Antitumor agents 283. Further elaboration of desmosdumotin C analogs as potent antitumor agents: activation of spindle assembly checkpoint as possible mode of action

In our ongoing study of the desmosdumotin C (1) series, twelve new analogues, 21-32, mainly with structural modifications in ring-A, were prepared and evaluated for in vitro antiproliferative activity against several human tumor cell lines. Among them, the 4'-iodo-3,3,5-tripropyl-4-methoxy analogue (31) showed significant antiproliferative activity against multiple human tumor cell lines with ED(50) values of 1.1-2.8 μM. Elongation of the C-3 and C-5 carbon chains reduced activity relative to propyl substituted analogues; however, activity was still better than that of natural compound 1. Among analogues with various ether groups on C-4, compounds with methyl (2) and propyl (26) ethers inhibited cell growth of multiple tumor cells lines, while 28 with an isobutyl ether showed selective antiproliferative activity against lung cancer A549 cells (ED(50) 1.7 μM). The gene expression profiles showed that 3 may modulate the spindle assembly checkpoint (SAC) and chromosome separation, and thus, arrest cells at the G2/M-phase.     

Role of Cholesterol Pathways in Norovirus Replication

Norwalk virus (NV) is a prototype strain of the noroviruses (family Caliciviridae) that have emerged as major causes of acute gastroenteritis worldwide. I have developed NV replicon systems using reporter proteins such as a neomycin-resistant protein (NV replicon-bearing cells) and a green fluorescent protein (pNV-GFP) and demonstrated that these systems were excellent tools to study virus replication in cell culture. In the present study, I first performed DNA microarray analysis of the replicon-bearing cells to identify cellular factors associated with NV replication. The analysis demonstrated that genes in lipid (cholesterol) or carbohydrate metabolic pathways were significantly (P < 0.001) changed by the gene ontology analysis. Among genes in the cholesterol pathways, I found that mRNA levels of hydroxymethylglutaryl-coenzyme A (HMG-CoA) synthase, squalene epoxidase, and acyl-CoA:cholesterol acyltransferase (ACAT), ACAT2, small heterodimer partner, and low-density lipoprotein receptor (LDLR)-related proteins were significantly changed in the cells. I also found that the inhibition of cholesterol biosynthesis using statins (an HMG-CoA reductase inhibitor) significantly increased the levels of NV proteins and RNA, whereas inhibitors of ACAT significantly reduced the replication of NV in replicon-bearing cells. Up- or downregulation of virus replication with these agents significantly correlated with the mRNA level of LDLR in replicon-bearing cells. Finally, I found that the expression of LDLR promoted NV replication in trans by transfection study with pNV-GFP. I conclude that the cholesterol pathways such as LDLR expression and ACAT activity may be crucial in the replication of noroviruses in cells, which may provide potential therapeutic targets for viral infection.Human noroviruses are now the leading cause of food- or waterborne gastroenteritis illnesses responsible for more than 60% of outbreaks (10). It has been estimated that noroviruses cause 23 million cases of illness, 50,000 hospitalizations, and 300 deaths each year in the United States alone (19). Molecular epidemiological studies have confirmed a global distribution of these viruses (13). The major public health concern with human noroviruses is their ability to cause large outbreaks in group settings such as schools, restaurants, summer camps, military units, hospitals, nursing homes, and cruise ships. Human noroviruses are currently classified as NIAID category B priority pathogens (category B bioterrorism agents). Noroviruses generally cause mild to moderate gastroenteritis, but the disease can be severe to life-threatening in the young, the elderly, and immunocompromised patients. During the last decade, noroviruses have gained media attention for causing large-scale outbreaks of gastroenteritis on cruise ships, in nursing homes, etc. Although noroviruses do not multiply in food or water, they can cause large outbreaks because as few as 10 to 100 virions are sufficient to cause illness in a healthy adult (12). Recent reports of noroviral gastroenteritis outbreaks among hurricane Katrina evacuees underscores the importance of preventive and therapeutic measures for noroviruses to promote public health (32). However, no vaccines or antivirals are currently available for the prevention or treatment of norovirus disease in humans, which is largely due to the absence of a cell culture system for human noroviruses. The recent development of replicon-bearing cells for Norwalk virus (NV) (7) has made possible the study of NV replication in cells and the discovery of antivirals. We recently demonstrated that the system provides an excellent platform for screening small molecules for antivirals (3, 7). We also reported another NV replicon system with reporter genes (green fluorescent protein [GFP] or luciferase) to study virus replication (4).As a component of membrane structures and a precursor for the steroid hormones and bile acids, cholesterol is one of the most essential biological molecules in the body (8). Cholesterol levels are maintained by controlling both de novo synthesis (major) and dietary uptake (minor) of cholesterol (8). De novo synthesis of cholesterol is subject to complex regulatory controls by various enzymes such as 3-hydroxy-3-methyl glutaryl-coenzyme A (HMG-CoA) reductase and acyl-CoA:cholesterol acyltransferase (ACAT) (1, 8, 21). The synthesis of bile acids from cholesterol is also tightly controlled and represents an important factor the cholesterol homeostasis (14, 22, 23). In the present study, I first performed DNA microarray analysis of replicon-bearing cells to identify cellular factors associated with NV replication. Analysis showed genes in lipid (cholesterol) or carbohydrate metabolic pathways were significantly (P < 0.001) changed by the gene ontology analysis. Because it has been shown that bile acids are essential for the replication of porcine enteric calicivirus (PEC) in cells (6) and important natural modulators of cholesterol pathways, I was particularly interested in potential regulation genes in the cholesterol pathways. I demonstrate here that the modulation of the cholesterol pathways via inhibitors of HMG-CoA reductase or ACAT led to either up- or downregulation of the replication of NV. I also show that the expression level of low-density lipoprotein receptor (LDLR) was positively correlated with NV replication in cells. These studies suggest that the cholesterol pathway is crucial for norovirus replication and provide potential therapeutic targets for noroviral infection.     

Synthesis, structure-activity relationships and biological evaluation of caudatin derivatives as novel anti-hepatitis B virus agents

A series of caudatin derivatives were synthesized, and their anti-hepatitis B virus (HBV) activity was evaluated in HepG 2.2.15 cells. Most of the 3-O-substituted caudatin derivatives showed effective anti-HBV activity. Among the tested compounds, six compounds (2e-2h, 2l, 2r) exhibited significantly inhibitory activity against HBV DNA replication with IC(50) values in the range of 2.82-7.48 μM. Interestingly, two compounds (2e, 2f) had potent activity inhibiting not only the secretion of HBsAg (IC(50)=18.68 μM, 21.71 μM), HBeAg (IC(50)=13.16 μM, 33.73 μM), but also HBV DNA replication (IC(50)=7.48 μM, 3.63 μM). The structure-activity relationships (SARs) of caudatin derivatives had been discussed, which were useful for caudatin derivatives to be explored and developed as novel anti-HBV agents.     

10T024A, a new phenazine derivative, as a radical scavenger and a prostaglandin, leukotriene release suppressor

We identified a new radical scavenger, 10T024A (C(15)H(12)N(2)O(4)), from a culture of the Streptomyces sp. Spectroscopic elucidation indicated that this compound is a new phenazine derivative. 10T024A showed radical-scavenging activity with an ED(50) of 125 μM. Moreover, it showed prostaglandin D(2) (PGD(2)) and leukotriene B(4) (LTB(4)) release suppressive activity in rat basophilic leukemia (RBL-2H3) cells, at IC(50): 8 μM and 10 μM respectively.     

Antitumor agents 281. Design, synthesis, and biological activity of substituted 4-amino-7,8,9,10-tetrahydro-2H-benzo[h]chromen-2-one analogs (ATBO) as potent in vitro anticancer agents

In our exploration of new biologically active chemical entities, we designed and synthesized a novel class of antitumor agents, substituted 4-amino-7,8,9,10-tetrahydro-2H-benzo[h]chromen-2-one (ATBO) analogs. We evaluated their cytotoxic activity against seven human tumor cell lines from different tissues, and established preliminary structure-activity relationships (SAR). All analogs, except 8, 9, and 25-27, displayed potent tumor cell growth inhibitory activity. Especially, compounds 15 and 33 with a 4-methoxyphenyl group at position C-4 were extremely potent with ED(50) values of 0.008-0.064 and 0.035-0.32 μM, respectively. Compound 15 was the most potent analog compared with structurally related neo-tanshinlactone (e.g., 1) and 4-amino-2H-benzo[h]chromen-2-one (ABO, e.g., 4) analogs, and thus merits further exploration as an anti-cancer drug candidate.     

Potent inhibition of Norwalk virus by cyclic sulfamide derivatives

A new class of compounds that exhibit anti-norovirus activity in a cell-based system and embody in their structure a cyclosulfamide scaffold has been identified. The structure of the initial hit (compound 2a, ED(50) 4 μM, TD(50) 50 μM) has been prospected by exploiting multiple points of diversity and generating appropriate structure-activity relationships.     

Synthesis, evaluation and structural studies of antiproliferative tubulin-targeting azetidin-2-ones

A series of azetidin-2-ones substituted at positions 1, 3 and 4 of the azetidinone ring scaffold were synthesised and evaluated for antiproliferative, cytotoxic and tubulin-binding activity. In these compounds, the cis double bond of the vascular targeting agent combretastatin A-4 is replaced with the azetidinone ring in order to enhance the antiproliferative effects displayed by combretastatin A-4 and prevent the cis/trans isomerisation that is associated with inactivation of combretastatin A-4. The series of azetidinones was synthetically accessible via the Staudinger and Reformatsky reactions. Of a diverse range of heterocyclic derivatives, 3-(2-thienyl) analogue 28 and 3-(3-thienyl) analogue 29 displayed the highest potency in human MCF-7 breast cancer cells with IC(50) values of 7 nM and 10nM, respectively, comparable to combretastatin A-4. Compounds from this series also exhibited potent activity in MDA-MB-231 breast cancer cells and in the NCI60 cell line panel. No significant toxicity was observed in normal murine breast epithelial cells. The presence of larger, bulkier groups at the 3-position, for example, 3-naphthyl derivative 21 and 3-benzothienyl derivative 26, resulted in relatively lower antiproliferative activity in the micromolar range. Tubulin-binding studies of 28 (IC(50)=1.37 μM) confirmed that the molecular target of this series of compounds is tubulin. These novel 3-(thienyl) β-lactam antiproliferative agents are useful scaffolds for the development of tubulin-targeting drugs.     

Synthesis and Anticancer Activity of 3-(3-Oxoprop-1-enyl)-Substituted Analogues of Carbocyclic Pyrimidine Nucleosides and 2′,3′-Dideoxy Pyrimidine Nucleosides

Abstract

Various 2′, 3′ -dideoxy and carbocyclic pyrimidine nucleosides, and their corresponding 3-(3-oxoprop-1-enyl) derivatives, have been synthesized and evaluated against murine L1210 and P388 leukemias and Sarcoma 180 and human CCRF-CEM lymphoblastic leukemia. Among the compounds tested, 3-(3-oxoprop-1-enyl)-3′ -fluoro-3′ -deoxythymidine (17), 3-(3-oxoprop-1-enyl)-3′ -azido-3′ -deoxythymidine (15) and 3-(3-oxoprop-1-eny!)-(+)-1-[(lα, 3β, 4α)-3-hydroxy-4-(hydroxymethyl)cyclopentyl]-5-methyl-2,4 (lH,3H)pyrimidinedione (6) were found to be the most active with ED₅₀, values of 0.5,0.2,0.1, and 0.3 μM; 1.2, 0.5,1.0 and 1.0 μM; and 0.8,0.7,1.5, and 3.0μM, respectively. Our preliminary findings indicate that the 3-(3-oxoprop-1-enyl) derivative of carbocyclic thymidine is approximately 7 times more active than the 3-(3-oxoprop-1-enyl) derivative of carbocyclic thymine riboside against L1210 leukemia cells in vitro, with ED₅₀ values of 0.8 μM and 5.5 μM, respectively. These findings suggest that the cytotoxicity of these compounds not only is dependent upon the 3-(3-oxoprop-1-enyl)-substituted group, but also may vary with the sugar moiety.     

Immunosuppressive Bibenzyl-phenylpropane Hybrids from Dendrobium devonianum

Two new bibenzyl-phenylpropane hybrids, dendrophenols A and B ( 1 and 2 ), along with nine known bibenzyls, were isolated from the aerial part of Dendrobium devonianum Paxt. Their structures were determined by extensive spectroscopic methods and methylation. Bioassays revealed that compounds 1–9 were specifically immunosuppressive to T lymphocytes with IC₅₀ values ranging from 0.41 to 9.4 μM, of which compounds 1 (IC₅₀=1.62 μM) and 2 (IC₅₀=0.41 μM) were promising immunosuppressive agents for T lymphocytes with the selectivity indices of 19.9 and 79.5, respectively.     

Chemical Constituents of Primulina eburnea (Gesneriaceae) and Their Cytotoxic Activities

Two new ursane-type triterpenes, eburnealactones A and B ( 1 and 2 ), one new flavonoid, eburneatin A ( 6 ), and one new phenylethanoid glycoside, chiritoside D ( 7 ), along with 9 known compounds ( 3–5 , 8–13 ) were isolated from the whole plant of Primulina eburnea. Their structures were elucidated by comprehensive spectroscopic data analysis (IR, UV, NMR, and HR-ESI-MS). All the compounds were evaluated for their cytotoxic activities. Compound 1 showed significant cytotoxic activities against MKN-45 cell lines and 5637 cell lines with the IC₅₀ values of 9.57 μM and 8.30 μM, respectively. Compound 1 exhibited moderate cytotoxic activities against A549 and PATU8988T cell lines with the IC₅₀ values of 30.70 μM and 38.22 μM, respectively. Compound 6 exhibited moderate cytotoxic activities against MKN-45, HCT116, PATU8988T, 5637 and A-673 cell lines with the IC₅₀ values of 19.69 μM, 16.44 μM, 18.07 μM, 11.51 μM and 18.15 μM, respectively. Compound 5 showed moderate cytotoxic activities against A549 cell lines with the IC₅₀ values of 24.06 μM.     

6-Nitrobenzimidazole derivatives: potential phosphodiesterase inhibitors: synthesis and structure-activity relationship

6-Nitrobenzimidazole derivatives (1-30) synthesized and their phosphodiesterase inhibitory activities determined. Out of thirty tested compounds, ten showed a varying degrees of phosphodiesterase inhibition with IC(50) values between 1.5±0.043 and 294.0±16.7 μM. Compounds 30 (IC(50)=1.5±0.043 μM), 1 (IC(50)=2.4±0.049 μM), 11 (IC(50)=5.7±0.113 μM), 13 (IC(50)=6.4±0.148 μM), 14 (IC(50)=10.5±0.51 μM), 9 (IC(50)=11.49±0.08 μM), 3 (IC(50)=63.1±1.48 μM), 10 (IC(50)=120.0±4.47 μM), and 6 (IC(50)=153.2±5.6 μM) showed excellent phosphodiesterase inhibitory activity, much superior to the standard EDTA (IC(50)=274±0.007 μM), and thus are potential molecules for the development of a new class of phosphodiesterase inhibitors. A structure-activity relationship is evaluated. All compounds are characterized by spectroscopic parameters.     

Design and synthesis of benzo[c,d]indolone-pyrrolobenzodiazepine conjugates as potential anticancer agents

A series of benzo[c,d]indol-2(1H)one-PBD conjugates (11a-l) have been designed and synthesized as potential anticancer agents. These compounds were prepared by linking the C8-position of DC-81 with a benzo[c,d]indol-2(1H)one moiety through different alkane spacers in good yields and confirmed by (1)H NMR, mass and HRMS data. The DNA binding ability of these conjugates was evaluated by thermal denaturation studies and interestingly, compound 11l showed enhanced DNA binding ability. These compounds were also evaluated for their anticancer activity in selected human cancer cell lines of lung, skin, colon and prostate by using MTT assay method. These new conjugates showed promising anticancer activity with IC(50) values ranging from 1.05 to 36.49 μM. Moreover, cell cycle arrest in SubG1 phase was observed upon treatment of A549 cells with 1 and 2 μM (IC(50)) concentrations of compound 11l and it induced apoptosis. This is confirmed by Annexin V-FITC, Hoechst staining, caspase-3 activity as well as DNA fragmentation analysis.     

Synthesis and evaluation of 1-(substituted)-3-prop-2-ynylureas as antiangiogenic agents

Novel urea derivatives of alkynes have been designed, synthesized, and evaluated as potential cancer therapeutics leads. The most active 1-((3-chloromethyl)phenyl)-3-prop-2-ynylurea (1) exhibited cytotoxic effect against HELA and MCF-7 cell lines with IC(50) values of 1.55 μM and 1.48 μM, respectively. Further investigation on tube formation assay in human vein umbilical cells (HUVEC) demonstrated that 1 and methyl 4-(3-(3-ethynylureido)benzyloxy) benzoate (6) possess antiangiogenic activity, with minimum effective dose of 25 nM (for 1) and 6.25 μM (for 6). The ED(50) of 1 and 6 were found to be 0.26 μM and 17.52 μM, respectively. The results from in vitro tyrosine kinase assay indicated the EGFR inhibition of 1 over other kinases (VEGFR2, FGFR1 and PDGFRβ). The cytotoxicity of 1 against EGFR overexpressing cell line A431 (IC(50) 36 nM) was comparable to that of erlotinib. The binding mode of 1 from docking simulation in the EGFR active site revealed that the urea motif formed hydrogen bonding with Lys745, Thr854 and Asp855 in hydrophobic pocket of EGFR. Compound 1 is considered as a potential lead for further optimization.     

Novel quinoline-3-carboxamides (Part 2): Design,optimization and synthesis of quinoline based scaffold as EGFR inhibitors with potent anticancer activity

EGFR has a key role in cell growth. Its mutation and overexpression share in epithelial malignancies and tumor growth. Quinazoline and quinoline derivatives are common anticancer intracellular inhibitors of EGFR kinase, and their optimization is an important issue for development of potent targeted anticancer agents. Based on these facts, different strategies were used for optimizing our reported quinoline-3-carboxamide compound III (EGFR IC₅₀ = 5.283 µM and MCF-7 IC₅₀ = 3.46 µM) through different molecular modeling techniques. The optimized compounds were synthesized and subjected to EGFR binding assay and accordingly some more potent inhibitors were obtained. The most potent quinoline-3-carboxamides were the furan derivative 5o; thiophene derivative 6b; and benzyloxy derivative 10 showing EGFR IC₅₀ values 2.61, 0.49 and 1.73 μM, respectively. Furthermore, the anticancer activity of compounds eliciting potent EGFR inhibition (5o, 5p, 6b, 8a, 8b, and 10) was evaluated against MCF-7 cell line where they exhibited IC₅₀ values 3.355, 3.647, 5.069, 3.617, 0.839 and 10.85 μM, respectively. Compound 6b was selected as lead structure for further optimization hoping to produce more potent EGFR inhibitors.     

Identification and evaluation of apoptotic compounds from Garcinia oligantha

Four new compounds, oliganthins A-D (1-4), and one known caged xanthone gaudichaudione H (5) were isolated from the stems of Garcinia oligantha. The structures of the new compounds were elucidated by spectroscopic evidences. All of the five compounds were evaluated for their apoptosis-inducing effects using HeLa-C3 cells which have been genetically engineered to produce a fluorescent biosensor capable of detecting caspase-3 activation. All of them induced cell apoptosis at 10 μM or lower concentrations. The apoptotic activity of oliganthins A, B and gaudichaudione H were further confirmed by detecting the cleavage of PARP, which is the substrate of activated caspase-3, in these compounds-treated cells using the method of Western blot. Moreover, the values of IC(50) were measured for all five compounds on HeLa cells using the MTT assay. Among them, gaudichaudione H had the lowest IC(50) value of 0.90 μM, while the other four new compounds had IC(50) values of 1.58, 1.52, 4.15, and 7.82 μM, respectively. These results show that gaudichaudione H has the strongest apoptosis-inducing effect and cell growth inhibition effect among these xanthones and it may have the potential to be developed into a new anticancer agent.     

New bichalcone analogs as NF-κB inhibitors and as cytotoxic agents inducing Fas/CD95-dependent apoptosis

A series of novel bichalcone analogs were synthesized and evaluated in lipopolysaccharide (LPS)-activated microglial cells as inhibitors of nitric oxide (NO) and for in vitro anticancer activity using a limited panel of four human cancer cell lines. All analogs inhibited NO production. Compounds 4 and 11 exhibited optimal activity with IC(50) values of 0.3 and 0.5 μM, respectively, and were at least 38-fold better than the positive control. A mechanism of action study showed that both compounds significantly blocked the nuclear translocation of NF-κB p65 and up-regulation of iNOS at 1.0 μM. Compound 4 and three other analogs (3, 20, and 23) exerted significant in vitro anticancer activity GI(50) values ranging from 0.70 to 13.10 μM. A mode of action study using HT-29 colon cancer cells showed that 23 acts by inducing apoptosis signaling.     

Synthesis, α-glucosidase inhibition and molecular docking study of coumarin based derivatives

We have synthesized seventeen Coumarin based derivatives (1–17), characterized by ¹HNMR, ¹³CNMR and EI-MS and evaluated for α-glucosidase inhibitory potential. Among the series, all derivatives exhibited outstanding α-glucosidase inhibition with IC₅₀ values ranging between 1.10 ± 0.01 and 36.46 ± 0.70 μM when compared with the standard inhibitor acarbose having IC₅₀ value 39.45 ± 0.10 μM. The most potent derivative among the series is derivative 3 having IC₅₀ value 1.10 ± 0.01 μM, which are many folds better than the standard acarbose. The structure activity relationship (SAR) was mainly based upon by bring about difference of substituent’s on phenyl part. Molecular docking studies were carried out to understand the binding interaction of the most active compounds.     

The inhibition study of human UDP-glucuronosyltransferases with cytochrome P450 selective substrates and inhibitors

Human uridine-5'-diphosphoglucuronosyltransferases (UGTs) are the major phase II metabolizing enzymes. In the present study, five human UGTs (UGT1A1, 1A4, 1A6, 2B7, and 2B10) were individually expressed and used to examine the inhibition IC(50) values of 20 selective substrates and inhibitors of major cytochromes P450 (CYPs). The inhibition kinetics of UGT1A1 was also analyzed. The results showed that some compounds like α-naphthoflavone, paclitaxel, midazolam, cyclosporine A, and ketoconazole displayed strong inhibitions on UGT activities with their IC(50) values in a range of 4.1-26 μM. Especially, the IC(50) values were 4.1?±?0.8 μM for ketoconazole in inhibiting UGT1A1-mediated β-estradiol-3-glucuronidation, and 4.9?±?0.3 μM for paclitaxel towards UGT1A4-mediated midazolam-N-glucuronidation. Additionally, the IC(50) values of bupropion, tolbutamide, and testosterone in inhibiting UGT-mediated metabolisms were similar with the K(m) values of respective CYPs. Some kinetic behaviours of UGTs were following Michaelis-Menten kinetics, while some were not.     

New Dibenzocyclooctadiene Lignans and Phenolics from Kadsura heteroclite with Anti-Inflammatory Activity

A chemical investigation of K. heteroclite led to isolation of two new dibenzocyclooctadienes ( 1 and 2 ) together with 14 known compounds ( 3 – 16 ) by using multiple chromatographic techniques. New compounds ( 1 and 2 ) were obtained and identified by spectroscopic methods (HR-ESI-MS, 1D and 2D NMR, and ECD) as well as by comparison of their experimental data with those reported in the literatures. All the isolates were evaluated for their ability to modulate TNF-α production in lipopolysaccharide (LPS) stimulated RAW264.7 cells. Among them, compound 5 displayed the most inhibition against tumor necrosis factor (TNF)-α production with IC₅₀ value of 6.16±0.14 μM. Whereas, compounds ( 1 , 3 , and 6 ) showed the significant inhibition (IC₅₀ values ranging from 9.41 to 14.54 μM), and compounds ( 2 , 4 , 9 , 10 , 13 , 15 , and 16 ) exhibited moderate inhibition (IC₅₀ values ranging from 19.27 to 40.64 μM) toward TNF-α production, respectively.