di nuovi derivati benzamidici variamente sostituiti incorporanti la porzione 1-aril-pirazolonica (1), di cui recentemente la FDA (2017) ha approvato la commercializzazione dell' EDARAVONE (2), un analogo strutturale, come nuovo agente neuroprotettivo. Lo scopo di questa ricerca mira a verificare gli effetti derivanti da una componente potenzialmente neuroprotettiva modulata da una componente benzamidica variamente sostituita(2) per l'ottenimento di nuovi derivati (3) di cui investigarne l'attività biologica. Un preliminare screening di vitalità cellulare permetterà di mettere in evidenza i composti a carattere citotossico, la cui l'attività biologica verrà approfondita e supportata da studi in silico2,3. 1 Mingoia F., Panzeca G., Gentile C., Martorana A., Lauria A.; MedChemSicily2018, Palermo, 17-20 July 2018, pp159. 2 Lauria A., Gentile C., Mingoia F., Palumbo Piccionello A., Bartolotta R., Delisi R., Buscemi S., Martorana A., Chem. Biol. Drug Des. 2018, 91, 39. 3 Lauria A., Mingoia F., Garcia-Argaez A. N., Delisi R., Martorana A., Dalla Via L., Chem. Biol. Drug Des. 2018, 91, 463.

Presentazione orale sugli ultimi sviluppi di ricerca condotta su intermedi 1-arilamino-pirazolonici che in studi precedenti hanno permesso l'accesso a derivati di interesse antitumorale. Dal sistema triciclico "5-6-6", si è progettato il sistema "5-7-6", generando una espansione di anello nella porzione centrale dell'anello. Lo scopo è quello di valutare gli effetti di tale espansione sull'attività antiproliferativa. I preliminari risultati di screening antitumorale eseguiti da NCI (USA), non sono confortanti se riferiti allo scaffold non sostituito. Di contro, l'introduzione di una catena cloro-propilica sullazoto amidico conduce a nuovi derivati attivi e particolarmente selettive sulle linee leucmiche. Così, viene individuata una nuova classe di composti triciclici ad attività antileucemica degna di ulteriori approfondimenti.

A crucial point in Medicinal Chemistry is the discovery of promising lead compounds and their optimization, in order to obtain new molecules with a good bioavailability and a high affinity for focused biological target. In lead optimization matter, the medicinal chemists have at their disposal several approaches more or less intuitive, such as the synthesis of drug analogues, isomer and isosteres, the modification of ring systems or the rational pharmacophore identification by computer-assisted drug design. In this context, the drug repurposing is a fashionable strategy to assign new therapeutic uses to known drugs or to promising molecules halted during the clinical development process.(1) In this work we propose a series of new naphthodithiophene compounds of type 1 and 2, firstly targeted as G-quadruplex (G4) stabilizers, and now repositioned as PTP1B and BCL2 inhibitors. The naphthodithiophene prototypes resulted as output molecules of in silico virtual screening structure-based studies (molecular docking, induced fit docking, molecular dynamics) applied on an in house database, that allow to correlate, with a good affinity score, these planar heterocyclic compounds to the secondary DNA structure rich in guanine, G4. Even if the preliminary in vitro studies suggested a good antiproliferative activity of the synthetized naphthodithiophenes 1 and 2 against the HeLa human cancer cell line, the consequent binding tests with G4s isoforms were unsatisfactory. Herein, we illustrate the description of the synthetic pathway carried out for the isolation and characterization of the whole new series of naphthodithiophene compounds 1 and 2 and their biological re-tasking as potential PTP1B and BCL2 inhibitors, with the help of new and innovative chemoinformatic tools

Pyrazolone derivatives are five membered important heterocyclic ring having one or two additional keto (CO) groups. Recently, pyrazol-3-ones containing tricycles have been investigated for their anticancer effects (1) especially as DNA and topoisomerase II modulators (2). Due to the easy preparation and to their interesting antiproliferative activity, in relation to our promising outcomes, we focused our studies on the synthesis and the biological evaluation of selected substituted-1-Aryl-Pyrazol-3-ones as new promising lead compounds. The nature and position of various substituents on the 1-aryl moiety have a crucial influence on the modulation of the tautomeric equilibrium, typical of this class of compounds. Thus, we explored adequate reaction conditions to allow a convenient regioselective control for the introduction of electrophilic substituents on a specific position, kept approachable by the tautomeric equilibrium. In particular, preliminary series of reactions with selected electrophiles have been carried out on derivatives a-c and further on e-g in order to clarify the substitution site. Herein we present our preliminary results on the regioselective reaction with selected strategic electrophiles along with the antiproliferative activity (NCI) of derivatives e-g.

The search of new small molecules behaving as free radical scavengers has attracted growing interest in drug discovery efforts because this fieldrepresents one of the most appealing therapeutic strategiesfor neuroprotection. Identifying new effective and low toxic neurotherapeutics is a challenging way for preventing neurodegeneration and mitigating against neuronal damage. A variety of central nervous system (CNS) disorders, including neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), dementia, stroke, and others, could have new therapeutic perspectives with new effective neuroprotective agents. A first step used by usfor evidencing the radical scavenger activity of the small molecules, is a DCF based assay. Further, cytocompatibility test has been performedby using different cell lines. Here, we propose our preliminary results ona structure activity relationship (SAR)of a series of variously substituted 1-Aryl-pyrazol-3-one derivatives as potential neuroprotective agents. This class of small molecules is related toEdaravone, arecentlyapproved drugfor neural damageby FDA (2017). In this way, nature and effect of substituents are under evaluationin order to get insights on the activity profile, useful for future developments.The chemical synthetic route followed involves the use of commercially available substituted reagents,providing access to a wide range of suitably substituted pyrazol-3-one derivatives.Thus, this scaffold could serve as a valuable lead for detailed SAR studies.In addition, in silico approacheswill be also considered.

A new series of 3-benzoylamino-5-(1H-imidazol-4-yl)methylaminobenzo[b]furans were synthesized and screened as antitumor agents. As a general trend, tested compounds showed concentration-dependent antiproliferative activity against HeLa and MCF-7 cancer cell lines, exhibiting GI(50) values in the low micromolar range. In most cases, insertion of a methyl substituent on the imidazole moiety improved the antiproliferative activity. Therefore, methyl-imidazolyl-benzo[b]furans compounds were tested in cell cycle perturbation experiments, producing cell cycle arrest with proapoptotic effects. Their core similarity to known colchicine binding site binders led us to further study the structure features as antitubulin agents by in silico protocols.

Due to the scarce biological profile, the pyrazolo[1,2-a]benzo[1,2,3,4]tetrazine-3-one scaffold (PBT) has been recently explored as promising core for potential anticancer candidates. Several suitably decorated derivatives (PBTs) exhibited antiproliferative activity in the low-micromolar range associated with apoptosis induction and cell cycle arrest on S phase. Herein, we selected the most active derivatives and submitted them to further biological explorations to deepen the mechanism of action. At first, a DNA targeting is approached by means of flow Linear Dichroism experiments so as to evaluate how small planar molecules might interact with DNA, including the interference with the catalytic cycle of topoisomerase II and the influence on the cleavable complex stabilization (poisoning effect). In support of the experimental data, in silico studies have been achieved to better understand the chemical space of the interactions. Interestingly some meaningful structural features, useful for further developments, were found. The 8,9-di-Cl substituted derivative revealed as the most effective in the intercalative process, as well as on the inhibition of catalytic activity of topoisomerase II. Predicted ADME studies confirm that PBTs are promising as potential drug candidates.

This work aims to fill the gap in the present knowledge about the structure of pectin from Opuntia ficus-indica. The water-soluble pectin (WSP) fraction, extracted with the Microwave Assisted Extraction (MAE), was further deproteinated (dWSP) and analyzed through several biophysical and biochemical techniques. HPSEC, light scattering and FTIR data showed that dWSP is low methylated high molecular weight pectin. The biochemical structure of dWSP, after methanolysis, silylation, carboxyl reduction showed that dWSP belongs to rhamnogalacturonan I class. Then, dWSP was heat-modified (HM) to obtain small-molecular weight deproteinated fraction (HM-dWSP). Both species, dWSP and HM-dWSP, were tested in LAN5 and NIH 3T3 model cells to study their biological effect. Results indicated that both dWSP and HM-dWSP exerted cytotoxic activity affecting selectively LAN5 cancer cells, without any effect on NIH 3T3 normal cells.

From chemical organic synthesis expertises ...to potential anticancer candidates: Molecular Scaffolds currently under study to ISMN-Pa The pyrazolo[1,2-a]benzo[1,2,3,4]tetrazin-3-one derivatives PBTs have been previously  designed as new alkylating agent because of their peculiar chemical behavior in soft acid environment towards charged or neutral nucleophiles. The tricycle is related to azolo-fused tetrazines such as Temozolomide, currently employed in cancer diseases. A preliminary NCI (USA) screening performed against more than 50 types of human tumor cell lines showed antiproliferative activity reaching in some cases sub-micromolar values [1]. DESIGN: Recently, through the in silico VLAK protocol [2], a second series of new PBTS derivatives have been designed. Four derivatives have been selected, synthesized and evaluated on peculiar cellular events including proliferation, apoptosis induction and cell cycle perturbation [3]. Several of the new synthesized derivatives screened against apoptosis induction (HeLa) showed promising anticancer potential reaching in some cases activity in the micromolar range and arresting the cell cycle in the S phase. Useful candidates to develop as potential anticancer agents ! They act simultaneously on multiple biological pathways. Further investigations on DNA /topoisomerase complex approached by Flow Linear Dicroism techniques. Additional dichroic signals appear at higher wavelengths (300-360 nm and 400-520 nm). The occurrence of a dichroic signal at these wavelengths, where no contribution from DNA base pairs can be detected, indicates that this compound becomes oriented in the flow field, that is the formation of a molecular complex with the macromolecule. Experimental validation on in vitro catalytic cycle of DNA/topoisomerase [4] . Only M382 , M347 Derivatives inhibited the catalytic cycle of topoisomerase II, according to LD outcomes. These data suggest that the intercalation process could be the reason for the cell cycle phase arrest on S phase and inhibiting topoisomerase II catalytic cycle. In silico investigations on DNA interactions. REMARKS & PERSPECTIVES Taking into account all the chemical diversity points in the scaffold, along with the commercial availability of the reactants, the extension of the library of new compounds coupled with further in silico optimization studies could bring to improved results, so as to consider these compounds for preclinical studies [5]. References [1] A.M. Almerico, F. Mingoia, P. Diana, P. Barraia, A. Lauria, A. Montalbano, G. Cirrincione, G. Dattolo, J. Med. Chem., 2005, 48, 2859-2866. [2] A. Lauria, M. Tutone, A.M. Almerico, Eur. J. Med. Chem., 2011, 46, 4274-4280. [3] F. Mingoia, C. Di Sano, F. Di Blasi, M. Fazzari, A. Martorana, A.M. Almerico, A. Lauria, Eur. J. Med Chem., 2013, 64, 345-356. [4] F. Mingoia et al 2016, manuscript in preparation. [5] This work was supported by RSTL. 654 2008 Francesco Mingoia.

The development of new strategies aimed to discover new molecules able to act simultaneously upon multiple biotargets in fighting cancer, is an attractive approach to achieve synergistic effects for new therapeutically perspectives. Recently a new series of pyrazolo[1,2-a]benzo[1,2,3,4]tetrazine-3-one derivatives (PBTs), has been explored as potential anticancer candidates because of their promising antiproliferative activity, apoptosis induction in the low ?M range, as well as cell cycle arrest promoters [1,2]. As an hopeful extension of these preliminary findings, we planned to investigate in depth on the mode of action the selected most active derivatives in an effort to get new insights on their anticancer potential. At first, a DNA targeting is approached by means of flow linear dichroism (LD) experiments to evaluate the ability of a molecule to form an intercalative molecular complex. Additionally, we investigated the capacity of compounds to interfere with the catalytic cycle of topoisomerase II. Depending on the choice of functional groups, PBT scaffold exhibit diverse modes of action, including DNA complexation or inhibition of topoisomeraseII activity. Preliminary in silico insights will also be shown on poster session. [1] A.M. Almerico, F. Mingoia, P. Diana, P. Barraia, A. Lauria, A. Montalbano, G. Cirrincione, G. Dattolo, J. Med. Chem., 2005, 48, 2859-2866. [2] F. Mingoia, C. Di Sano, F. Di Blasi, M. Fazzari, A. Martorana, A.M. Almerico, A. Lauria, Eur. J. Med. Chem.,2013, 64, 345-356.

Thanks to a new synthetic approach recently developed by us1, a new series of pyrrolo[3,2-c]quinoline (PQ) derivatives, has been synthesized permitting us to introduce suitable functional groups in different positions of the scaffold. The latter known for possessing a wide range of biological activities, including anticancer2. The one dose screening (10 ?M) performed by the National Cancer Institute (NCI, USA) against 60 types of tumor cell lines including several panel such as leukemia, lung, renal, CNS, prostate, breast, melanoma, ovarian and colon cancers, allowed us to identify a clear profile of selectivity and potency. Thus, for the most promising derivatives, we undertaken in dept biological investigations. In detail, the 4-substituted piperonil one, which exhibited a preliminary 58% and 18% of growth inhibition in the NCI testing against MCF-7 and MDA-MB-231, respectively, are here submitted to further investigations in order to get new insights on the mode of action. Therefore, a dose response profile at 24, 48 and 72 h was undertaken. More details will be shown at the poster session

Recently, a novel series of derivatives 2a-d, incorporating the new ring system pyrazolo[1,2-a][1,2,5]benzotriazepin-3,6(7H)-dione, has been synthesized and evaluated as anticancer agents. Several derivatives, screened against more than 50 types of human tumor cell lines (NCI), showed moderate antiproliferative activity coupled with a promising selective CNS tumor cell tropism. These derivatives, were designed as evolution of the previously proposed first series of pyrazolo[1,2-a]benzo-[1,2,3,4]tetrazin-3-one derivatives 1 [1] endowed with antiproliferative activity in low micromolar range. A further second series highlighted these derivatives as effective on apoptosis induction, cell cycle arrest on S phase (HeLa). [2] Thus, to investigate the effect of a ring expansion, moving from a tetrazine central moiety to a triazepinone one, we planned to introduce a series of side chains in order to modulate their biological profile. Exploiting the synthetic pathway, an easy access in the functionalization on ring Z was reached obtaining a small library of 20 compounds. Several 6/7-(3-R4 -propyl)-1-methyl-9-R1 -3H,5H-pyrazolo[1,2-a]benzo[1,2,5]triazepin-3-one (3a-d)(1'-5') derivatives were submitted to the MTT assay to assess their grown inhibition activity against HeLa cells. Among them, the most part resulted not active or showed low antiproliferative activity, while the 3c2' derivative revealed as the most effective showing IG50 value of about 27mM. Interestingly, none of the screened derivatives, tested in the concentration range 50-1 ?M, affected the cell membrane integrity, as determined preliminary by the Trypan Blu exclusion method. References 1. A.M. Almerico, F. Mingoia, P. Diana, P. Barraja, A. Lauria, A. Montalbano, G. Dattolo, G. Cirrincione, J. Med. Chem., 2005, 48, 2859-2866. 2. F. Mingoia, C. Di Sano, F. Di Blasi, M. Fazzari, A. Martorana, A. M. Almerico, A. Lauria, Eur. J. Med. Chem., 2013, 64, 345-356.

Protein kinasesfamily plays pivotal roles in nearly every aspectof cellular function. They control metabolism, transcription, cell division and programmed cell death through modification of the protein target function. Typically, the reaction catalyzed by protein kinaseis MgATP1- + protein-O: H -> protein-O: PO32- + MgADP + H+.Protein kinases family has becomeone of the most important drug targets over the past two decades.In particular, most of the FDA-approved protein kinase inhibitors are competitive with respect to ATP. IKK?/?is a member of the protein kinase family and has a central role in NF-?B activationin response to different pro-inflammatory stimuli, since the active form of the enzyme leads to IkB?/?phosphorylation and ultimately to the activation of NF-?B.Our data suggest that ferulic acid (FA) may interfere with IKK/I?B/NF-?B signalling, both reducing NF-?B nuclear translocation and the phosphorylation of IKK?/? and IkB? in LPS-induced Raw 264.7 cells.The regulatory mechanisms governing this phosphorylation event are not well understood. It is known that the molecular mechanism underlying the inhibition of kinase proteins consists of the formation of a ligand-receptor complex. In support, of these experimental outcomes we carried out computational studies by means of Molecular Docking and Induced Fit Docking (IFD) protocols, in order to get new insights at molecular level on the interaction between FA and the human IKK-? (PDB: 4KIK). Furthermore, to better understand the binding mode and the molecular pose, we selected some IKK-beta known ligands such as Staurosporine and BMS 345541, both known as potent ATPase competitors, and also ATP. These studies permitted us to get the preliminary binding affinity on IKK-beta of all the analysed ligands.

Recently, a new series of pyrazolo[1,2-a]benzo[1,2,3,4]tetrazin-3-one derivatives 1, synthesized and submitted to the NCI one dose screening, showed valuable antiproliferative activity reaching in some cases sub-micromolar values. Further investigations, focused on getting new insights on the mechanism of action, provided further support for their proposition as anticancer candidates. Other than the attractive antiproliferative activity, effective apoptosis induction was observed, together with a cell cycle arrest in S phase (HeLa cell). Thus, encouraged by these results, we planned new structural modifications on structure 1 in order to modulate the biological profile. In detail, a ring expansion was approached.The tetrazinecentral moiety was replaced with aseven membered ring [1,2,5]-triazepinoneone(2). Therefore, ten derivatives, variously substituted, were synthesized and submitted to NCI one dose screening. Although literature survey encouraged our research scope because of the wide range of biological properties exhibited by this class of compounds, including anticancer activity, from the NCI screening, only moderate antiproliferative activity was observed. The best value of growth inhibition(GI) was 23% against SNB-75 (CNS cancer), suggesting a central nervous system tropism. Due to the moderate antiproliferative activity showed by the phenyl decorated tricycle (2), we decided to introduce a Chloro-propyl side chain on the expanded ring (3), as shown in Fig.1 with the aim to increase the activity. The biological outcomes provided by the NCI one dose screening evidenced a selective increased antiproliferative activity only towards leukemic tumor cell lines. Detailed results will be shown on poster session.