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The iminoquinone is an important moiety of a large number of antineoplastic drugs and plays a significant role in the nucleus of actinomycins, powerful, highly toxic, natural antibiotics that target DNA as intercalating agents. A series of polycyclic iminoquinonic compounds, 2-amino-3H-phenoxazin-3-one (1), 2-amino-1,9-diacetyl-3H-phenoxazin-3-one (2), 2-acetylamino-3H-phenoxazin-3-one (3), 3H-phenoxazin-3-one (4), 5H-pyrido[3,2-a]phenoxazin-5-one (5), and 5H-pyrido[3,2-a]phenothiazin-5-one (6), strictly related to the actinomycin chromophore, were synthesized for developing new anticancer intercalating drugs. The antiproliferative activity of these compounds, evaluated against representative human liquid and solid neoplastic cell lines, showed that 5 and its isoster 6 were the most active compounds inhibiting cell proliferation in a submicromolar range. Compound 5 was also evaluated against KB subclones (KB^MDR, KB^7D, and KB^V20C), which overexpress the MDR1/P-glycoprotein drug efflux pump responsible for drug resistance. All the above KB subclones did not show altered sensitivity to the antiproliferative activity of 5. UV−vis and ¹H NMR spectroscopy experiments support the phenoxazinone 5/DNA binding. Molecular mechanics methods were used to build a three-dimensional model of the 5/[d(GAAGCTTC)]2 complex. Electrostatic interactions between the hydrogen of the positively charged pyridine nitrogen of 5 and the negatively charged oxygen atoms (O4‘ and O5‘) of the cytosine C5 residue together with stacking forces contribute to the high antiproliferative activity. The metal(II)-assisted synthesis procedure of 5 is described, and the formation mechanism is proposed.

Molecular modeling analysis of compounds belonging to the recently published series of dihydro-alkoxy-benzyl-oxopyrimidines (DABOs), such as S-DABOs and DATNOs, gave support to the design of new 2,6-disubstituted benzyl-DABO derivatives as highly potent and specific inhibitors of the HIV-1 reverse transcriptase (RT). To follow up on the novel DABO derivatives, we decided to investigate the effect of electron-withdrawing substituents in the benzyl unit of the S-DABO skeleton versus their anti-HIV-1 activity. Such chemical modifications impacted the inhibitory activity, especially when two halogen units were introduced at positions 2 and 6 in the phenyl portion of the benzyl group bound to C-6 of the pyrimidine ring. Various 5-alkyl-2-(alkyl(or cycloalkyl)thio)-6-(2,6-dichloro(or 2,6-difluoro)phenylmethyl)-3,4-dihydropyrimidin-4(3H)-ones were then synthesized and tested as anti-HIV-1 agents in both cell-based and enzyme (recombinant reverse transcriptase, rRT) assays. Among the various mono- and disubstituted phenyl derivatives, the most potent were those containing a 6-(2,6-difluorophenylmethyl) substituent (F-DABOs), which showed EC50's ranging between 40 and 90 nM and selectivity indexes up to ≥5000. An excellent correlation was found between EC50 and IC50 values which confirmed that these compounds act as inhibitors of the HIV-1 RT. The structure−activity relationships of the newly synthesized pyrimidinones are presented herein.

Tetrahydroimidazo[4,5,1-jk][1,4]benzodiazepin-2(1H)-one and -thione (TIBO) derivatives (e.g., R82150) are potent, human immunodeficiency virus-1 (HIV-1)-specific, inhibitors of reverse transcriptase (RT) that are undergoing initial evaluation in clinical trials. Because HIV-1 has become resistant to other RT inhibitors, we investigated the potential for viral resistance to TIBO R82150 by serial in vitro passage of HIV-1IIIB in the presence of drug. R82150-resistant variants (> 100-fold increase in IC50) dominated the replicating virus population after only three or four passages. R82150-resistant virus was partially cross-resistant to other HIV-1-specific RT inhibitors, including nevirapine (approximately 10-fold increase in IC50) and 1-[(2-hydroxyethoxy)methyl]-6-(phenylthio)thymine (approximately 3.5-fold increase) but remained susceptible to 2',3'-dideoxynucleosides and phosphonoformate. DNA sequencing of cloned resistant RT, combined with site-specific mutational analyses and construction of mutant recombinant proviruses, demonstrated that a single, conservative amino acid substitution (Leu100 to Ile) in HIV-1 RT is responsible for high level R82150 resistance and partial nevirapine resistance. These studies indicate that a subtle mutation in HIV-1 RT can dramatically affect viral susceptibility to an HIV-1-specific RT inhibitor. The clinical efficacy of TIBO derivatives and other HIV-1-specific RT inhibitors may be limited by the emergence of drug-resistant viral strains.

Various cinnammoyl-based structures were synthesized and tested in enzyme assays as inhibitors of the HIV-1 integrase (IN). The majority of compounds were designed as geometrically or conformationally constrained analogues of caffeic acid phenethyl ester (CAPE) and were characterized by a syn disposition of the carbonyl group with respect to the vinylic double bond. Since the cinnamoyl moiety present in flavones such as quercetin (inactive on HIV-1-infected cells) is frozen in an anti arrangement, it was hoped that fixing our compounds in a syn disposition could favor anti-HIV-1 activity in cell-based assays. Geometrical and conformational properties of the designed compounds were taken into account through analysis of X-ray structures available from the Cambridge Structural Database. The polyhydroxylated analogues were prepared by reacting 3,4-bis(tetrahydropyran-2-yloxy)benzaldehyde with various compounds having active methylene groups such as 2-propanone, cyclopentanone, cyclohexanone, 1,3-diacetylbenzene, 2,4-dihydroxyacetophenone, 2,3-dihydro-1-indanone, 2,3-dihydro-1,3-indandione, and others. While active against both 3‘-processing and strand-transfer reactions, the new compounds, curcumin included, failed to inhibit the HIV-1 multiplication in acutely infected MT-4 cells. Nevertheless, they specifically inhibited the enzymatic reactions associated with IN, being totally inactive against other viral (HIV-1 reverse transcriptase) and cellular (RNA polymerase II) nucleic acid-processing enzymes. On the other hand, title compounds were endowed with remarkable antiproliferative activity, whose potency correlated neither with the presence of catechols (possible source of reactive quinones) nor with inhibition of topoisomerases. The SARs developed for our compounds led to novel findings concerning the molecular determinants of IN inhibitory activity within the class of cinnamoyl-based structures. We hypothesize that these compounds bind to IN featuring the cinnamoyl residue C&dbd;C−C&dbd;O in a syn disposition, differently from flavone derivatives characterized by an anti arrangement about the same fragment. Certain inhibitors, lacking one of the two pharmacophoric catechol hydroxyls, retain moderate potency thanks to nonpharmacophoric fragments (i.e., a m-methoxy group in curcumin) which favorably interact with an “accessory” region of IN. This region is supposed to be located adjacent to the binding site accommodating the pharmacophoric dihydroxycinnamoyl moiety. Disruption of coplanarity in the inhibitor structure abolishes activity owing to poor shape complementarity with the target or an exceedingly high strain energy of the coplanar conformation.

Several newly discovered potent and selective non-nucleoside inhibitors of human immunodeficiency virus-1 reverse transcriptase (RT) are undergoing evaluation in clinical trials. We studied the potential for development of viral resistance to one of the prototype compounds, BI-RG-587, a dipyridodiazepinone derivative. Human immunodeficiency virus-1 resistant to BI-RG-587 emerged after only one cycle of in vitro infection in the presence of the drug. Resistant virus was cross-resistant to the non-nucleoside tetrahydroimidazo[4,5,1-jk][1,4]benzodiazepin-2(1H)-thione derivative R82150 but remained susceptible to 2',3'-dideoxynucleosides and phosphonoformate. Both native (virion-associated) and recombinant RT derived from resistant virus were insensitive to BI-RG-587 and R82150. Nucleotide sequence analysis of multiple drug-resistant and -sensitive recombinant RT clones identified a single predicted amino acid change common to all resistant clones (tyrosine-181----cysteine). These studies suggest that the viral resistance to non-nucleoside RT inhibitors may develop in vivo. This possibility should be carefully monitored in clinical trials of these compounds.

In der vorliegenden Untersuchung werden neue Frequenzdaten bezüglich der häufigen und seltenen Varianten der polymorphen Enzymsysteme ADA, AK-1, 6-PGD, EsA, EsB, EsD, PGM-1, PGM-2, SOD-A, SEP, GPT und PGI in der italienischen Bevölkerung mitgeteilt. Außerdem wird eine umfassende Übersicht über die elektrophoretischen Enzymvarianten in Italien gegeben. Daraus ergibt sich eine beträchtliche genetische Heterogenität zwischen den verschiedenen Subpopulationen auf der italienischen Halbinsel sowie zwischen der Bevölkerung dieser und derjenigen Sardiniens. Schätzungen der average heterozygosity zeigen für Sardinien beträchtlich geringere Werte als für die Halbinsel bzw. für Sizilien. Schließlich wird über die Häufigkeit verschiedener seltener Enzymvarianten berichtet, die insgesamt der für nordeuropäische Bevölkerungen (Harris et al. 1974) entspricht.