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Twenty-six 9-aminoacridine derivatives were evaluated in cell-based assays for cytotoxicity and antiviral activity against a panel of 10 RNA and DNA viruses. While seven compounds (9, 10, 14, 19, 21, 22, 24) did not affect any virus and two (6, 11) were moderately active against CVB-5 or Reo-1, 17 compounds exhibited a marked specific activity against BVDV, prototype of pestiviruses which are responsible for severe diseases of livestock. Most anti-BVDV agents showed EC(50) values in the range 0.1-8 μM, thus comparing favorably with the reference drugs ribavirine and NM 108. Some compounds, particularly those bearing a quinolizidinylalkyl side chain, displayed pronounced cytotoxicity. Further studies are warranted in order to achieve still better anti-BVDV agents, and to explore the potential antiproliferative activity of this kind of compounds., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Unsymmetrical methylene derivatives 5 were prepared following a known method, by reaction of the Mannich bases of 2-naphthols 4 with indoles. All synthesized compounds were tested against a wide panel of viruses, since previous work showed that Mannich bases on 7-hydroxycoumarin 1 and unsymmetrical methylene derivatives 2 were endowed with some antiviral activities. The symmetrical Mannich bases 4 were completely inactive, whereas the unsymmetrical methylene derivatives 5, although possessing a certain degree of toxicity, showed a significant activity against RSV. Some of compounds 5 showed a moderate antiviral activity against HIV-1, BVDV, YFV and CVB-2. The lack of activity of Mannich bases 4 demonstrates the crucial importance for antiviral activity of coumarin moiety present in Mannich bases 1.

Life-threatening RNA viruses emerge regularly, and often in an unpredictable manner. Yet, the very few drugs available against known RNA viruses have sometimes required decades of research for development. Can we generate preparedness for outbreaks of the, as yet, unknown viruses? The VIZIER (VIral enZymes InvolvEd in Replication) (http://www.vizier-europe.org/) project has been set-up to develop the scientific foundations for countering this challenge to society. VIZIER studies the most conserved viral enzymes (that of the replication machinery, or replicases) that constitute attractive targets for drug-design. The aim of VIZIER is to determine as many replicase crystal structures as possible from a carefully selected list of viruses in order to comprehensively cover the diversity of the RNA virus universe, and generate critical knowledge that could be efficiently utilized to jump-start research on any emerging RNA virus. VIZIER is a multidisciplinary project involving (i) bioinformatics to define functional domains, (ii) viral genomics to increase the number of characterized viral genomes and prepare defined targets, (iii) proteomics to express, purify, and characterize targets, (iv) structural biology to solve their crystal structures, and (v) pre-lead discovery to propose active scaffolds of antiviral molecules.

The human immunodeficiency virus-type 1 (HIV-1) reverse transcriptase (RT) is a multifunctional enzyme which displays DNA polymerase activity, which recognizes RNA and DNA templates, and a degradative ribonuclease H (RNase H) activity. While both RT functions are required for retroviral replication, until now only the polymerase function has been widely explored as drug target. We have identified a novel diketo acid derivative, 6-[1-(4-fluorophenyl)methyl-1H-pyrrol-2-yl)]-2,4-dioxo-5-hexenoic acid ethyl ester (RDS 1643), which inhibits in enzyme assays the HIV-1 RT-associated polymerase-independent RNase H activity but has no effect on the HIV-1 RT-associated RNA-dependent DNA polymerase (RDDP) activity and on the RNase H activities displayed by the Avian Myeloblastosis Virus and E. coli. Time-dependence studies revealed that the compound is active independently on the order of its addition to the reaction mixture, and inhibition kinetics studies demonstrated that RDS 1643 inhibits the RNase H activity noncompetitively, with a K(I) value of 17 microM. When RDS 1643 was combined with non-nucleoside RT inhibitors (NNRTI), such as efavirenz and nevirapine, results indicated that RDS 1643 does not affect the NNRTIs anti-RDDP activity and that, vice versa, the NNRTIs do not alter the RNase H inhibition by RDS 1643. When assayed on the viral replication in cell-based assays, RDS 1643 inhibited the HIV-1(IIIB) strain with an EC(50) of 14 microM. Similar results were obtained against the Y181C and Y181C/K103N HIV-1 NNRTI resistant mutant strains. RDS 1643 may be the first HIV-1 inhibitor selectively targeted to the viral RT-associated RNase-H function.

Pursuing our searches on quinolonecarboxylic acids we used a simple three-step one pot procedure to synthesize novel 1,7-disubstituted-6-nitroquinolones. The new derivatives were tested against Mycobacterium tuberculosis and Mycobacterium avium complex (MAC) as well as against both gram-positive and gram-negative bacteria. In vitro assays showed some derivatives were endowed with good inhibiting activities against tested mycobacteria. Some derivatives were also found more potent than ciprofloxacin and ofloxacin (used as reference drugs) against gram-positive bacteria.

A SAR study has been performed in order to evaluate how much the ester function could be a determinant for the anti-human immunodeficiency virus type-1 activity of pyrryl aryl sulfones (PASs), a potent family of non-nucleoside reverse transcriptase (RT) inhibitors discovered in the last years. Twenty-three new esters were prepared with the aim to enhance the inhibitory potency of 4a and 4c, two PAS agents endowed with good activity (EC50 = 0.14 microM) and deprived of cytotoxicity up to >200 microM. None of test derivatives was as potent as 4a and 4c and lacked of selectivity due to their higher cytotoxicity (compounds 22-25). Antiviral activity correlate with an ester ramified chain.

Twenty eight pyrrolo[1,2-a]quinoxalines bearing at position 4 various substituents related to the moieties present in classical and non classical antifolic agents were prepared and evaluated in vitro for antiproliferative activity. In an in vitro screening performed at NCI, several compounds emerged as potent antiproliferative agents at concentrations ranging between 10 and 100 microM. Interestingly, some of these compounds proved active also against bovine and murine DHFR (Farmaco 53 (1998) 480). More recently, a compound of classical antifolate type has been reported to be a potent inhibitor of hDHFR in vitro (Farmaco 58 (2003) 51). We then synthesized new derivatives that, in our hands, were endowed with in vitro antiproliferative activities as low as 3.4 microM against a panel of cell lines derived from hematological and solid tumours. In addition, a complete screening of cytotoxicity, antiretroviral HIV-1 and antimicrobial activity has been carried out.

In the course of a study on 1H-imidazol-1-amine derivatives as antifungal agents, we found that N-[(1,1'-biphenyl)-4-ylmethyl]-N-[(2,4-dichlorophenyl)methyl]-1H-imidazol-1-amine (1a) exhibited promising activities. In order to explore more in detail the structure-activity relationship of this new class of antifungal agents, we report now the synthesis and the biological activity of new analogues (1b-k) of compound 1a. The synthesis was performed using N-[(1,1'-biphenyl)-4-ylmethyl]-1H-imidazol-1-amine as starting material which was reacted with the proper arylmethyl halide. Most of the newly synthesized imidazolamines exhibited both fungal growth inhibition activity and cellular selectivity.

A series of azole derivatives, isoxazole or pyrimidine analogues of the antifungal drug bifonazole, were synthesized and tested in vitro against representative human pathogenic fungi (Candida albicans, Cryptococcus neoformans and Aspergillus fumigatus). They were also evaluated as antibacterial agents against Staphylococcus aureus and Salmonella spp. Only 5-(imidazol-1-yl-phenylmethyl)-2,4-diphenyl-pyrimidine 7c showed weak antimicrobial activity (MIC = 66 microM) against C. albicans, C. neoformans and S. aureus. Results of biological tests proved, therefore, that replacement of the biphenyl portion of the bifonazole with a phenylisoxazolyl or phenylpyrimidinyl moiety is not profitable for antimicrobial properties.

A series of pyrazole analogues of bifonazole, an antifungal drug used in clinical practice, 2a-h and 4a-h were synthesized and tested in vitro against Candida albicans, Cryptococcus neoformans and Aspergillus fumigatus, with no significant results. Imidazoles 2a-h were also tested in vivo for antiarrhythmic and antihypertensive activities; two of these compounds showed moderate activity against ventricular fibrillation caused by aconitine in rats. The above compounds were prepared by reaction of phenyl-[5 substituted 1-phenyl (or 1-methyl)-1H-pyrazol-4-yl]methanols with N,N'-carbonyldiimidazole (2a-h) or of the respective chloro derivatives with 1H-1,2,4-triazole (4a-h).

Acyclic glycosidopyrroles of type 1, synthesized in good overall yields, were evaluated for anti-viral activity. Compound 10i was found to inhibit the HIV-1 replication at concentrations that were very close to those cytotoxic for MT-4 cells. Compounds 10a,f,i inhibited both strains HSV-1 and HSV-2 at concentrations slightly below those cytotoxic for Vero cells. However for this series of glycosidopyrroles some relationship between calculated log P values and the observed cytotoxicity was found.

Following the discovery of anti-HIV properties of suramin great efforts were devoted to design novel NNRT agents with the aims to find novel drugs for the clinical therapeutic management of AIDS. Sulfone and sulfonamide derivatives were studied by NCI at Bethesda as potential anti-HIV-1 agents and nitrophenyl phenyl sulfone (NPPS) was selected as lead compound for further investigations. At the same time Merck Laboratories discovered L-737,126, a potent indolyl aryl sulfone with inhibitory activity against reverse transcriptase. These studies stimulated novel search in the sulfone series and both diarylsulfones and cyclic sulfone derivatives were investigated. Our decennial interest in chemotherapeutic agents containing a pyrrole ring pulsed us to synthesize and test as anti-HIV-1 agents a number of pyrryl aryl sulfones (PASs), pyrrolobenzothiadiazepine (PBTDs) and pyrrolobenzothiazepine related sulfones. The new sulfone derivatives inhibit selectively HIV-1 and were inactive against HIV-2. Most of them were as active as, if not more active than, nevirapine.

Acyclic glycosidopyrroles of type 3 were synthetized in good overall yields, according to the Scheme. When evaluated for antiviral activity against DNA and RNA viruses, only compound in which R1 = R2 = Ph, R3 = NH2 was found to inhibit the HIV-1 replication at concentrations that were not cytotoxic for MT-4 cells.

A series of novel 3,6-disubstituted 1,2,4-triazolo[3,4-b][1,3,4]thiadiazole derivatives was prepared and tested to evaluate their antimycotic, antibacterial and anti-HIV-1 activities. The reaction of thiocarbohydrazide with carboxylic acids at the melting temperature allows an improved preparation of the 5-substituted 4-amino-3-mercapto-1,2,4-triazole heterocycles which in turn allows an easier preparation of the 1,2,4-triazolo[3,4-b] [1,3,4]thiadiazole ring system. All tested compounds didn't show any significant activity.

The synthesis and the in vitro anti-HIV-1 activity of novel pyrrolo annulated benzothiadiazepine acetic acids and some related derivatives are reported. The new compounds share chemical features with pyrrolo[1,2-d][1,4]benzodiazepin-6-one 1 and Ro 5-3335 pyrrylbenzodiazepinone 4, two inhibitors of HIV-replication at the level of reverse transcriptase (RT) and transcriptional transactivation by Tat, respectively. Two derivatives, namely methyl 10,11-dihydropyrrolo[1,2-b][1,2,5]benzothiadiazepine-11-acetic-5,5 -dioxide (5a) and 1,12b-dihydro-2H-azeto[2,1-d]pyrrolo[1,2-b][1,2,5]benzoth iadiazepin-2-one 8,8-dioxide (7a), were found to exhibit a significant, although not very potent, activity against human immunodeficiency virus Type 1 (HIV-1).

Previous studies showed that some 2'-alkyloxyisoxazoles 2b-d obtained from 3-(diethylamino)-5-(2'hydroxy-4'-methoxyphenyl)isoxazole 2a were endowed with an interesting anti-group B rhinovirus activity (action on HRV-2 serotype). Other isoxazoles (WIN compounds) are well known to have anti-group A rhinovirus activity (action on HRV-14 serotype). To obtain an action similar to that of WIN compounds, starting from 2a, the 2'-acyl (3,4,5) and 2'alkyl (6,8) derivatives were synthesized. Also some Mannich bases (9,10) and bisisoxazoles (7,11,13,14) were studied. Though some of the tested compounds mainly exhibited anti-group B rhinovirus activity, their potency was less intense with respect to the above mentioned compounds 2b-d. The only N-methylpiperazinomethyl derivative 10 was slightly active against both tested serotypes.

N-acetyl-L-cysteine (NAC) is known to antagonize the PMA- or cytokine-stimulated HIV-1 replication in latently and acutely infected monocytic and lymphocytic cell lines, and to reduce the virus multiplication in acutely infected, PHA-stimulated PBMC. We here report on the modulatory effects of NAC on the HIV-1 multiplication in both chronically and acutely infected lymphocytes that produce high virus levels independently from cytokine activation. In both cases, NAC doses of 0.12 and 0.25 mM decreased, whereas doses of 0.5-2 mM increased the infectious HIV-1 yield. At these concentrations, the modulatory effect of NAC on the HIV-1 multiplication paralleled that on cell proliferation, suggesting a close correlation between the two phenomena; in fact, under conditions where NAC could not modulate the cell growth, the drug also failed to modulate the HIV-1 multiplication. High NAC concentrations (4-16 mM), which were able to increase the proliferative rate of both chronically infected H9/IIIB and normal T lymphocytes, increased up to 6-fold the virus multiplication in H9/IIIB cells but were inhibitory to HIV-1 in acutely infected cells. This inhibition was due to the fact that, like dextran sulfate, NAC interfered with an early event in the virus growth cycle. The finding that high NAC doses were also capable of preventing syncytium formation in H9/IIIB and C8166 (or MT-4) cocultures further indicated an interference of the drug with receptor-binding-related events.

-2',3'-Seco nucleosides 5 carrying fluorine and sulfur substituents at C-3' and C-5', respectively, of acyclic sugar moiety were synthesized in enantiomerically and diastereoisomerically pure form. These products and some structurally similar 1',2'-seco-2'-nor-and 1',2'-seco-nucleosides 3 and 4 were tested in vitro for cytotoxicity and antiviral activity. At non-cytotoxic concentrations the compounds were inactive against human immunodeficiency virus and herpes simplex virus type-1.

A series of 4-diazo-5-alkylsulphonamidopyrazoles (5) was prepared and tested for antitumor, antiviral and antimicrobial activity. Compounds (5a) and (5b) showed a selective, although not very potent cytostatic activity against L1210 and a human T lymphoblastoid cell line (C8166). Compounds (5a) and (5d-h) showed a selective anti-coxsackie B1 virus activity, whereas 5b was also endowed with some activity against Bacillus subtilis.