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Sono stati sintetizzati composti a struttura policiclica complessa. Tutti i composti sintetizzati sono stati saggiati dall'NCI (USA) su un panel di 60 linee cellulari tumorali umane. Sula base dei risultati ottenuti è stato possibile effettuare uno studio SAR che ha evidenziato come la presenza di gruppi idrofili è necessaria ai fini dell'attività antitumorale. E' stato inoltre studiato il meccanismo d'azione di queste molecole. E' stato notato un arresto delle cellule MDA-MB231 in fase G0/G1 correlato con la defosforilazione ed attivazione, indotta dai composti, della proteina pRb. Il meccanismo comunque varia arrestando il ciclo cellulare in fase G2/M quando per effetto della glicosilazione del derivato.

There is an urgent need of anti-virulence agents effective in the prevention or eradication of biofilms that are intrinsically resistant to conventional antibiotics. If we consider that the first step of staphylococcal pathogenesis and of biofilm formation is the bacterial adhesion, promoted by the surface exposed proteins at the cell wall, we believe that new anti-virulence agents could be developed by using as a target the Sortase A (SrtA), the enzyme responsible of linking surface exposed proteins to peptidoglycan. Therefore, SrtA inhibitors could act as anti-adhesion agents useful to prevent Gram positive virulence mechanisms as well as a virulence mechanism based on biofilm formation. Starting from these consideration and in continuing our research work on antibacterial/antibiofilm agents, we thought it was of interest to obtain novel SrtA inhibitors, showing a molecular diversity than the known ones.The esters and the acids were tested for their inhibitory activity on SrtA, utilizing the FRET technics (Fluorescenze Resonance Energy Transfer) [1]. Some of them showed IC50 values in the range 50-100 μM. For a preliminary assessment of antivirulence properties the most active SrtA inhibitors were tested for their ability to interfere with biofilm formation of S. aureus ATCC 29213, S. aureus ATCC 25923, S.aureus ATCC 6538 and S. epidermidis RP62A. We found that the above derivatives interfere with biofilm formation

Gram-positive bacteria are a significant cause of nosocomial and community-acquired infections associated with diseases of high morbidity and mortality. Moreover, antibiotic resistance of important Gram-positive pathogens, such as Staphylococcus aureus, Streptococcus pneumoniae and Enterococcus faecalis is one of the major worldwide health problems. Over the last decade, many studies have focused on agents that target the virulence of important pathogens without killing or inhibiting their growth therefore imposing limited selective pressure to promote the development and spread of resistance mechanisms [1]enterococci and streptococci, sortase A plays a critical role in Gram-positive bacterial pathogenesis. It is thus considered a promising target for the development of new anti-infective drugs that aim to interfere with important Gram-positive virulence mechanisms, such as adhesion to host tissues, evasion of host defences, and biofilm formation. The additional properties of sortase A as an enzyme that is not required for Gram-positive bacterial growth or viability and is conveniently located on the cell membrane making it more accessible to inhibitor targeting, constitute additional reasons reinforcing the view that sortase A is an ideal target for anti-virulence drug development. Many inhibitors of sortase A have been identified to date using high-throughput or in silico screening of compound libraries (synthetic or natural. We synthesized two classes of molecules: curcumin analogues and N-phenyl-1H-pyrazole-4-carboxamides able to interfere with the bacterial adhesion, a fundamental step of Gram-positive pathogenesis [2].All derivatives obtained were tested for both the planktonic growth inhibition activity and the inhibition of biofilm formation against the following reference bacterial strains: S. aureus ATCC 6538, S. aureus ATCC 25923 and S. aureus ATCC 29213.

The reaction of the diazonium salt 12 derived from N-(2-aminophenyl)-N,3-dimethyl-1-phenyl-1H-pyrazole-5-carboxamide with copper sulfate and sodium chloride in the presence of ascorbic acid afforded the unexpected products 3,5-dimethyl-1-phenyl-1,5-dihydro-4H-pyrazolo[4,3-c]¬quinolin-4-one (17) and N-methyl-2-(3-methyl-1-phenyl-1H-pyrazol-5-yl)aniline (19), accompanied by N-(2-chlorophenyl)-N,3-dimethyl-1-phenyl-1H-pyrazole-5-carboxamide (18). Products 17 and 19 are formed via a non-classical Pschorr reaction. The formation of 17 represents an alternative to the literature synthesis of this biologically active compound. The molecular structure of 18 was confirmed by single-crystal X-ray analysis.

Staphylococcus aureus, due to its wide arsenal of virulence factors, is a very versatile pathogen responsible for a wide variety of infectious diseases. The virulence factors include the cell-wall associated proteins that have a direct role in the first stage of pathogenesis. The Sortase A (SrtA) transpeptidase is responsible for covalent anchoring to the cell wall of various surface proteins and it is considered a good target to design new antivirulence agents. In this study, we report the identification of an inhibitor of SrtA afforded from the random screening of a small molecular library of around 150 synthetic compounds, through a high throughput assay by using the standard Dabcyl-QALPETGEE-Edans fluorescence resonance energy transfer (FRET)-peptide substrate for measurement of enzyme activity. Such hit compound showed an IC50 value of 80µM. A group of derivatives of the hit compound has been obtained and evaluated for their activity as SrtA inhibitors. The efficacy of the most interesting SrtA inhibitors needs to be evaluated by in vivo models of infection, but we used a simple methodology in vitro, such as inhibition of biofilm formation, for a preliminary assessment of antivirulence properties of some of these novel inhibitors of SrtA. An interesting correlation between inhibition of SrtA and interference with the formation of staphylococcal biofilms has been observed in many cases especially at a concentration equal or more than IC50 determined as SrtA inhibitors.

By reacting methylaminopyrazoles with hexane-2,5-dione in 1,4-dioxane in the presence of p-toluensulfonic acid, new derivatives containing the policyclic sistem 5,7:7,13-dimethanopyrazolo[3,4-b]pyrazolo[3'.4':2,3]azepino[4,5-f]azocine were obtained. The new synthesized compounds were tested preliminarly at 10 microM against five human cancer cell lines showing a range of inhibition of 20-62% against the most susceptible cell lines K562 and HCT116.

Indazole nucleus represents a very attractive scaffol to obtain new molecole endowed with antineoplastic activity. On the basis of these literature data we have designed some indazole derivatives such as N-indazolylbenzamides and N-indazolyl-N’-phenylureas as potential CDK1 inhibitors. In fact the above compounds contain the structural feature, common to the majority of CDK inhibitors, requested to make hydrogen bonds with the molecular forks present in the hinge region of CDKs. The N-indazolylbenzamides 1 were obtained by reacting aminoindazoles and substituted benzoylchlorides. Among the synthesized compounds some derivatives 1 resulted to be CDK1 inhibitors showing IC50 values in the range 4.4-10.0 M

The cinnamoyl anthranilamides represent a class of biological active substances of great importance in medicinal chemistry. Moreover, despite their wide range of biological activities, a review of the literature revealed that no anticancer activity is described for this kind of substances. Starting from the 2-cinnamamido-5-iodobenzamide, resulted able to inhibit the leukemic cell line K-562 proliferation with a percent of inhibition of 74% at 10M concentration, we undertake the following structural modifications on cinnamamidobenzamide skeleton: the introduction of various substituents both on the benzamido and the cinnamamido moieties, the substitution of olefinic bond with the ethane, ethyne, cyclopropane and phenyl groups. Synthesized compounds caused growth inhibition against many tumor cell lines at low micromolar and submicromolar concentrations against every tumor cell line investigated The best activity was obtained when the 5 position of the benzamido moiety was substituted with an iodine moiety. COMPARE analysis, effects on tubulin polymerization in cells and with purified tubulin, and effects on cell cycle distribution (G2-M phase block), including induction of apoptosis, indicate that these new antiproliferative compounds act as antitubulin agents. Preliminary biological data on 2-(3-phenylpropiolamido)benzamides, bearing an ethyne bond, showed a different mechanism of action considering that they caused a G0-G1 block of the cell cycle on K526 cell line.

Recently our research group has reported the synthesis of some 3-amino-N-phenyl-1H-indazole-1-carboxamides able to inhibit at low micromolar concentrations the cell growth of many neoplastic cell lines. The above compounds are unsubstituted in the indazole nucleus and this gives the hope to obtain more active compounds if appropriate substituents are beared by the above nucleus. So, several new N-phenyl-1H-indazole-1-carboxamides 1c-h and 4l,m were prepared by reacting phenyl isocyanates 3a,b with 3-amino-1H-indazoles 2c,e,g, or 1H-indazole 2l respectively. Chemical trasformations of compounds 1a,b and 1g,h gave 3-acetamido-N-phenyl-1H-indazole-1-carboxamides 5a,b, and 3,5-diamino-N-phenyl-1H-indazole-1-carboxamides 4i,j respectively. Finally, 3,5-diacetamido-N-phenyl-1H-indazole-1-carboxamides 6a,b were prepared by acethylation of 4i,j. Some of synthesized compounds were evaluated for their in vitro antiproliferative activity against 60 human tumoral cell lines derived from nine clinically isolated cancer types (leukemia, non-small cell lung, colon, CNS, melanoma, ovarian, renal, prostate and breast). Compound 1c, the most active of the series, was able to inhibit cell growth showing GI50 values in the 0.041-33.6 μM range, being very effective against colon and melanoma cell lines. Compound 1c induced an increase of cells in G0-G1 peak and a decrease of cells in S and G2-M phases when used at a concentration of or higher than 40µM which is the TGI calculated after 24 h of 1c exposure. This could be due to a block of G1-S transition as indicated by the ability of 1c to increase the hypophosphorylate pRb/ total pRb ratio.

Many infections such as otitis media, sinusitis, cholesteatoma, tonsillitis and adenoiditis are caused by biofilm forming mucosal pathogens (P. aeruginosa, S. aureus, S. peneomoniae, H. influenzae and M. catarrhalis). Moreover, the role of biofilms in the chronic otolaryngologic infections has been recognized for otitis media, tonsillitis and rhinosinusitis. Finally, bacterial biofilms of S. aureus, S. epidermis and E. faecalis are the leading cause of medical device-related infections. Pathogens growing as biofilms are intrinsically resistant to conventional antibiotics and therefore the discovery of new compounds able to act against biofilm aggregated micro organisms is an urgent task. Previously, we reported the study on the antibiofilm activity of 4-diazopyrazole derivatives of type 1 against S. aureus ATCC 29213, methycillin-resistant S. aureus ATCC 43866 and methycillin-resistant S. epidermis RP626 preformed biofilms. The compounds were active at the maximum tested concentration of 25 microg/ml. On the basis of these results, we have prepared a new series of 4-diazopyrazole derivatives 2a-i and 3 which take the phenylureido or phenylacetamido moiety at 5-position of the pyrazole nucleus. The compounds has been tested preliminarly against planktonic (single cells) strains of S. aureus (ATCC 25923), S. aureus (ATCC 29213), E. coli (ATCC 25922) and P. aeruginosa (ATCC 9027), showing MIC’s values in the range 50-0.097 microg/ml. Experimental work aimed to verify the effect of the above compounds on the corresponding microbial biofilms are in progress.

Some natural and synthetic related pyrrolomycins, a family of halogenated pyrrole antibiotics, showed anti-biofilm properties in vitro at low concentration (0.045μg/mL) against preformed staphylococcal biofilms. Moreover, considering the human cell toxicity, the selectivity indexes (ratio of cytotoxicity to antibiofilm activity) of some of them were very interesting. The present study aims to investigate if the pyrrolomycins could also prevent staphylococcal biofilm formation. The evaluation of S.aureus ATCC 25923 biofilm formation inhibition was conducted by safranin staining method. At tested concentrations of 0.18, 0.09, 0.045 μg/mL (concentrations much lower than MIC value determined on planktonic strain) the novel pyrrolomycin derivative IV resulted effective as biofilm inhibitor showing inhibition percentages ranging from 56.5 to 29% against S.aureus ATCC 25923. We are investigating if sortase A which is responsible for the anchoring of surface proteins to Gram positive cell wall, could be the rational target of pyrrolomycins. The surface proteins play pivotal roles in the adhesion to host’s tissues, and the evasion of host-immune responses, moreover they facilitate attachment on biological and abiotic surfaces in the first steps of biofilm formation. It has been observed that inhibition of sortase A by different chemicals resulted in decrease of virulence and staphylococcal biofilm formation. A molecular modeling study conducted by using a crystal structure of sortase A, recovered from protein data bank, and by studying docking properties of a known sortase A inhibitor and of pyrrolomycins, is in progress. In vitro experiments on sortase inhibition activity are needed to confirm the computational results

The pyrazolo[3,4-d]pyrimidine system shows a multitude of interesting pharmacological properties. Owing to the potential anti-inflammatory activity of 5-benzamido-pyrazolo[3,4-d]pyrimidin- 4-one derivatives and considering the easy synthesis of this class of compounds, a set of new 5- benzamido-1H-pyrazolo[3,4-d]pyrimidin-4-ones has been prepared in 42-80% yields by reacting 5- aminopyrazole-4(N-benzoyl)carbohydrazide derivatives and the opportune triethylorthoesters. Compounds 8a, b, 10a–d, and 11a, b revealed a superior inhibitory profile against COX-2, when compared to that of reference standards NS398 and indomethacin. Molecular modelling studies confirmed the obtained biological results.