Synthesis and Evaluation of Anticancer Activity of New 4-Acyloxy Derivatives of Robustic Acid.

In the present study, a series of 4-acyloxy robustic acid derivatives were synthesized and characterized for evaluation of their anti-cancer activity. The structures of these derivatives were elucidated by mass spectra (MS) nuclear magnetic resonance spectra (NMR). The single-crystal X-ray diffraction structure of one of these compounds was obtained, for further validation of the target compound structures. The anticancer activities of the target products were evaluated against human leukemic cells HL-60, human non-small cell lung carcinoma cells A-549, human hepatic carcinoma cells SMMC-7721, human hepatocellular carcinoma cells HepG2, and human cervical carcinoma cells Hela. Three compounds among them exhibited potent in-vitro cytotoxicity and excellent DNA topoisomerase I inhibitory activity, even at 0.1 mM concentrations. The most noteworthy observation was the minor toxicity of two of these compounds to normal cells, with an activity similar to the positive control in cancerous cells. A Surflex-Dock docking study was performed to investigate the topoisomerase I activity of all compounds. Of all the other compounds, the most sensitive compound was selected for further investigation of its effect on apoptosis induction and cell cycle regulation in HL-60 cells. Our results suggest that the anticancer effects of these compounds can be attributed to their pharmacological effects on topoisomerase I, cell apoptosis, and cell cycle. These findings suggest that robustic acid derivatives could be used as potential antitumor drugs.

Anticancer effects of O-aminoalkyl derivatives of alloxanthoxyletin and seselin.

Seselin and alloxanthoxyletin, naturally occurring pyranocoumarins, were recently isolated from a number of plant sources, such as family of Rutaceae. It was previously reported that their natural and synthetic derivatives show cytotoxic and antitumor activity. In the present study new series of O-aminoalkyl substituted alloxanthoxyletins and seselins were synthesized and evaluated for their anticancer toxicity. Microwave assisted synthesis was used, and the structures of the compounds were confirmed by 1H NMR, 13C NMR and MS spectroscopic data. The molecular and crystal structure of 3a was analyzed by single crystal X-ray diffraction. Alloxanthoxyletin derivatives 2a, 2b, and 2d showed the highest cytotoxic potential against HTB-140 cells with IC50 of 2.48, 2.80 and 2.98µM, respectively. In vitro drug sensitivity testing in HaCaT, A549 and HTB-140 cells were also performed. Tumor cells showed a higher sensitivity to tested compounds than normal cells. Compounds 2a, 2b and 2d inhibited cell migration and exerted stronger effect on A549 and HTB-140 cells than on HaCaT cells. In order to explain the basic mechanism of cell death induction we have investigated the effect of derivatives 2a, 2b and 2d on early and late apoptosis using annexin V-FITC/7-AAD flow cytometry analysis. Derivatives 2a and 2b were much more potent inducers of early apoptosis in HTB-140 cells compared to HaCaT and A549 cells.

Naturally occurring calanolides: an update on their anti-HIV potential and total syntheses.

Calanolides are naturally occurring pyranocoumarins found particularly in Calophyllum species (family Clusiaceae/ Guttiferae), and are well known for their potent anti-human immunodeficiency virus (HIV) activity. Various preclinical and clinical studies with this class of compounds are going on, and the National Cancer Institute (NCI) has been playing an active and supportive role in this regard. The present review covers an up-to-date literature of naturally occurring calanolides in view of their anti-HIV potential and total syntheses including information on related patents.

F18, a novel small-molecule nonnucleoside reverse transcriptase inhibitor, inhibits HIV-1 replication using distinct binding motifs as demonstrated by resistance selection and docking analysis.

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are one of the key components of antiretroviral therapy drug regimen against human immunodeficiency virus type 1 (HIV-1) replication. We previously described a newly synthesized small molecule, 10-chloromethyl-11-demethyl-12-oxo-calanolide A (F18), a (+)-calanolide A analog, as a novel anti-HIV-1 NNRTI (H. Xue et al., J. Med. Chem. 53:1397-1401, 2010). Here, we further investigated its antiviral range, drug resistance profile, and underlying mechanism of action. F18 consistently displayed potent activity against primary HIV-1 isolates, including various subtypes of group M, circulating recombinant form (CRF) 01_AE, and laboratory-adapted drug-resistant viruses. Moreover, F18 displayed distinct profiles against 17 NNRTI-resistant pseudoviruses, with an excellent potency especially against one of the most prevalent strains with the Y181C mutation (50% effective concentration, 1.0 nM), which was in stark contrast to the extensively used NNRTIs nevirapine and efavirenz. Moreover, we induced F18-resistant viruses by in vitro serial passages and found that the mutation L100I appeared to be the dominant contributor to F18 resistance, further suggesting a binding motif different from that of nevirapine and efavirenz. F18 was nonantagonistic when used in combination with other antiretrovirals against both wild-type and drug-resistant viruses in infected peripheral blood mononuclear cells. Interestingly, F18 displayed a highly synergistic antiviral effect with nevirapine against nevirapine-resistant virus (Y181C). Furthermore, in silico docking analysis suggested that F18 may bind to the HIV-1 reverse transcriptase differently from other NNRTIs. This study presents F18 as a new potential drug for clinical use and also presents a new mechanism-based design for future NNRTI.

Enantioselective total synthesis of (+)-scuteflorin A using organocatalytic asymmetric epoxidation.

We report the first enantioselective total synthesis of (+)-scuteflorin A in 14% overall yield, employing a chiral iminium salt to effect an organocatalytic asymmetric epoxidation of xanthyletin in >99% ee as the key step.

One-pot enantioselective synthesis of functionalized pyranocoumarins and 2-amino-4H-chromenes: discovery of a type of potent antibacterial agent.

Function-oriented design and synthesis of chiral small molecules with novel activity is a key goal in modern organic chemistry. As multiple antibiotic-resistant pathogens are emerging and causing serious diseases, the need for practical routes for the development of new types of antibacterial agents is very urgent. Herein, we present a highly efficient process for the synthesis of optically active pyranocoumarins and 2-amino-4H-chromenes through an organocatalytic Knoevenagel/Michael/cyclization sequence, and the preliminary biological studies of these new heterocyclic compounds revealed potent antibacterial activity. This study provides a novel strategy for further research and development of new types of antibacterial agents effective against human pathogens.

[Anti-HIV-1 activity and structure-activity relationship of pyranocoumarin analogs].

The purpose of this study is to find out anti-HIV-1 reverse transcriptase (RT)/protease (PR) activity and inhibition of virus replication in cell cultures of novel coumarin analogs and determine their structure-activity relationship. Coumarin derivatives have been demonstrated to inhibit the activity of HIV-1 RT/PR in cell free system. It also shows inhibition effects to HIV-1 replication in cell culture. Based on the Chinese traditional pharmacological characteristics and protein three dimension computer aided design, analogs of tetracyclic dipyranocoumarin were synthesized from natural leading compounds. We studied the relationship of antiviral effects and chemical structures via HIV-1 PR/RT enzyme models and cell culture model system. Seven compounds were designed and tested. Several compounds showed anti-HIV-1 activity in varying degrees, especially V0201 showed much higher anti-HIV-1 activity with 3.56 and 0.78 micromol x L(-1) of IC50 against HIV-1 PR/RT and 0.036 micromol x L(-1) against HIV-1 replication in PBMC cultures. V0201 with a novel structure may be a new leading compound. These new compounds are valuable for development of new anti-HIV drugs in the future.

Highly suppressing wild-type HIV-1 and Y181C mutant HIV-1 strains by 10-chloromethyl-11-demethyl-12-oxo-calanolide A with druggable profile.

We herein report a new compound: 10-chloromethyl-11-demethyl-12-oxo-calanolide A (20, EC(50) = 7.4 nM, SI = 1417), which demonstrates a druggable profile with 32.7% oral bioavailability in rat, tolerated oral single dose toxicity in mice, and especially the feature of highly efficient suppression of the wild-type HIV-1 and Y181C mutant HIV-1 at an EC(50) = 7.4 nM and EC(50) = 0.46 nM, respectively.

Anti-HBV and cytotoxic activities of pyranocoumarin derivatives.

Four natural pyranocoumarins clausenidin (1), nordentatin (2), clausarin (3), and xanthoxyletin (4) were isolated from the medicinal plant Clausena excavata. Recently, we found that 1 and 2 suppressed hepatitis B virus surface antigen in HepA2 cells, and in addition, 1-3 showed cytotoxic activity against four human cancer cell lines (A549, MCF7, KB, and KB-VIN). To explore the SAR of 1-4, 17 pyranocoumarin analogues (5-21) were designed and synthesized. Among these analogues, 5 and 10 were the most potent against hepatitis B virus with EC(50) values of 1.14 and 1.34microM, respectively. The most interesting result in the cytotoxicity assay was the significant activity of 1, 5, and 6 against the multi-drug resistant cell line, KB-VIN, without activity against the KB cell line. These data suggest that these three compounds could be useful hits for developing MDR-inverse drugs.

Synthesis and evaluation of novel substituted 5-hydroxycoumarin and pyranocoumarin derivatives exhibiting significant antiproliferative activity against breast cancer cell lines.

A library of novel 5-hydroxycoumarin and pyranocoumarin derivatives was constructed via silica sulfuric acid-catalyzed pechmann reaction and Pd(0)-catalyzed suzuki coupling in tandem, and their antiproliferative activities against breast cancer cells MCF-7 and MDA-MB-231 were evaluated. The results showed that compounds such as 6b, 6d, 6h, and 6k possess significant antiproliferative activity against MCF-7 cell line with the IC(50) values of 7.2, 5.3, 3.3, and 6.5 microM, respectively.

Improved synthesis and pharmacological evaluation of racemic 11 -demethylcalanolide A.

An improved and practical synthesis of racemic 11-demethylcalanolide A [(+/-)-1] was developed. This improved process involved Pechmann reaction on phloroglucinol with ethyl butyrylacetate to give 5,7,-dihydroxy4-n-propylcoumarin (3). Poly phosphoric acid (PPA) catalyzed acylation of compound (3) with crotonic acid, then intramolecular cyclization was achieved simultaneously in one step to afford the key intermediate chromanone (4). A microwave assisted synthetic method preparing chromene (6) using chromenynation of chromanone (4) with 1, 1-diethoxy-methyl-2-butene was conducted. Luche reduction of chromene (6) using NaBH4 with CeCl3 x 7H2O preferably gave (+/-)-1. The overall yield of this four step synthesis of (+/-)-1 was around 32% increasing one fold more than that of the previous method. An in vitro investigation showed that (+/-)-1 exhibited inhibitory activities against both wild-type and drug-resistant HIV-1 in HIV-1 RT and cell culture assay, and significant synergistic effects in combination with AZT, T-20, and indinavir. Its LD50 of acute toxicity in mice by intragastric administration and by intraperitoneal injection were 735.65 mg kg(-1) and 525.10 mg x kg(-1), respectively. The Cmax and AUC(0-infinity) were 0.54 microg x mL(-1) and 1.08 (microg x mL(-1) x h, respectively. The dynamics study of the inhibition of mice sera on HIV-1 RT showed that mice treated with 100 mg x kg(-1 (+/-)-1 once intraperitoneally were similar to that of 5 mg x kg(-1) of known clinical effective anti-HIV-1 drug neverapine. The results suggested that further investigation of the anti-HIV candidate (+/-)-1 was warranted.

Synthesis and complete assignment of the 1H and 13C NMR signals of some oxopyrancoumarin and oxofuropyrancoumarin derivatives.

The synthesis of four pyranocoumarins starting from phloroglucinol and the complete (1)H and (13)C NMR assignment of seven pyranocoumarins has been performed using 1D and 2D NMR techniques including COSY, HMQC and HMBC experiments.

Chemical library and structure-activity relationships of 11-demethyl-12-oxo calanolide A analogues as anti-HIV-1 agents.

(+)-Calanolide A ( 1) as a natural product was previously found as an inhibitor of HIV-1 reverse transcriptase. In our further investigation of its template, racemic 11-demethyl-12-oxo calanolide A ( 15), which had two fewer chiral carbon centers at the C-11 and C-12 positions than (+)-calanolide A, had a comparably inhibitory activity and better therapeutic index (EC 50 = 0.11 microM, TI = 818) against HIV-1 in vitro. A library based on its structural core was then designed and synthesized with introduction of nine diversity points in this article. The evaluations of anti-HIV-1 activity in vitro concluded their structure-activity relationships (SARs). A novel compound (10-bromomethyl-11-demethyl-12-oxo calanolide A, 123) was identified to have much higher inhibitory potency and therapeutic index (EC 50 = 2.85 nM, TI > 10,526) than those of the class compound against HIV-1. This finding provided a very important clue that modifications of the C ring at the C-10 position may be conducted to obtain drug candidates with better activity against HIV-1.

Chemical resolution of +/- -calanolide A, +/- -cordatolide A and their 11-demethyl analogues.

The chemical resolution of (+/-)-calanolide A and (+/-)-cordatolide A into their corresponding optically active enantiomers is described. Their inhibitory activities against HIV-1 are tested in vitro.

Pyranocoumarin, a novel anti-TB pharmacophore: synthesis and biological evaluation against Mycobacterium tuberculosis.

Pyranocoumarin compounds were identified to embody a novel and unique pharmacophore for anti-TB activity. A systematic approach was taken to investigate the structural characteristics. Focused libraries of compounds were synthesized and evaluated for their anti-TB activity in primary screening assays. Compounds shown to be active were further determined for MIC and MBC values. Three of the four bactericidal compounds (16, 17c, and 18f) were amino derivatives, with MIC values of 16 microg/mL and respective MBC values of 32, 32, and 64 microg/mL.

Interactions of a series of novel spiropyranocoumarin derivatives with reactive oxygen species.

A series of new spiro-substituted pyranocoumarin derivatives have been synthesized starting from the commercially available 7-hydroxycoumarin and the conformation of the pyran ring was investigated. The antioxidant activity of the compounds was evaluated in-vitro, by means of three different tests: the interaction with the stable free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH), the competition with DMSO for hydroxyl radicals scavenging ability and the quenching of superoxide anions generated by the enzymic xanthine-xanthine oxidase system. In the DPPH test the spiroadamantane derivative 13 was the most active and possessed a 40% inhibition at a concentration of 400 microM. All compounds successfully compete with DMSO for hydroxyl radicals generated by the Fe(3+)/ascorbic acid system. Compound 13 inhibited the oxidation of DMSO (3.125 mM) by 93% at 2 mM and by 71% at 0.25 mM. The corresponding second-order rate constants have been estimated and all compounds demonstrated higher rate constants compared with the reference compounds, 7-hydroxycoumarin and mannitol. Derivatives possessing extended conjugation showed the highest inhibitory activity for superoxide anions generated by the xanthine-xanthine oxidase system, although the results of this experiment possessed partial parallelism with the results observed in the other two tests. The overall obtained data indicate that the size of the different spiro- substituents influence the degree of free radical scavenging and demonstrate the importance of extended conjugation for the antioxidant activity. Due to its multiple mechanism of protective action, derivative 13 may serve as a lead for the development of analogues that could be useful for the treatment of pathophysiological processes dependent upon reactive oxygen species.

Coumarins and pyranocoumarins, potential novel pharmacophores for inhibition of measles virus replication.

A series of coumarin and pyranocoumarin analogues were evaluated in vitro for antiviral efficacy against measles virus (MV), strain Chicago. Of the 22 compounds tested for inhibition, six were found to have selectivity indices greater than 10. These were compounds 5-hydroxy-7-propionyloxy-4-propylcoumarin (2a), 5,7-bis(tosyloxy)-4-propylcoumarin (7); 5-hydroxy-4-propyl-7-tosyloxy-coumarin (8); 6,6-dimethyl-9-propionyloxy-4-propyl-2H,6H-benzo[1,2-b:3,4-b']dipyran-2-one (9); 6,6-dimethyl-9-pivaloyloxy-4-propyl-2H,6H-benzo[1,2-b:3,4-b']dipyran-2-one (10); and 7,8-cis-10,11,12-trans-4-propyl-6,6,10,11-tetramethyl-7,8,9-trihydroxy-2H,6H,12H-benzo[1 ,2-b:3,4-b':5,6-b'']tripyran-2-one (18). Three of the active drugs were propyl coumarin analogues (2a, 7 and 8), two were dipyranone or chromeno-coumarins (9 and 10), and one was a benzotripyranone with a coumarin nucleus (18). Some appeared to be rather specific and potent inhibitors of MV with EC50 values ranging from 0.2 to 50 microg/ml and the majority of the EC50 values being less than 5 pg/ml. The compounds inhibited an additional nine strains of MV, and in virucidal tests the drugs did not physically disrupt the virion to inhibit virus replication. The inhibitory activity for one of the compounds tested (7) was somewhat dependent on virus concentration and it was still active when added to cells up to 24 h after virus exposure. When used in combination with ribavirin, compound 7 appeared not to profoundly affect the antiviral efficacy of ribavirin or its cell-associated toxicity. However, a slightly antagonistic MV-inhibitory effect was observed at the highest concentration of ribavirin used in combination with most concentrations of compound 7 tested. This and related compounds may be valuable leads in the development of a potent and selective class of MV inhibitors that could be used in future in the clinic.