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The phytochemical investigation of the methanol extract of the seeds of Magydaris pastinacea afforded two undescribed benzofuran glycosides, furomagydarins A-B ( 1 , 2 ), together with three known coumarins. The structures of the new isolates were elucidated after extensive 1D and 2D NMR experiments as well as HR MS. Compound 1 was able to inhibit the COX-2 expression in RAW264.7 macrophages exposed to lipopolysaccharide, a pro-inflammatory stimulus. RT-qPCR and luciferase reporter assays suggested that compound 1 reduces COX-2 expression at the transcriptional level. Further studies highlighted the capability of compound 1 to suppress the LPS-induced p38MAPK, JNK, and C/EBPβ phosphorylation, leading to COX-2 down-regulation in RAW264.7 macrophages.

A small library of 1-(4-nitrophenyl)-3-arylprop-2-en-1-one derivatives was synthesized to identify new human monoamine oxidase B selective inhibitors. Their inhibitory activity toward MAO-A and MAO-B isoforms was evaluated to determine their potency and selectivity. All newly synthesized compounds were nanomolar inhibitors of the B isoform with IC 50 concentrations ranging from 120 to 2.2 nM. Conversely, their activity toward the A isozyme was only observed at micromolar concentrations. Our results bear out the hypothesis that the 1,3-diarylpropenone scaffold could represent a valuable starting point for designing efficient and selective MAO-B inhibitors., Competing Interests: The authors declare no competing financial interest., (© 2024 American Chemical Society.)

Human Carbonic Anhydrases (hCA) are enzymes that contribute to cancer's development and progression. Isoforms IX and XII have been identified as potential anticancer targets, and, more specifically, hCA IX is overexpressed in hypoxic tumor cells, where it plays an important role in reprogramming the metabolism. With the aim to find new inhibitors towards IX and XII isoforms, the hybridization of the privileged scaffolds isatin, dihydrothiazole, and benzenesulfonamide was investigated in order to explore how it may affect the activity and selectivity of the hCA isoforms. In this respect, a series of isatin thiazolidinone hybrids have been designed and synthesized and their biological activity and selectivity on hCA I, hCA II, hCA IX, and hCA XII explored. The new compounds exhibited promising inhibitory activity results on isoforms IX and XII in the nanomolar range, which has highlighted the importance of substituents in the isatin ring and in position 3 and 5 of thiazolidinone. In particular, compound 5g was the most active toward hCA IX, while 5f was the most potent inhibitor of hCA XII within the series. When both potency and selectivity were considered, compound 5f appeared as one of the most promising. Additionally, our investigations were supported by molecular docking experiments, which have highlighted the putative binding poses of the most promising compound.

Chemotherapeutic agents have remained the first-line treatment option for advanced-stage cancers when surgery or radiation therapy is not viable. Human carbonic anhydrase (hCA) isoforms IX and XII have been validated as anticancer targets. In particular, hCA IX is overexpressed in several solid tumor cells. As a result, selective isoform inhibitors with high potency and low toxicity are sought after. Pursuing our investigation on new scaffolds as hCA-selective inhibitors, a new series of isatin thiazolidinone hybrids has been designed and synthesized. Their biological activity and selectivity toward hCA I, hCA II, hCA IX, and hCA XII were investigated. The results revealed an inhibitory activity in the nanomolar range on carbonic anhydrases IX and XII, and the nature of substitution in positions 3 and 5 of thiazolidinone appears to be crucial for the compounds' selectivity. Docking experiments have been applied to predict the binding mode of these new, promising derivatives., Competing Interests: The authors declare no competing financial interest., (© 2024 American Chemical Society.)

Tumour associated carbonic anhydrases (CAs) IX and XII have been recognised as potential targets for the treatment of hypoxic tumours. Therefore, considering the high pharmacological potential of the chromene scaffold as selective ligand of the IX and XII isoforms, two libraries of compounds, namely 2H-chromene and 7H-furo-chromene derivatives, with diverse substitution patterns were designed and synthesised. The structure of the newly synthesised compounds was characterised and their inhibitory potency and selectivity towards human CA off target isoforms I, II and cancer-associated CA isoforms IX and XII were evaluated. Most of the compounds inhibit CA isoforms IX and XII with no activity against the I and II isozymes. Thus, while the potency was influenced by the substitution pattern along the chromene scaffold, the selectivity was conserved along the series, confirming the high potential of both 2H-chromene and 7H-furo-chromene scaffolds for the design of isozyme selective inhibitors.

In the effort to identify and develop new HIV-1 inhibitors endowed with innovative mechanisms, we focused our attention on the possibility to target more than one viral encoded enzymatic function with a single molecule. In this respect, we have previously identified by virtual screening a new indolinone-based scaffold for dual allosteric inhibitors targeting both reverse transcriptase-associated functions: polymerase and RNase H. Pursuing with the structural optimization of these dual inhibitors, we synthesized a series of 35 new 3-[2-(4-aryl-1,3-thiazol-2-ylidene)hydrazin-1-ylidene]1-indol-2-one and 3-[3-methyl-4-arylthiazol-2-ylidene)hydrazine-1-ylidene)indolin-2-one derivatives, which maintain their dual inhibitory activity in the low micromolar range. Interestingly, compounds 1a, 3a, 10a, and 9b are able to block HIV-1 replication with EC 50 < 20 µM. Mechanism of action studies showed that such compounds could block HIV-1 integrase. In particular, compound 10a is the most promising for further multitarget compound development., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Corona, Meleddu, Delelis, Subra, Cottiglia, Esposito, Distinto, Maccioni and Tramontano.)

Ebola virus (EBOV) is a highly infectious and lethal pathogen responsible for sporadic self-limiting clusters of Ebola virus disease (EVD) in Central Africa capable of reaching epidemic status. 100% protection from lethal EBOV-Zaire in Balb/c mice was achieved by rintatolimod (Ampligen) at the well tolerated human clinical dose of 6 mg/kg. The data indicate that the mechanism of action is rintatolimod's dual ability to act as both a competitive decoy for the IID domain of VP35 blocking viral dsRNA sequestration and as a pathogen-associated molecular pattern (PAMP) restricted agonist for direct TLR3 activation but lacking RIG-1-like cytosolic helicase agonist properties. These data show promise for rintatolimod as a prophylactic therapy against human Ebola outbreaks., Competing Interests: Declaration of competing interest Authors AC, SD, GLD, AP, and EZ declare no conflict of interest. DS is the Medical Director of AIM ImmnunoTech. WMM is a member of Board of Directors for AIM ImmunoTech. The funders had no role in the design of the study, in the collection, analyses, or interpretation of data, in the writing of the manuscript, or in the decision to publish the results., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Ebola virus (EBOV) is one of the deadliest infective agents whose lethality is linked to the ability to efficiently bypass the host's innate antiviral response. EBOV multifunctional protein VP35 plays a major role in viral replication both as polymerase cofactor and interferon (IFN) antagonist. By hiding the non-self 5'-ppp dsRNA from the cellular receptor RIG-I, VP35 prevents its activation and inhibits IFN-β production. Blocking VP35-dsRNA interaction and IFN-β suppression is a validated drug target. We screened a library of natural extracts and found that cynarin inhibits dsRNA-VP35 binding with an IC 50 value of 8.5 μM. It reverts the EBOV VP35 inhibition of IFN-β production, while it does not induce IFN production by itself. Docking experiments suggest that the molecule can bind on the end-capping pocket of VP35 C-terminal Interferon Inhibitory domain (IID), and differential scanning fluorimetry confirmed that cynarin interacts with VP35-IID with a K D of 12 μM. Cynarin was further tested in an EBOV minigenome assay but did not inhibit VP35 polymerase cofactor activity. When evaluated during challenge of IFN-susceptible A549 cells with EBOV isolate derived from the 2014 West African outbreak, cynarin was able to inhibit viral replication with an EC 50 value of 9.1 μM, showing no significant cytotoxicity. Our findings show that cynarin blocks EBOV replication by acting directly on VP35 and subverting its IFN antagonism function but not cofactor function, and as such identify the first EBOV inhibitor with this mode of action., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Bioassay-guided fractionation of the ethyl acetate extract from Teucrium flavum subsp. glaucum , endowed with inhibitory activity towards the HIV-1 reverse transcriptase-associated RNase H function, led to the isolation of salvigenin ( 1 ), cirsimaritin ( 2 ) and cirsiliol ( 3 ) along with the neo -clerodanes teuflavin ( 4 ) and teuflavoside ( 5 ). Acid hydrolysis of the inactive teuflavoside provided three undescribed neo -clerodanes, flavuglaucins A-C ( 7-9 ) and one known neo -clerodane ( 10 ). Among all neo -clerodanes, flavuglaucin B showed the highest inhibitory activity towards RNase H function with a IC 50 value of 9.1 μM. Molecular modelling and site-directed mutagenesis analysis suggested that flavuglaucin B binds into an allosteric pocket close to RNase H catalytic site. This is the first report of clerodane diterpenoids endowed with anti-reverse transcriptase activity. Neo -clerodanes represent a valid scaffold for the development of a new class of HIV-1 RNase H inhibitors.

A small library of coumarin and their psoralen analogues EMAC10157a-b-d-g and EMAC10160a-b-d-g has been designed and synthesised to investigate the effect of structural modifications on their inhibition ability and selectivity profile towards carbonic anhydrase isoforms I, II, IX, and XII. None of the new compounds exhibited activity towards hCA I and II isozymes. Conversely, both coumarin and psoralen derivatives were active against tumour associated isoforms IX and XII in the low micromolar or nanomolar range of concentration. These data further corroborate our previous findings on analogous derivatives, confirming that both coumarins and psoralens are interesting scaffolds for the design of isozyme selective hCA inhibitors.

Current therapeutic protocols for the treatment of HIV infection consist of the combination of diverse anti-retroviral drugs in order to reduce the selection of resistant mutants and to allow for the use of lower doses of each single agent to reduce toxicity. However, avoiding drugs interactions and patient compliance are issues not fully accomplished so far. Pursuing on our investigation on potential anti HIV multi-target agents we have designed and synthesized a small library of biphenylhydrazo 4-arylthiazoles derivatives and evaluated to investigate the ability of the new derivatives to simultaneously inhibit both associated functions of HIV reverse transcriptase. All compounds were active towards the two functions, although at different concentrations. The substitution pattern on the biphenyl moiety appears relevant to determine the activity. In particular, compound 2-{3-[(2-{4-[4-(hydroxynitroso)phenyl]-1,3-thiazol-2-yl} hydrazin-1-ylidene) methyl]-4-methoxyphenyl} benzamide bromide ( EMAC2063 ) was the most potent towards RNaseH (IC 50 = 4.5 mM)- and RDDP (IC 50 = 8.0 mM) HIV RT-associated functions.

Quorum sensing (QS) plays an essential role in the production of virulence factors, in biofilm formation and antimicrobial resistance. Consequently, inhibiting QS is being considered a promising target for antipathogenic/anti-virulence therapies. This study aims to screen 2-nitrovinylfuran derivatives structurally related to Furvina (a broad-spectrum antibiotic already used for therapeutic purposes) for their effects on QS and in biofilm prevention/control. Furvina and four 2-nitrovinylfuran derivatives (compounds 1 - 4 ) were tested to assess the ability to interfere with QS of Staphylococcus aureus using bioreporter strains ( S. aureus ALC1742 and ALC1743). The activity of Furvina and the most promising quorum-sensing inhibitor (QSI) was evaluated in biofilm prevention and in biofilm control (combined with fusidic acid). The biofilms were further characterized in terms of biofilm mass, viability and membrane integrity. Compound 2 caused the most significant QS inhibition with reductions between 60% and 80%. Molecular docking simulations indicate that this compound interacts preferentially with the protein hydrophobic cleft in the LytTR domain of AgrA pocket. Metabolic inactivations of 40% for S. aureus ALC1742 and 20% for S. aureus ALC1743 were reached. A 24 h-old biofilm formed in the presence of the QSI increased the metabolic inactivation by fusidic acid to 80%, for both strains. The overall results highlight the effects of compound 2 as well as the potential of combining QSI with in-use antibiotics for the management of skin and soft tissues infections.

In an in vitro screening for human carbonic anhydrase (hCA) inhibiting agents from higher plants, the petroleum ether and ethyl acetate extracts of Magydaris pastinacea seeds selectively inhibited hCA IX and hCA XII isoforms. The phytochemical investigation of the extracts led to the isolation of ten linear furocoumarins ( 1 - 10 ), four simple coumarins ( 12 - 15 ) and a new angular dihydrofurocoumarin ( 11 ). The structures of the isolated compounds were elucidated based on 1 D and 2 D NMR, MS, and ECD data analysis. All isolated compounds were inactive towards the ubiquitous cytosolic isoform hCA I and II ( K i  > 10,000 nM) while they were significantly active against the tumour-associated isoforms hCA IX and XII. Umbelliprenin was the most potent coumarin inhibiting hCA XII isoform with a K i of 5.7 nM. The cytotoxicity of the most interesting compounds on HeLa cancer cells was also investigated.

HIV-1 infection requires life-long treatment and with 2.1 million new infections/year, faces the challenge of an increased rate of transmitted drug-resistant mutations. Therefore, a constant and timely effort is needed to identify new HIV-1 inhibitors active against drug-resistant variants. The ribonuclease H (RNase H) activity of HIV-1 reverse transcriptase (RT) is a very promising target, but to date, still lacks an efficient inhibitor. Here, we characterize the mode of action of N' -(2-hydroxy-benzylidene)-3,4,5-trihydroxybenzoylhydrazone (compound 13 ), an N-acylhydrazone derivative that inhibited viral replication (EC 50 = 10 µM), while retaining full potency against the NNRTI-resistant double mutant K103N-Y181C virus. Time-of-addition and biochemical assays showed that compound 13 targeted the reverse-transcription step in cell-based assays and inhibited the RT-associated RNase H function, being >20-fold less potent against the RT polymerase activity. Docking calculations revealed that compound 13 binds within the RNase H domain in a position different from other selective RNase H inhibitors; site-directed mutagenesis studies revealed interactions with conserved amino acid within the RNase H domain, suggesting that compound 13 can be taken as starting point to generate a new series of more potent RNase H selective inhibitors active against circulating drug-resistant variants.

Ebola virus (EBOV) is among the most devastating pathogens causing fatal hemorrhagic fever in humans. The epidemics from 2013 to 2016 resulted in more than 11,000 deaths, and another outbreak is currently ongoing. Since there is no FDA-approved drug so far to fight EBOV infection, there is an urgent need to focus on drug discovery. Considering the tight correlation between the high EBOV virulence and its ability to suppress the type I interferon (IFN-I) system, identifying molecules targeting viral protein VP24, one of the main virulence determinants blocking the IFN response, is a promising novel anti-EBOV therapy approach. Hence, in the effort to find novel EBOV inhibitors, a screening of a small set of flavonoids was performed; it showed that quercetin and wogonin can suppress the VP24 effect on IFN-I signaling inhibition. The mechanism of action of the most active compound, quercetin, showing a half-maximal inhibitory concentration (IC 50 ) of 7.4 μM, was characterized to significantly restore the IFN-I signaling cascade, blocked by VP24, by directly interfering with the VP24 binding to karyopherin-α and thus restoring P-STAT1 nuclear transport and IFN gene transcription. Quercetin significantly blocked viral infection, specifically targeting EBOV VP24 anti-IFN-I function. Overall, quercetin is the first identified inhibitor of the EBOV VP24 anti-IFN function, representing a molecule interacting with a viral binding site that is very promising for further drug development aiming to block EBOV infection at the early steps., (Copyright © 2020 American Society for Microbiology.)

In the present study we investigated the structure-activity relationships of a new series of 4-[(3-ethyl-4-aryl-2,3-dihydro-1,3-thiazol-2-ylidene)amino]benzene-1-sulfonamides ( EMAC10101a - m ). All synthesized compounds, with the exception of compound EMAC10101k , preferentially inhibit off-target hCA II isoform. Within the series, compound EMAC10101d , bearing a 2,4-dichorophenyl substituent in position 4 of the dihydrothiazole ring, was the most potent and selective toward hCA II with an inhibitory activity in the low nanomolar range., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)

The paper focussed on a step-by-step structural modification of a cycloheptathiophene-3-carboxamide derivative recently identified by us as reverse transcriptase (RT)-associated ribonuclease H (RNase H) inhibitor. In particular, its conversion to a 2-aryl-cycloheptathienoozaxinone derivative and the successive thorough exploration of both 2-aromatic and cycloheptathieno moieties led to identify oxazinone-based compounds as new anti-RNase H chemotypes. The presence of the catechol moiety at the C-2 position of the scaffold emerged as critical to achieve potent anti-RNase H activity, which also encompassed anti-RNA dependent DNA polymerase (RDDP) activity for the tricyclic derivatives. Benzothienooxazinone derivative 22 resulted the most potent dual inhibitor exhibiting IC 50 s of 0.53 and 2.90 μM against the RNase H and RDDP functions. Mutagenesis and docking studies suggested that compound 22 binds two allosteric pockets within the RT, one located between the RNase H active site and the primer grip region and the other close to the DNA polymerase catalytic centre.

A library of 4-[(3-methyl-4-aryl-2,3-dihydro-1,3-thiazol-2-ylidene)amino]benzene-1-sulphonamides ( EMAC8002a-m ) was designed and synthesised to evaluate the effect of substituents in the positions 3 and 4 of the dihydrothiazole ring on the inhibitory potency and selectivity toward human carbonic anhydrase isoforms I, II, IX, and XII. Most of the new compounds preferentially inhibit the isoforms II and XII. Both electronic and steric features on the aryl substituent in the position 4 of the dihydrothiazole ring concur to determine the overall biological activity of these new derivatives.

Nine compounds, including two undescribed withanolides, withasomniferolides A and B (1 and 2), three known withanolides (3-5), a ferulic acid dimeric ester (6), and an inseparable mixture of three long alkyl chain ferulic acid esters (7-9), were isolated from a GABA A receptor positive activator methanol extract of the roots of Withania somnifera. The structures of the isolated compounds were elucidated based on NMR, MS, and ECD data analysis. In order to bioassay the single ferulic acid derivatives, compounds 6-9 were also synthesized. The most active compound, docosanyl ferulate (9), was able to enhance the GABA A receptor inhibitory postsynaptic currents with an IC 50 value of 7.9 μM. These results, by showing an ability to modulate the GABA A receptor function, cast fresh light on the biological activities of the secondary metabolites of W. somnifera roots.

In this work we report a parallel application of both docking- and shape-based virtual screening (VS) methods, followed by Molecular Dynamics simulations (MDs), for discovering new compounds able to inhibit the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) RNA-dependent DNA polymerase activity. Specifically, we screened more than 143000 natural compounds commercially available in the ZINC database against the best five RT crystallographic models, taking into account the five approved NNRTIs as query compounds. As a result, 20 hit molecules were selected and tested on biochemical assays for the inhibition of the RNA dependent DNA polymerase RT function and, among them, an indoline pyrrolidine (hit1), an indonyl piperazine (hit2) and an indolyl indolinone (hit3) derivatives were identified as novel non-nucleoside RT inhibitors in the low micromolar range., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

A series of isatin-dihydropyrazole hybrids have been synthesized in order to assess their potential as anticancer agents. In particular, 12 compounds were evaluated for their antiproliferative activity toward A549, IGR39, U87, MDA-MB-231, MCF-7, BT474, BxPC-3, SKOV-3, and H1299 cell lines, and human foreskin fibroblasts. Four compounds exhibited interesting antiproliferative activity and were further examined to determine their EC 50 values toward a panel of selected tumor cell lines. The best compounds were then investigated for their induced mechanism of cell death. Preliminary structure-activity relationship indicates that the presence of a substituent such as a chlorine atom or a methyl moiety in position 5 of the isatin nucleus is beneficial for the antitumor activity. EMAC4001 proved the most promising compound within the studied series with EC 50 values ranging from 0.01 to 0.38 μM., Competing Interests: The authors declare no competing financial interest.

Ebola virus (EBOV) is a filovirus that causes a severe and rapidly progressing hemorrhagic syndrome; a recent epidemic illustrated the urgent need for novel therapeutic agents because no drugs have been approved for treatment of Ebola virus. A key contribution to the high lethality observed during EBOV outbreaks comes from viral evasion of the host antiviral innate immune response in which viral protein VP35 plays a crucial role, blocking interferon type I production, first by masking the viral double-stranded RNA (dsRNA) and preventing its detection by the pattern recognition receptor RIG-I. Aiming to identify inhibitors of the interaction of VP35 with the viral dsRNA, counteracting the VP35 viral innate immune evasion, we established a new methodology for high-yield recombinant VP35 (rVP35) expression and purification and a novel and robust fluorescence-based rVP35-RNA interaction assay ( Z' factor of 0.69). Taking advantage of such newly established methods, we screened a small library of Sardinian natural extracts, identifying Limonium morisianum as the most potent inhibitor extract. A bioguided fractionation led to the identification of myricetin as the component that can inhibit rVP35-dsRNA interaction with an IC 50 value of 2.7 μM. Molecular docking studies showed that myricetin interacts with the highly conserved region of the VP35 RNA binding domain, laying the basis for further structural optimization of potent inhibitors of VP35-dsRNA interaction.

A novel series of of 4-[(3-phenyl-4-aryl-2,3-dihydro-1,3-thiazol-2-ylidene)amino]benzene-1-sulfonamides ( EMAC10111a-g ) was synthesized and assayed toward both human carbonic anhydrase isozymes I, II, IX, and XII and cyclooxygenase isoforms. The majority of these derivatives preferentially inhibit hCA isoforms II and XII and hCOX-2 isozyme, indicating that 2,3,4-trisubstituted 2,3-dihydrothiazoles are a promising scaffold for the inhibition of hCA isozymes and of hCOX-2 enzyme. The nature of the substituent at the dihydrothiazole ring position 4 influenced the activity and selectivity toward both enzyme families. EMAC10111g resulted as the best performing compound toward both enzyme families and exhibited preferential activity toward hCA XII and hCOX-2 isozymes., Competing Interests: The authors declare no competing financial interest.

A small library of psoralen carboxylic acids and their corresponding benzenesulfonamide derivatives were designed and synthesized to evaluate their activity and selectivity toward tumor associated human carbonic anhydrase (hCA) isoforms IX and XII. Both psoralen acids and sulfonamides exhibited potent inhibition of IX and XII isozymes in the nanomolar concentration range. However, psoralen acids resulted as the most selective in comparison with the corresponding benzenesulfonamide derivatives. Our data indicate that the psoralen scaffold is a promising starting point for the design of highly selective tumor associated hCA inhibitors., Competing Interests: The authors declare no competing financial interest.

A series of 3-3-{2-[2-3-methyl-4-phenyl-2,3-dihydro-1,3-thiazol-2-ylidene]hydrazin-1-ylidene-2,3-dihydro-1H-indol-2-one derivatives has been designed and synthesized to study their activity on both HIV-1 (Human Immunodeficiency Virus type 1) RT (Reverse Transcriptase) associated functions. These derivatives are analogs of previously reported series whose biological activity and mode of action have been investigated. In this work we investigated the influence of the introduction of a methyl group in the position 3 of the dihydrothiazole ring and of a chlorine atom in the position 5 of the isatin nucleus. The new synthesized compounds are active towards both DNA polymerase and ribonuclease H in the µM range. The nature of the aromatic group in the position 4 of the thiazole was relevant in determining the biological activity.

3,5-Diaryl-4,5-dihydroisoxazoles were synthesized and evaluated as monoamine oxidase (MAO) enzyme inhibitors and iron chelators. All compounds exhibited selective inhibitory activity towards the B isoform of MAO in the nanomolar concentration range. The best performing compound was preliminarily evaluated for its ability to bind iron II and III cations, indicating that neither iron II nor iron III is coordinated. The best compounds racemic mixtures were separated and single enantiomers inhibitory activity evaluated. Furthermore, none of the synthesised compounds exhibited activity towards MAO A. Overall, these data support our hypothesis that 3,5-diaryl-4,5-dihydroisoxazoles are promising scaffolds for the design of neuroprotective agents.

The isatin scaffold is the constitutive fragment of several natural and synthetic bioactive molecules. Albeit several benzene sulphonamide-based carbonic anhydrase inhibitors (CAIs) have been reported, only recently isatin benzene sulphonamides have been studied and proposed as CAIs. In this study we have designed, synthesised, and evaluated the biological activity of a series of differently substituted isatin-based benzene sulphonamides which have been designed for the inhibition of carbonic anhydrase isoforms. The activity of all the synthesised compounds was evaluated towards human carbonic anhydrase I, II, IX, and XII isozymes. Our results indicate that the nature and position of substituents on the isatin ring can modulate both activity and isozyme selectivity.

The dichloromethane extract of the leaves of Bupleurum fruticosum was found to inhibit the replication of human rhinovirus (HRV) serotypes 14 and 39. Bioassay-guided fractionation led to the isolation of seven phenylpropenol derivatives (3-9), two polyacetylenes (1 and 2), and one monoterpene (10). Compounds 1 and 10 were identified as previously undescribed secondary metabolites after extensive 1D and 2D NMR experiments as well as high-resolution mass spectrometry. Compounds 2, 4, and 5 showed a selective inhibition of viral replication against HRV39 serotype, with 2 and 4 being the most active, with EC 50 values of 1.8 ± 0.02 and 2.4 ± 0.04 μM. Mechanism of action studies indicated that 4 behaves not only as a capsid binder, interfering with the early phases of virus replication, but also as a late-phase replication inhibitor. Docking experiments were performed to confirm the ability of the antiviral phenylpropenoids to selectively fit into the hydrophobic pocket of VP1-HRV39.

In the continuous effort to identify new HIV-1 inhibitors endowed with innovative mechanisms, the dual inhibition of different viral functions would provide a significant advantage against drug-resistant variants. The HIV-1 reverse transcriptase (RT)-associated ribonuclease H (RNase H) is the only viral-encoded enzymatic activity that still lacks an efficient inhibitor. We synthesized a library of 3,5-diamino-N-aryl-1H-pyrazole-4-carbothioamide and 4-amino-5-benzoyl-N-phenyl-2-(substituted-amino)-1H-pyrrole-3-carbothioamide derivatives and tested them against RNase H activity. We identified the pyrazolecarbothioamide derivative A15, able to inhibit viral replication and both RNase H and RNA-dependent DNA polymerase (RDDP) RT-associated activities in the low micromolar range. Docking simulations hypothesized its binding to two RT pockets. Site-directed mutagenesis experiments showed that, with respect to wt RT, V108A substitution strongly reduced A15 IC50 values (12.6-fold for RNase H inhibition and 4.7-fold for RDDP), while substitution A502F caused a 9.0-fold increase in its IC50 value for RNase H, not affecting the RDDP inhibition, reinforcing the hypothesis of a dual-site inhibition. Moreover, A15 retained good inhibition potency against three non-nucleoside RT inhibitor (NNRTI)-resistant enzymes, confirming a mode of action unrelated to NNRTIs and suggesting its potential as a lead compound for development of new HIV-1 RT dual inhibitors active against drug-resistant viruses., (© FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Despite the availability of several anti-retrovirals, there is still an urgent need for developing novel therapeutic strategies and finding new drugs against underexplored HIV-1 targets. Among them, there are the HIV-1 reverse transcriptase (RT)-associated ribonuclease H (RNase H) function and the cellular α-glucosidase, involved in the control mechanisms of N-linked glycoproteins formation in the endoplasmic reticulum. It is known that many natural compounds, such as pentacyclic triterpenes, are a promising class of HIV-1 inhibitors. Hence, here we tested the pentacyclic triterpene Lupeol, showing that it inhibits the HIV-1 RT-associated RNase H function. We then performed combination studies of Lupeol and the active site RNase H inhibitor RDS1759, and blind docking calculations, demonstrating that Lupeol binds to an HIV-1 RT allosteric pocket. On the bases of these results and searching for potential multitarget active drug supplement, we also investigated the anti-HIV-1 activity of Hemidesmus indicus, an Ayurveda medicinal plant containing Lupeol. Results supported the potential of this plant as a valuable multitarget active drug source. In fact, by virtue of its numerous active metabolites, H. indicus was able to inhibit not only the RT-associated RNase H function, but also the HIV-1 RT-associated RNA-dependent DNA polymerase activity and the cellular α-glucosidase., (© FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

A series of N -acylbenzenesulfonamide dihydro-1,3,4-oxadiazole hybrids ( EMAC8000a-m ) was designed and synthesized with the aim to target tumor associated carbonic anhydrase (hCA) isoforms IX and XII. Most of the compounds were selective inhibitors of the tumor associated hCA XII. Moreover, resolution of EMAC8000d racemic mixture led to the isolation of the levorotatory eutomer exhibiting an increase of hCA XII inhibition potency and selectivity with respect to hCA II. Computational studies corroborated these data. Overall our data indicate that both substitution pattern and stereochemistry of dihydro-1,3,4-oxadiazole could be considered as key factors to determine activity and selectivity toward hCA isozymes. These results can provide further indication for the design and optimization of selective hCA inhibitors.

Background: Recent findings demonstrated that, in mammalian cells, telomere DNA (Tel) is transcribed into telomeric repeat-containing RNA (TERRA), which is involved in fundamental biological processes, thus representing a promising anticancer target. For this reason, the discovery of dual (as well as selective) Tel/TERRA G-quadruplex (G4) binders could represent an innovative strategy to enhance telomerase inhibition., Methods: Initially, docking simulations of known Tel and TERRA active ligands were performed on the 3D coordinates of bimolecular G4 Tel DNA (Tel 2 ) and TERRA (TERRA 2 ). Structure-based pharmacophore models were generated on the best complexes and employed for the virtual screening of ~257,000 natural compounds. The 20 best candidates were submitted to biophysical assays, which included circular dichroism and mass spectrometry at different K + concentrations., Results: Three hits were here identified and characterized by biophysical assays. Compound 7 acts as dual Tel 2 /TERRA 2 G4-ligand at physiological KCl concentration, while hits 15 and 17 show preferential thermal stabilization for Tel 2 DNA. The different molecular recognition against the two targets was also discussed., Conclusions: Our successful results pave the way to further lead optimization to achieve both increased selectivity and stabilizing effect against TERRA and Tel DNA G4s., General Significance: The current study combines for the first time molecular modelling and biophysical assays applied to bimolecular DNA and RNA G4s, leading to the identification of innovative ligand chemical scaffolds with a promising anticancer profile. This article is part of a Special Issue entitled "G-quadruplex" Guest Editor: Dr. Concetta Giancola and Dr. Daniela Montesarchio., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Using an HIV-1 Reverse Transcriptase (RT)-associated RNase H inhibition assay as lead, bioguided fractionation of the dichloromethane extract of the Ocimum sanctum leaves led to the isolation of five triterpenes (1-5) along with three 3-methoxy-4-hydroxy phenyl derivatives (6-8). The structure of this isolates were determined by 1D and 2D NMR experiments as well as ESI-MS. Tetradecyl ferulate (8) showed an interesting RNase H IC 50 value of 12.4 μM and due to the synthetic accessibility of this secondary metabolite, a structure-activity relationship study was carried out. A series of esters and amides of ferulic and caffeic acids were synthesized and, among all, the most active was N-oleylcaffeamide displaying a strong inhibitory activity towards both RT-associated functions, ribonuclease H and DNA polymerase. Molecular modeling studies together with Yonetani-Theorell analysis, demonstrated that N-oleylcaffeamide is able to bind both two allosteric site located one close to the NNRTI binding pocket and the other close to RNase H catalytic site., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Cyclohexyliden- and 2-methylcyclohexyliden-hydrazo-4-arylthiazoles were synthesized and tested as antifungal agents. All compounds exhibited minimal inhibitory concentration (MIC) values comparable with those of fluconazole (FLC). Moreover, some compounds showed fungicidal activity at low concentration. Worth noting five out of nine compounds were active towards Candida albicans 25 FLC resistant isolated from clinical specimens. The cellular toxicity was evaluated and none of the compounds is toxic at the MIC. On the basis of our data we can conclude that these derivatives are promising agents for the treatment of resistant C. albicans.

Background: Despite the availability of effective antiretroviral therapies, drugs for HIV-1 treatment with new mode of action are still needed. An innovative approach is aimed to identify dual HIV-1 inhibitors, small molecules that can inhibit two viral functions at the same time. Rhubarb, originated from Rheum palmatum L. and Rheum officinale Baill., is one of the earliest and most commonly used medicinal plants in Traditional Chinese Medicine (TCM) practice. We wanted to explore TCM for the identification of new chemical scaffolds with dual action abilities against HIV-1., Methods: R. palmatum L. and R. officinale Baill. extracts along with their main single isolated constituents anthraquinone derivatives were tested on both HIV-1 Reverse Transcriptase (RT)-associated DNA Polymerase (RDDP) and Ribonuclease H (RNase H) activities in biochemical assays. Active compounds were then assayed for their effects on HIV-1 mutated RTs, integrase (IN) and viral replication., Results: Both R. palmatum L. and R. officinale Baill. extracts inhibited the HIV-1 RT-associated RNase H activity. Among the isolated constituents, Sennoside A and B were effective on both RDDP and RNase H RT-associated functions in biochemical assays. Sennoside A was less potent when tested on K103N, Y181C, Y188L, N474A and Q475A mutated RTs, suggesting the involvement of two RT binding sites for its antiviral activity. Sennoside A affected also HIV-1 IN activity in vitro and HIV-1 replication in cell-based assays. Viral DNA production and time of addition studies showed that Sennoside A targets the HIV-1 reverse transcription process., Conclusion: Sennoside A is a new scaffold for the development of HIV-1 dual RT inhibitors., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

It is well known that G-quadruplexes are targets of great interest for their roles in crucial biological processes, such as aging and cancer. Hence, a promising strategy for anticancer drug therapy is the stabilization of these structures by small molecules. We report a high-throughput in silico screening of commercial libraries from several different vendors by means of a combined structure-based pharmacophore model approach followed by docking simulations. The compounds selected by the virtual screening procedure were then tested for their ability to interact with human telomeric G-quadruplex folding by circular dichroism, fluorescence spectroscopy, and fluorescence intercalator displacement. Our approach resulted in the identification of a 13-[(dimethylamino)methyl]-12-hydroxy-8H-benzo[c]indolo[3,2,1-ij][1,5]naphthyridin-8-one derivative as a novel promising stabilizer of G-quadruplex structures within the human telomeric and the c-myc promoter sequences., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)