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Tyrosine kinase inhibitors (TKIs) represent one of the most advanced class of therapeutics for cancer treatment. Most of them are also cytochrome P450 (CYP) inhibitors and/or substrates thereof. Accordingly, their efficacy and/or toxicity can be affected by CYP-mediated metabolism and by metabolism-derived drug-drug interactions. In order to enhance the therapeutic performance of these drugs, we developed a prodrug (Pro962) of our TKI TK962 specifically designed for liposome loading and pH-controlled release in the tumor. A cholesterol moiety was linked to TK962 through pH-sensitive hydrazone bond for anchoring to the liposome phospholipid bilayer to prevent leakage of the prodrug from the nanocarrier. Bioactivity studies performed on isolated target kinases showed that the prodrug maintains only partial activity against them and the release of TK962 is required. Biopharmaceutical studies carried out with prodrug loaded liposomes showed that the prodrug was firmly associated with the vesicles and the drug release was prevented under blood-mimicking conditions. Conversely, conventional liposome loaded with TK962 readily released the drug. Flow cytometric studies showed that liposomes efficiently provided for intracellular prodrug delivery. The use of the hydrazone linker yielded a pH-controlled drug release, which resulted in about 50% drug release at pH 4 and 5 in 2 h. Prodrug, prodrug loaded liposomes and active lead compound have been tested against cancer cell lines in either 2D or 3D models. The liposome formulation showed higher cytotoxicity than the unformulated lead TK962 in both 2D and 3D models. The stability of prodrug, prodrug loaded liposomes and active lead compound in human serum and against human, mouse, and rat microsomes was also assessed, demonstrating that liposome formulations impair the metabolic reactions and protect the loaded compounds from catabolism. The results suggest that the liposomal formulation of pH releasable TKI prodrugs is a promising strategy to improve the metabolic stability, intracellular cancer cell delivery and release, and in turn the efficacy of this class of anticancer drugs.

A small library of new angelicin derivatives was designed and synthesized with the aim of bypassing the side effects of trimethylangelicin (TMA), a promising agent for the treatment of cystic fibrosis. To prevent photoreactions with DNA, hindered substituents were inserted at the 4 and/or 6 positions. Unlike the parent TMA, none of the new derivatives exhibited significant cytotoxicity or mutagenic effects. Among the synthesized compounds, the 4-phenylderivative 12 and the 6-phenylderivative 25 exerted a promising F508del CFTR rescue ability. On these compounds, preliminary in vivo pharmacokinetic (PK) studies were carried out, evidencing a favorable PK profile per se or after incorporation into lipid formulations. Therefore, the selected compounds are good candidates for future extensive investigation to evaluate and develop novel CFTR correctors based on the angelicin structure.

A series of new-generation TMA (4,6,4′-trimethyl angelicin) analogues was projected and synthetized in order to ameliorate anti-inflammatory activity, with reduced or absent toxicity. Since the NF-κB transcription factor (TF) plays a critical role in the expression of IL-8 (Interluekin 8), a typical marker of lung inflammation in Cystic Fibrosis (CF), the use of agents able to interfere with the NF-κB pathway represents an interesting therapeutic strategy. Through preliminary EMSA experiments, we identified several new TMA derivatives able to inhibit the NF-κB/DNA complex. The selected active molecules were then analyzed to evaluate the anti-inflammatory effect using both Pseudomonas aeruginosa (PAO1) infection and TNF-alpha stimulus on the CF IB3-1 cell line. It was demonstrated that mainly two TMA analogues, GY971a mesylate salt (6-p-minophenyl-4,4′-dimethyl-angelicin) and GY964 (4-phenyl-6,4′-dimethyl-angelicin), were able to decrease the IL-8 gene expression. At the same time, these molecules were found to have no pro-apoptotic, mutagenic and phototoxic effects, facilitating our decision to test the efficacy in vivo by using a mouse model of acute P. aeruginosa lung infection. The anti-inflammatory effect of GY971a was confirmed in vivo; this derivative was able to deeply decrease the total number of inflammatory cells, the neutrophil count and the cytokine/chemokine profile in the P. aeruginosa acute infection model, without evident toxicity. Considering all the obtained and reported in vitro and in vivo pre-clinical results, GY971a seems to have interesting anti-inflammatory effects, modulating the NF-κB pathway, as well as the starting lead compound TMA, but without side effects.

Simple Summary The Epidermal Growth Factor Receptor (EGFR) is a receptor protein involved in many types of cancers. EGFR can be inhibited by monoclonal antibodies (protein drugs acting on the extracellular domain of the protein) or by ATP-mimic compounds (small molecule drugs blocking the intracellular domain). Here we report the identification of a novel potential class of drugs, i.e., small molecules acting on the extracellular domain of EGFR. The identified compounds modified the trafficking of EGFR and induced cytotoxicity in cells overexpressing EGFR and insensitive to monoclonal antibodies being active in cell lines bearing KRAS mutations. The Epidermal Growth Factor Receptor (EGFR) is a transmembrane glycoprotein belonging to the protein kinase superfamily. It is composed of an extracellular domain, a transmembrane anchoring region and a cytoplasmic region endowed with tyrosine kinase activity. Genetic mutations of EGFR kinase cause higher activity thereby stimulating downstream signaling pathways that, in turn, impact transcription and cell cycle progression. Due to the involvement of mutant EGFR in tumors and inflammatory diseases, in the past decade, several EGFR inhibitory strategies have been extensively studied, either targeting the extracellular domain (through monoclonal antibodies) or the intracellular kinase domain (through ATP-mimic small molecules). Monoclonal antibodies impair the binding to growth factor, the receptor dimerization, and its activation, whereas small molecules block the intracellular catalytic activity. Herein, we describe the development of a novel small molecule, called DSF-102, that interacts with the extracellular domain of EGFR. When tested in vitro in KRAS mutant A549 cells, it impairs EGFR activity by exerting (i) dose-dependent toxicity effects; (ii) a negative regulation of ERK, MAPK p38 and AKT; and (iii) a modulation of the intracellular trafficking and lysosomal degradation of EGFR. Interestingly, DSF-102 exerts its EGFR inhibitory activity without showing interaction with the intracellular kinase domain. Taken together, these findings suggest that DSF-102 is a promising hit compound for the development of a novel class of anti-EGFR compounds, i.e., small molecules able to interact with the extracellular domain of EGFR and useful for overcoming the KRAS-driven resistance to TKI treatment., Cancers, 14 (15), ISSN:2072-6694

Glioblastoma multiforme (GBM), a malignant tumor of the central nervous system, has a high mortality rate. No curative treatment is presently available, and the most commonly used chemotherapeutic drug, the alkylating agent temozolomide (TMZ), is only able to increase life expectancy and is often associated with drug resistance. Therefore, an urgent need does exist for novel drugs aimed at treating gliomas. In the present study, we obtained three major results using corilagin: (a) demonstrated that it inhibits the growth of U251 glioma cells through activation of the apoptotic pathway; (b) demonstrated that it is also active on TMZ-resistant T98G glioma cells; and (c) demonstrated that when used in combination with TMZ on T98G glioma cells, a higher level of proapototic and antiproliferative effects is observed. Our study indicates that corilagin should be investigated in more detail to determine whether it can be developed as a potential therapeutic agent. In addition, our results suggest that corilagin could be used in combination with low doses of other standard anticancer chemotherapeutic drugs against gliomas (such as TMZ) with the aim of obtaining enhanced anticancer effects.

A series of trimethylangelicin (TMA) derivatives were designed and synthesized to overcome the unwanted effects of TMA, promising agent for treatment of inflammation-related diseases and other pathologies, such as cystic fibrosis. The new generation TMA analogues bore hindered substituents at the 4 position in order to minimize or avoid the photoreactions with DNA. Among them, the 4-isopropyl-6-ethyl derivative 23 exhibited TMA-like inhibitory activity on NF-κB/DNA interactions but it proved unable to photoreact with pyrimidine bases of DNA, nor to induce any other DNA damage. The isopropyl analogue 23 was proven to lack mutagenicity when assayed through Ames test and exhibited no anti-proliferative activity on cystic fibrosis IB3-1 cells, displaying at the same time inhibition of the TNF-α induced release of the NF-κB regulated PDGF-B chain, IL-10, IL-15, IL-17 and IFN-γ. Therefore compound 23 deserves further assay to determine its anti-inflammatory properties, since it lacks photoreaction properties and mutagenicity-related side effects.

Recent studies have identified and characterized a novel putative transcriptional repressor site in a 5' untranslated region of the Aγ-globin gene that interacts with the Ly-1 antibody reactive clone (LYAR) protein. LYAR binds the 5'-GGTTAT-3' site of the Aγ-globin gene, and this molecular interaction causes repression of gene transcription. In β-thalassemia patients, a polymorphism has been demonstrated (the rs368698783 GA polymorphism) within the 5'-GGTTAT-3' LYAR-binding site of the Aγ-globin gene. The major results gathered from surface plasmon resonance based biospecific interaction analysis (SPR-BIA) studies (using crude nuclear extracts, LYAR-enriched lysates, and recombinant LYAR) support the concept that the rs368698783 GA polymorphism of the Aγ-globin gene attenuates the efficiency of LYAR binding to the LYAR-binding site. This conclusion was fully confirmed by a molecular docking analysis. This might lead to a very important difference in erythroid cells from β-thalassemia patients in respect to basal and induced levels of production of fetal hemoglobin. The novelty of the reported SPR-BIA method is that it allows the characterization and validation of the altered binding of a key nuclear factor (LYAR) to mutated LYAR-binding sites. These results, in addition to theoretical implications, should be considered of interest in applied pharmacology studies as a basis for the screening of drugs able to inhibit LYAR-DNA interactions. This might lead to the identification of molecules facilitating induced increase of γ-globin gene expression and fetal hemoglobin production in erythroid cells, which is associated with possible reduction of the clinical severity of the β-thalassemia phenotype. Graphical abstract.

An unpredicted condensation of naphthylamine with two molecules of ethyl propiolate yields directly carbethoxy benzoquinoline in high yield. Some benzoquinoline carboxamide derivatives with protonatable side chains were then synthesized and evaluated for antiproliferative activity on human tumor cell lines. The most active compound (7a) demonstrated to intercalate into DNA and to inhibit the relaxation activity mediated by topoisomerase II.

Introduction: Cell cycle and gene transcription are under the control of cyclin-dependent kinases (CDKs), whose activity depends on the binding with cyclins. Deregulated CDK activities have been reported in a majority of human cancers, representing potential therapeutic targets. Areas covered: This review provides preclinical and clinical (phase I/II) updates of promising therapeutic compounds targeting CDKs published between 2013 and 2016 Expert opinion: First generation pan-CDK inhibitors showed marked toxicity in clinical trials and most compounds were discontinued. Despite their failure was ascribed also to inadequate patient selection rules, novel pan-CDK inhibitors have entered clinical trials with still poorly defined selection strategies. The most interesting results have been obtained with dual CDK4/6 inhibitors and through a more accurate evaluation of predictive biomarkers, suggesting the usefulness of CDK inhibitors for personalized treatment. The increased knowledge on the roles of CDKs in cell cycle and gene transcription suggests to review also the anticancer potential of first generation CDK inhibitors by defining more appropriate rules for patients engagement. Recent findings has highlighted CDK8 as a novel target for cancer treatment. Indeed some biomarkers for CDK8 inhibition sensitivity have already been proposed. CDK8 inhibition is also supposed to prevent cancer metastasis.

Quinoline core has been shown to possess a promising role in the development of anticancer agents. However, the correlation between its broad spectrum of bioactivity and the underlying mechanism of actions is poorly understood. The present study, with the use of bioinformatics approaches, reported a series of designed molecules which integrated quinoline core and sulfonyl moiety, with the objective of evaluating the substituent and linker effects on anticancer activities and associated mechanistic targets. We identified potent compounds (1h, 2h, 5 and 8) exhibiting significant anticancer effects towards liver cancer cells (Hep3B) with the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) relative values of cytotoxicity below 0.40, a value in the range of doxorubicin positive control with the value of 0.12. Bulky substituents and the presence of bromine atom, as well as the presence of sulfonamide linkage, are likely the favorable structural components for molecules exerting a strong anticancer effect. To the best of our knowledge, our findings obtained from chemical synthesis, in vitro cytotoxicity, bioinformatics-based molecular docking analysis (similarity ensemble approach, SEA),and electrophoretic mobility shift assay provided the first evidence in correlation to the anticancer activities of the selected compound 5 with the modulation on the binding of transcription factor NF-κB to its target DNA. Accordingly, compound 5 represented a lead structure for the development of quinoline-based NF-κB inhibitors and this work added novel information on the understanding of the mechanism of action for bioactive sulfonyl-containing quinoline compounds against hepatocellular carcinoma.

Simplified analogues of previously reported NF-κB interaction inhibitors, lacking the furan moiety, were synthesized and evaluated by performing experiments based on electrophoretic mobility shift assay (EMSA). The synthetic modifications led to simpler coumarin derivatives with lower activity allowing to better understand the minimal structural requirement for the binding to NF-κB.

Oncogenic activation of RET kinase has been found in several neoplastic diseases, like medullary thyroid carcinoma, multiple endocrine neoplasia, papillary thyroid carcinoma, and non-small-cell lung cancer. Currently approved RET inhibitors were not originally designed to be RET inhibitors, and their potency against RET kinase has not been optimized. Hence, novel compounds able to inhibit both wild-type RET (

The angelicin analogue 4,6,4′-trimethylangelicin (TMA) was recently reported as a strong inhibitor of nuclear factor-κB (NF-κB) activity and of the expression of the interleukin-8 (IL-8) gene in bronchial epithelial cells in which the inflammatory response has been challenged withP. aeruginosa, the most common bacterium found in the airways of patients affected by cystic fibrosis (CF). These findings encouraged us to analyze new synthetic analogues of TMA in order to evaluate their biological activities on human bronchial epithelial CF IB3-1 cells and to find more potent anti-NF-κB agents exhibiting only minor antiproliferative effects. Analogues able to inhibit NF-κB/DNA interaction at lower concentration than TMA were found and selected to investigate their biological activity on IB3-1 cells induced with TNF-α. In this biological system, NF-κB-mediated IL-8 gene expression was investigated. Some analogues showed similar activity to the lead compound TMA. Other analogues displayed higher activities; in particular, the most interesting compounds showing relevant anti-inflammatory effects were found to cause 56–83% reduction of IL-8 mRNA expression at low concentrations (1–10 μM), without changes in cell proliferation pattern, demonstrating their potential interest for a possible development of anti-inflammatory therapy of cystic fibrosis.

Oncogenic activation of RET kinase has been found in several neoplastic diseases, like medullary thyroid carcinoma, multiple endocrine neoplasia, papillary thyroid carcinoma, and non-small-cell lung cancer. Currently approved RET inhibitors were not originally designed to be RET inhibitors, and their potency against RET kinase has not been optimized. Hence, novel compounds able to inhibit both wild-type RET (wt RET) and its mutants (e.g., V804M RET) are needed. Herein we present the development and the preliminary evaluation of a new sub-micromolar wt RET/V804M RET inhibitor, N-(2-fluoro-5-trifluoromethylphenyl)-N'-{4'-[(2''-benzamido)pyridin-4''-ylamino]phenyl}urea (69), endowed with a 4-anilinopyridine structure, starting from our previously identified 4-anilinopyrimidine hit compound. Profiling against a panel of kinases indicated 69 as a multi cKIT/wt RET/V804M RET inhibitor.

Several DNA polymorphisms have been associated with high production of fetal hemoglobin (HbF), although the molecular basis is not completely understood. In order to identify and characterize novel HbF-associated elements, we focused on five probands and their four families (from Egypt, Iraq and Iran) with thalassemia major (either β(0)-IVSII-1 or β(0)-IVSI-1) and unusual HbF elevation (98 %), congenital or acquired after rejection of bone marrow transplantation, suggesting an anticipated favorable genetic background to high HbF expression.Patient recruitment, genomic DNA sequencing, western blotting, electrophoretic mobility shift assays, surface plasmon resonance (SPR) biospecific interaction analysis, bioinformatics analyses based on docking experiments.A polymorphism of the Aγ-globin gene is here studied in four families with β(0)-thalassemia (β(0)-IVSII-1 and β(0)-IVSI-1) and expressing unusual high HbF levels, congenital or acquired after rejection of bone marrow transplantation. This (G→A) polymorphism is present at position +25 of the Aγ-globin genes, corresponding to a 5'-UTR region of the Aγ-globin mRNA and, when present, is physically linked in chromosomes 11 of all the familiar members studied to the XmnI polymorphism and to the β(0)-thalassemia mutations. The region corresponding to the +25(G→A) polymorphism of the Aγ-globin gene belongs to a sequence recognized by DNA-binding protein complexes, including LYAR (Ly-1 antibody reactive clone), a zinc-finger transcription factor previously proposed to be involved in down-regulation of the expression of γ-globin genes in erythroid cells.We found a novel polymorphism of the Aγ-globin gene in four families with β(0)-thalassemia and high levels of HbF expression. Additionally, we report evidence suggesting that the Aγ-globin gene +25(G→A) polymorphism decreases the efficiency of the interaction between this sequence and specific DNA binding protein complexes.

Some new 3-substituted quinazolinones were synthesized and evaluated as inhibitors of kinases involved in fibrogenic process. The compounds were tested against a panel of both tyrosine and serine-threonine kinases. The profile of selectivity of some representative compounds was investigated through molecular docking studies. The most interesting compounds were also evaluated in vitro as potential agents for the treatment of fibrotic diseases. Quinazolinone derivatives reduced proliferation and expression of genes involved in the fibrogenic process in hepatic stellate cells (HSCs) and intestinal subepithelial myofibroblasts (ISEMFs). Furthermore some compounds downregulated phosphorylation of p38MAPK. Our findings provide evidences that 3-substituted quinazolinones target multiple essential pathways of the fibrogenic process.

A new series of benzo[h]quinazoline and benzo[f]quinazoline derivatives was prepared and studied for the biological activity. The compounds carrying a dimethylaminoethyl side chain (6c, 8c and 12) inhibit cell growth. The ability to form a molecular complex with DNA and to interfere with topoII and topoI relaxation activity was evidenced for the most active 6c and 8c, along with the capacity to induce apoptosis on HeLa cells.

Some 4,8-dimethyl-3-psoralenacetic acids were synthesized and studied. All the designed psoralenacetic acids bear alkyl or cycloalkyl substituents at the furan ring. These psoralenacetic acids were shown to be a novel class of psoralen derivatives characterized by an interesting photobiological profile. The carboxylic group at the 3 position, useful to confer hydrophilic properties, appears to be detrimental to the classical intercalation into DNA, likely because of repulsive interactions with the negative surface of the macromolecule. Nevertheless, the new derivatives possess a notable photoantiproliferative activity, due to a peculiar mechanism of action consisting of a decarboxylation step before exerting their photobiological activity. The most active compound 2 is able to induce a noteworthy photocytotoxic effect, with GI50 values being submicromolar on human tumor cell lines and no effect in the dark. The involvement of DNA photoaddition after UVA light-mediated decarboxylation and ROS formation is responsible for its biological activity, as demonstrated comparing the activity profile of the decarboxylated analogue. However, other biological targets seem to be involved in the photooxidative damage, such as proteins. Compound 2 could thus be considered as a prodrug, inactive without UVA light but activated upon specific irradiation, thus preventing unselective side effects and opening new perspectives on agents useful in photochemotherapy.

Kinase inhibitors are attractive drugs/drug candidates for the treatment of cancer. The most recent literature has highlighted the importance of multi target kinase inhibitors, although a correct balance between specificity and non-specificity is required. In this view, the discovery of multi-tyrosine kinase inhibitors with subfamily selectivity is a challenging goal. Herein we present the synthesis and the preliminary kinase profiling of a set of novel 4-anilinopyrimidines. Among the synthesized compounds, the N-phenyl-N’-[4-(pyrimidin-4-ylamino)phenyl]urea derivatives selectively targeted some members of class III receptor tyrosine kinase family. Starting from the structure of hit compound19 we synthesized a further compound with an improved affinity toward the class III receptor tyrosine kinase members and endowed with a promising antitumor activity both in vitro and in vivo in a murine solid tumor model. Molecular modeling simulations were used in order to rationalize the behavior of the title compounds.

Simplified analogues of previously reported NF-κB interaction inhibitors, lacking the furan moiety, were synthesized and evaluated by performing experiments based on electrophoretic mobility shift assay (EMSA). The synthetic modifications led to simpler coumarin derivatives with lower activity allowing to better understand the minimal structural requirement for the binding to NF-κB.

A new synthetic pathway to quinazolines is described. This new method uses hexamethylenetetramine in TFA and potassium ferricyanide in aqueous ethanolic KOH, starting from simple N-protected anilines. The method affords substituted quinazolines with high selectivities and good yields, reducing reaction-time and work-up operations.

4-Hydroxymethyl-1,6,8-trimethylfuro[2,3-h]quinolin-2(1H)-one (HOFQ) was prepared by a new profitable way, which allowed to synthesize also 4-methoxymethyl-1,6,8-trimethylfuro[2,3-h]quinolin-2(1H)-one (MOFQ), and 4-hydroxymethyl-6,8-dimethylfuro[2,3-h]quinolin-2(1H)-one (HOHFQ). Some biological activities of the three compounds were studied in comparison with 8-MOP. In the dark, they inhibited topoisomerase II, leading to a moderate antiproliferative activity in mammalian cells. The antiproliferative activity was also tested upon UVA irradiation in mammalian cells: all compounds showed higher activity than 8-MOP, without mutagenicity and skin phototoxicity, with the best results for HOFQ. Photobinding to DNA was investigated, demonstrating a different sequence specificity for these furoquinolinones in comparison with furocoumarins. For all these features, HOFQ and the other analogues appeared very promising photochemotherapeutic agents, whose mechanism of action will be further investigated.

Electrospray mass spectrometry (ES-MS) was successfully employed for the structural differentiation of six isomeric trimethylfurocoumarins of possible pharmaceutical interest. Two different approaches were employed. The first was based on MSn experiments of MH+ ions. Although the product ion spectra of MH+ of the isomers are very similar, the MS3 spectra of the collisionally generated [MHCO]+ ions show some characteristic differences. The second approach was based on complexation of the molecules with Li+, Na+ and K+ using ESI-MS of sample solutions containing alkali ions in a 100:1 molar ratio with respect to the analyte. Significant differences were observed in complex production yields, and these were related to the dimension of the alkali ion and to the steric availability of chelating groups in the different isomers. Copyright © 2003 John Wiley & Sons, Ltd.

Cell cycle experiments with our previously reported 4-biphenylaminoquinazoline (1–3) multityrosine kinase inhibitors revealed an activity profile resembling that of known tubulin polymerization inhibitors. Novel 4-biarylaminoquinazoline analogues of compound 2 were synthesized and evaluated as inhibitors of several tyrosine kinases and of tubulin. Although compounds 1–3 acted as dual inhibitors, the heterobiaryl analogues possessed only anti-tubulin properties and targeted the colchicine site. Furthermore, molecular modeling studies allowed the rationalization of the pharmacodynamic properties of the compounds.

A new series of pyrroloquinolinones bearing different alkylamino side chains were synthesized and evaluated as cytotoxic compounds against three different human tumor cell lines (HeLa, HL-60 and A431). Some compounds showed interesting antiproliferative activity, in particular against A431 cells. The compounds were tested for their ability to counteract topoisomerase II relaxation activity and the most interesting one (3c) was tested also against topoisomerase I, resulting a dual inhibitor. The molecular interactions between 3c and the intracellular targets were finally investigated through molecular modeling simulations.

Linear (L) and angular (A) 4',5'-dimethylfuroquinazolines (FQZs) were synthesized and studied as furocoumarin analogs. These molecules proved to be photounstable with a photodegradation extent correlated to UVA light doses. Both compounds did intercalate inside the DNA double helix, but were not able to photobind DNA bases under UVA irradiation. This behavior was further rationalized through docking studies. The photosensitizing effects of these compounds were evaluated on Jurkat tumor cells and NCTC-2544 human keratinocytes, with and without antioxidants, to demonstrate the involvement of a photodynamic mechanism. Indeed, significant amounts of singlet oxygen and superoxide anion were generated in the presence of both compounds, that account for the oxidative damage induced to some isolated biological substrates (DNA, amino acids, proteins and lipids). Photophysical studies by use of a flash photolysis set up showed detectable triplet population and production of singlet reactive oxygen species for linear furoquinazoline, which can be responsible for the oxidation of biological substrates, and therefore can affect the cell proliferation.

Microtubules are high dynamic protein filaments fundamental for cells growth and proliferation. Hence, tubulin inhibitors are useful anticancer compounds. Three major binding site have been identified in tubulin, on the basis of known ligands: the vinca domain, the colchicine domain and the taxane domain. Several compounds able to bind the colchicine and the taxane domains have been to date synthesized and evaluated. In this review we give a description of the developed QSAR and 3D-QSAR models, giving particular attention to those studies that give structural insight in the binding modes of compounds with the target.

The selection of the most appropriate protein conformation is a crucial aspect in molecular docking experiments. In order to reduce the errors arising from the use of a single protein conformation, several authors suggest the use of several tridimensional structures for the target. However, the selection of the most appropriate protein conformations still remains a challenging goal. The protein 3D-structures selection is mainly performed based on pairwise root-mean-square-deviation (RMSD) values computation, followed by hierarchical clustering. Herein we report an alternative strategy, based on the computation of only two atom affinity map for each protein conformation, followed by multivariate analysis and hierarchical clustering. This methodology was applied on seven different kinases of pharmaceutical interest. The comparison with the classical RMSD-based strategy was based on cross-docking of co-crystallized ligands. In the case of epidermal growth factor receptor kinase, also the docking performance on 220 known ligands were evaluated, followed by 3D-QSAR studies. In all the cases, the herein proposed methodology outperformed the RMSD-based one.

In this work the synthesis and the biological evaluation of some novel anilinoquinazoline derivatives carrying modifications in the quinazoline scaffold and in the aniline moiety were reported. Preliminary cytotoxicity studies identified three derivatives, carrying dioxygenated rings fused on the quinazoline portion and the biphenylamino substituent as aniline portion, as the most effective compounds. Further investigations revealed that these compounds exhibited antiproliferative activity on a wide panel of human tumor cell lines through the inhibition of both receptor and nonreceptor TKs. Furthermore, the compound bearing the dioxolane nucleus was also able to inhibit in vivo tumor growth. Molecular modeling of these compounds into kinase domain suggested that the phenyl group allows favorable interaction energies with the target proteins: this feature is favored by fused dioxygenated ring at the 6,7 positions, whereas free rotating functions do not allow the correct placement of the molecule, thus impairing the inhibitory potency. Finally, the biphenylamino derivatives, at noncytotoxic concentrations, acted as antiangiogenic agents both in in vitro and in vivo assays.

Some new psoralen derivatives were synthesized and evaluated as inhibitors of NF-κB/DNA interaction, with the aim to investigate the structural determinants required to inhibit this interaction. Starting from molecular docking studies, several possible protein binding sites were proposed and several three-dimensional quantitative structure-activity relationship (3D-QSAR) models were built using the docked poses of 29 (the most active psoralen in the series) as templates for alignment of the inhibitors. The selected best model was validated through the prediction of the activity of 17 novel compounds. All the experimental data agreed with the computational experiments, supporting the reliability of the computational approach. The hypothesis about the interaction with NF-κB was also supported by surface plasmon resonance based assays using compound 29. All the collected data allowed the identification of compound 29 as a potential candidate for the development of pharmaceutical strategies against the inflammatory phenotype of cystic fibrosis.

A new tetracyclic derivative, difurocoumarin, was synthesized and studied in order to ascertain its possible use as a photochemotherapeutic agent alternative to psoralens. The compound proved able to photobind monofunctionally to DNA on irradiation with UV-A. A photocycloadduct with thymine was isolated and characterized spectroscopically.

Due to the increase in knowledge about cancer pathways, there is a growing interest in finding novel potential drugs. Quinazoline is one of the most widespread scaffolds amongst bioactive compounds. A number of patents and papers appear in the literature regarding the discovery and development of novel promising quinazoline compounds for cancer chemotherapy. Although there is a progressive decrease in the number of patents filed, there is an increasing number of biochemical targets for quinazoline compounds.This paper provides a comprehensive review of the quinazolines patented in 2007 - 2010 as potential anticancer agents. Information from articles published in international peer-reviewed journals was also included, to give a more exhaustive overview.From about 1995 to 2006, the anticancer quinazolines panorama has been dominated by the 4-anilinoquinazolines as tyrosine kinase inhibitors. The extensive researches conducted in this period could have caused the progressive reduction in the ability to file novel patents as shown in the 2007 - 2010 period. However, the growing knowledge of cancer-related pathways has recently highlighted some novel potential targets for therapy, with quinazolines receiving increasing attention. This is well demonstrated by the number of different targets of the patents considered in this review. The structural heterogeneity in the patented compounds makes it difficult to derive general pharmacophores and make comparisons among claimed compounds. On the other hand, the identification of multi-target compounds seems a reliable goal. Thus, it is reasonable that quinazoline compounds will be studied and developed for multi-target therapies.

Some photobiological properties of 2,6-dimethyl-9-methoxy-4H-pyrrolo [3,2,1-ij]quinolin-4-one (PQ) have been studied in comparison with 8-methoxypsoralen (8-MOP). In Ehrlich cells, PQ induced a moderate inhibition in DNA and RNA syntheses in the dark, which appeared to be more pronounced upon UVA irradiation. In contrast to 8-MOP, in the presence of UVA, PQ also affected protein synthesis. Likewise marked antiproliferative effects were also observed in the study of the clonal growth of CHO cells cultivated in vitro . Using alkaline elution and CHO cells, a moderate formation of single-strand breaks (SSBs) and of DNA-protein cross-links (DPCs) was observed by incubation in the dark; upon UVA irradiation the amount of both lesions increased greatly, whereas no inter-strand cross-links (ISCs) were formed. As expected, 8-MOP did not damage DNA in the dark, but induced SSBs, ISCs and DPCs in the presence of UVA. The induction of SSBs by both compounds seems to be directly related to a photochemical event rather than to incisions during DNA repair. As the induction of ISCs, and also the formation of DPCs by 8-MOP and UVA, appears to be based on a two-step reaction involving photo-bound 8-MOPDNA moieties. In contrast, the formation of DPCs by PQ and UVA seems to involve photosensitization by free PQ molecules connected with SSB and DPC formation rather than with a DNA photo-binding activity. The PQ activity observed in the dark could probably be ascribed to a moderate inhibition of topoisomerases.

The formation of C4-cycloadducts by photoreaction of eight methyl derivatives of 8-azapsoralen with DNA was studied. The main reaction involves the furan ring of the compounds and the 5,6 double bond of thymine giving cis-syn adducts, although a minor amount of a cycloadduct with cytosine was also isolated for 4,4'-dimethylazapsoralen. The role of the nitrogen atom appears to depend on the electronic effect, leading to a decrease in the reactivity of the pyrone ring. As with psoralens, the methyl groups increase both the lipophilicity and, with the exception of the 5,4',5'-trimethyl derivative, the photoreactivity. However, methyl groups do not appear to influence the chemistry of the photoaddition.

Nuclear Factor kappaB (NF-κB) plays a very important role in the control of gene expression and is deeply involved in several human pathologies. Accordingly, molecules targeting NF-κB dependent biological functions are considered of great interest. Virtual screening of furocoumarin libraries against NF-κB p50 allowed to rank compounds in respect to their expected ability to bind NF-κB and the identified compound might be considered for the development of analogs to be tested for biological activity on inhibition of NF-κB/DNA complex formation. The data reported in the present paper suggest that, following this approach, the best ranked compounds identified by virtual screening (a) strongly bind in silico to NF-κB and (b) efficiently inhibit the molecular interactions between 32P-labeled NF-κB double stranded DNA and p50 or p50/p65 complex. These data allowed to develop a novel lead of great interest for inhibiting NF-κB dependent biological functions. This novel molecule (compound 2), bearing a methyl group in the 9 position of the psoralen nucleus, exhibits high efficiency in inhibiting NF-κB/DNA interactions. In addition, we found that compound 2 is a potent inhibitor of IL-8 gene expression in TNF-α treated IB3-1 cystic fibrosis cells. Taken together, our data indicate that compound 2 might find an important place in the set of molecules of interest for the development of pharmaceutical strategies against the inflammatory phenotype of cystic fibrosis.

Quantitative Structure Activity Relationship (QSAR) is a well known cheminformatic tool for the discovery of novel biologically active compounds. However, when large and heterogeneous datasets are mined, it is not possible to derive a QSAR equation able to predict in a satisfactory manner the activity of the compounds. Thus, QSAR models are often inadequate for virtual screening purpose. Herein we present a novel approach to multitarget classification QSAR models, useful to assess the selectivity profile of the tyrosine kinases inhibitors. A descriptor-based clusterization process was employed, that allowed the generation of models with high accuracies and independent from the chemical classification of the compounds (i.e. from the scaffold type). The herein proposed methodology can lead to QSAR models useful for virtual screening processes.

The synthesis of new tetracyclic 4H-furo[3,2-g]pyrrolo[3,2,1-ij]quinolin-4-ones, 15–17, 1H,5H-benzofuro[5,6, 7-ij]quinolizin-5-ones, 18–20, and 3H,5H-benzofuro[5,6,7-ij]quinolizin-5-ones, 21–23 is described. The planarity of the structure of benzofuroquinolizin-5-one 18 was determined by X-ray single-crystal analysis.

The antiangiogenic effects of three novel anilinoquinazoline derivatives were studied with the aim to find new multi-kinase inhibitors as anticancer agents. The compounds are characterized by dioxolane, dioxane and dioxepine rings and bear the same aniline substituent in 4 position as Vandetanib, known antiangiogenic agent. The in vitro assays were carried out on human umbilical vein endothelial cells (HUVECs), whereas in vivo angiogenesis was evaluated by means of Matrigel plug assay. The results showed that these compounds exert, even though to different extents, antiangiogenic activity affecting the various step of the process that leads to the formation of new blood vessels. At high concentrations they induced antiproliferative effects, whereas at non-cytotoxic concentrations they inhibited cell migration and the formation of tubular structures in Matrigel. In in vitro assays the dioxolane derivative 1 was more effective than Vandetanib. Indeed, it inhibited the effects induced by exogenous VEGF and FGF-2 on both cell proliferation and morphogenesis, whereas Vandetanib was completely ineffective. Moreover, all the compounds, as Vandetanib, counteracted the FGF-2-induced increase in the hemoglobin content in the Matrigel plugs. Our results showed that all the three novel derivatives possess both in vitro and in vivo antiangiogenic activity, with compound 1 more effective than Vandetanib to inhibit in vitro angiogenesis induced by exogenous cytokines.

Tyrosine kinases constitute an eligible class of target for novel drug discovery. They resulted often overexpressed and/or deregulated in several cancer diseases. Thus, the development of novel tyrosine kinases inhibitors is of value, as well as the finding of novel cheminformatic tools for their design. Among the different ways to rationally design novel compounds, the Quantitative Structure-Activity Relationship (QSAR) plays a key role. The QSAR approach, in fact, allow the prediction of activity against a number of targets (multi-target QSAR), thus leading to models able to predict not only the activity of a compound, but also its selectivity versus a set of targets. Despite it is well known that tyrosine kinase inhibitors have to show multi-kinases inhibitory potency to be useful in anticancer therapy, only few multi-target computational tools have been developed to help medicinal chemists in the design of novel compounds. Herein we present the development of several multi-target classification QSAR (mtc-QSAR) models useful to assess the activity profile of the tyrosine kinases inhibitors.

A new convenient synthesis of pyrrolo[3,2,1-ij]quinolin-4-one derivatives is described. In this method, methyl-7-hydroxyquinoline-2-ones are the starting materials onto which the third pyrrolo ring is condensed directly, yielding dehydrogenated methyl-9-hydroxypyrrolo[3,2,1-ij]quinolin-4-ones

The synthesis of new tetracyclic 4H-furo[3,2-g]pyrrolo[3,2,1-ij]quinolin-4-ones, 15–17, 1H,5H-benzofuro[5,6, 7-ij]quinolizin-5-ones, 18–20, and 3H,5H-benzofuro[5,6,7-ij]quinolizin-5-ones, 21–23 is described. The planarity of the structure of benzofuroquinolizin-5-one 18 was determined by X-ray single-crystal analysis.