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Twenty-three natural jamunone analogues along with a series of jamunone-based derivatives were synthesized and evaluated for their inhibitory effects against breast cancer (BC) MDA-MB-231 and MCF-7 cells. The preliminary structure-activity relationship revealed that the length of aliphatic side chain and free phenolic hydroxyl group at the scaffold played a vital role in anti-BC activities and the methyl group on chromanone affected the selectivity of molecules against MDA-MB-231 and MCF-7 cells. Among them, jamunone M (JM) was screened as the most effective anti-triple-negative breast cancer (anti-TNBC) candidate with a high selectivity against BC cells over normal human cells. Mechanistic investigations indicated that JM could induce mitochondria-mediated apoptosis and cause G0/G1 phase arrest in BC cells. Furthermore, JM significantly restrained tumor growth in MDA-MB-231 xenograft mice without apparent toxicity. Interestingly, JM could downregulate phosphatidylinositide 3-kinase (PI3K)/Akt pathway by suppressing protein-tyrosine phosphatase 1B (PTP1B) expression. These findings revealed the potential of JM as an appealing therapeutic drug candidate for TNBC.

Disulfide bond formation is a critical post-translational modification of newly synthesized polypeptides in the oxidizing environment of the endoplasmic reticulum and is mediated by protein disulfide isomerase (PDIA1). In this study, we report a series of α-aminobenzylphenol analogues as potent PDI inhibitors. The lead compound, AS15 , is a covalent nanomolar inhibitor of PDI, and the combination of AS15 analogues with glutathione synthesis inhibitor buthionine sulfoximine (BSO) leads to synergistic cell growth inhibition. Using nascent RNA sequencing, we show that an AS15 analogue triggers the unfolded protein response in glioblastoma cells. A BODIPY-labeled analogue binds proteins including PDIA1, suggesting that the compounds are cell-permeable and reach the intended target. Taken together, these findings demonstrate an extensive biochemical characterization of a novel series of highly potent reactive small molecules that covalently bind to PDI.

A series of triphenylethylene bisphenol analogues of the selective estrogen receptor modulator (SERM) tamoxifen were synthesized and evaluated for their abilities to inhibit aromatase, bind to estrogen receptor α (ER-α) and estrogen receptor β (ER-β), and antagonize the activity of β-estradiol in MCF-7 human breast cancer cells. The long-range goal has been to create dual aromatase inhibitor (AI)/selective estrogen receptor modulators (SERMs). The hypothesis is that in normal tissue the estrogenic SERM activity of a dual AI/SERM could attenuate the undesired effects stemming from global estrogen depletion caused by the AI activity of a dual AI/SERM, while in breast cancer tissue the antiestrogenic SERM activity of a dual AI/SERM could act synergistically with AI activity to enhance the antiproliferative effect. The potent aromatase inhibitory activities and high ER-α and ER-β binding affinities of several of the resulting analogues, together with the facts that they antagonize β-estradiol in a functional assay in MCF-7 human breast cancer cells and they have no E/Z isomers, support their further development in order to obtain dual AI/SERM agents for breast cancer treatment.

Several new series of 5,6,7-trimethoxyindole derivatives were synthesized and their structure-activity relationships (SARs) were studied. Some of these compounds exhibited strong antiproliferative activities in the submicromolar range. N-Methyl-5,6,7-trimethoxylindoles 21 and 31 displayed the highest antiproliferative activities, with IC50 values ranging from 22 to 125 nM in four human cancer cell lines and activated human umbilical vein endothelial cells (HUVECs). In addition to vascular disrupting activity verified by in vitro assays, compounds 21 and 31 displayed much higher selectivity for activated HUVECs versus quiescent HUVECs than those of colchicine and combretastatinA-4. The polymerization of cancer cell tubulin was inhibited and the cell cycle was arrested in the G2/M phase after treatment with 21 and 31. It was showed that 21 disrupted tumor vasculature by use of in vivo assay. Our results suggest that these two new compounds we synthesized may become the promising leads for the development of vascular disrupting agents.

Here, we report the systematic synthesis and characterization of simple phenols bearing a trialkyl(aryl)silyl or trialkyl(aryl)germyl functional group as a hydrophobic substituent. These silicon and germanium analogues exhibited higher hydrophobicity than the corresponding carbon analogues, with a difference in log P value of approximately 0.6, independent of the alkyl(aryl) species. Trimethylsilylphenol and trimethylgermylphenol exhibited smaller pK(a) values than the corresponding carbon analogue or unsubstituted phenol, indicating that trialkylsilyl and trialkylgermyl functional groups have a negative substituent constant (σ). The trialkylsilyl- and trialkylgermylphenols exhibited more potent estrogenic activity as compared with the carbon analogues. The substituent parameters and structure-activity relationship reported here may be helpful for drug discovery utilizing the heavier group 14 elements.

Alzheimer's disease (AD) is a complex multifactorial syndrome. Metal chelator and Aβ inhibitor are showing promise against AD. In this report, three small hybrid compounds (1, 2, and 3) have been designed and synthesized utilizing salicylaldehyde (SA) based Schiff bases as the chelators and benzothiazole (BT) as the recognition moiety for AD treatment. These conjugates can capture Cu(2+) from Aβ and become dimers upon Cu(2+) coordination and show high efficiency for both Cu(2+) elimination and Aβ assembly inhibition. Besides, the complexes have superoxide dismutase (SOD) activity and significant antioxidant capacity and are capable of decreasing intracellular reactive oxygen species (ROS) and increasing cell viability. All these results indicate that the multifunctional metal complexes which have Aβ specific recognition moiety and metal ion chelating elements show the potential for AD treatment. Therefore, our work will provide new insights into exploration of more potent amyloid inhibitors.

Since the discovery of the endocannabinoid system, evidence has been progressively accumulating to suggest that 2-arachidonoylglycerol (2-AG) rather than anandamide (AEA) is the endogenous ligand for both cannabinoid (CB) receptors. Moreover, other studies have shown that another lipid molecule, 2-arachidonyl-glycerol ether (2-AGE, noladin ether), which acts as a full agonist at cannabinoid receptors, might occur in tissues. Having previously designed a resorcinol-AEA hybrid model, in this paper we have explored the cannabinoid receptor binding properties, the CB1 functional activity, and the stability to plasma esterases of a novel series of compounds characterized by the conversion of the amide head into the glycerol-ester or glycerol-ether head, typical of 2-AG or the "putative" endocannabinoid 2-AGE, respectively. Glyceryl esters 39 and 41 displayed greater potency for CB1 (Ki in the nanomolar range) than for CB2 receptors plus the potential to be exploited as useful hits for the development of novel 2-AG mimetics.

Estrogen deficiency in postmenopausal women or elderly men is often associated with the skeletal disease osteoporosis. The supplementation of estradiol (E2) in osteoporotic patients is known to prevent bone fracture but cannot be administered because of adverse effect. As 17β-hydroxysteroid dehydrogenase type 2 (17β-HSD2) oxidizes E2 to its inactive form estrone (E1) and has been found in osteoblastic cells, it is an attractive target for the treatment of osteoporosis. Twenty-one novel, naphthalene-derived compounds have been synthesized and evaluated for their 17β-HSD2 inhibition and their selectivity toward 17β-HSD1 and the estrogen receptors (ERs) α and β. Compound 19 turned out to be the most potent and selective inhibitor of 17β-HSD2 in cell-free assays and had a very good cellular activity in MDA-MB-231 cells, expressing naturally 17β-HSD2. It also showed marked inhibition of the E1-formation by the rat and mouse orthologous enzymes and strong inhibition of monkey 17β-HSD2. It is thus an appropriate candidate to be further evaluated in a disease-oriented model.

In order to discover novel probes that may help in the investigation and control of infectious diseases through a new mechanism of action, we have evaluated a library of phenol-based natural products (NPs) for enzyme inhibition against four recently characterized pathogen β-family carbonic anhydrases (CAs). These include CAs from Mycobacterium tuberculosis, Candida albicans, and Cryptococcus neoformans as well as α-family human CA I and CA II for comparison. Many of the NPs selectively inhibited the mycobacterial and fungal β-CAs, with the two best performing compounds displaying submicromolar inhibition with a preference for fungal over human CA inhibition of more than 2 orders of magnitude. These compounds provide the first example of non-sulfonamide inhibitors that display β over α CA enzyme selectivity. Structural characterization of the library compounds in complex with human CA II revealed a novel binding mode whereby a methyl ester interacts via a water molecule with the active site zinc.

Inhibition of 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1) is a novel and attractive approach to reduce the local levels of the active estrogen 17β-estradiol in patients with estrogen-dependent diseases like breast cancer or endometriosis. With the aim of optimizing the biological profile of 17β-HSD1 inhibitors from the hydroxyphenylnaphthol class, structural optimizations were performed at the 1-position of the naphthalene by introduction of different heteroaromatic rings as well as substituted phenyl groups. In the latter class of compounds, which were synthesized applying Suzuki-cross coupling, the 3-methanesulfonamide 15 turned out to be a highly potent 17β-HSD1 inhibitor (IC(50) = 15 nM in a cell-free assay). It was also very active in the cellular assay (T47D cells, IC(50) = 71 nM) and selective toward 17β-HSD2 and the estrogen receptors α and β. It showed a good membrane permeation and metabolic stability and was orally available in the rat.

Structure-based design was applied to the optimization of a series of 2-(quinazolin-2-yl)phenols to generate potent and selective ATP-competitive inhibitors of the DNA damage response signaling enzyme checkpoint kinase 2 (CHK2). Structure-activity relationships for multiple substituent positions were optimized separately and in combination leading to the 2-(quinazolin-2-yl)phenol 46 (IC(50) 3 nM) with good selectivity for CHK2 against CHK1 and a wider panel of kinases and with promising in vitro ADMET properties. Off-target activity at hERG ion channels shown by the core scaffold was successfully reduced by the addition of peripheral polar substitution. In addition to showing mechanistic inhibition of CHK2 in HT29 human colon cancer cells, a concentration dependent radioprotective effect in mouse thymocytes was demonstrated for the potent inhibitor 46 (CCT241533).

Starting from bisphenolic bis-styrylbenzene DF-9 (4), β-amyloid (Aβ) binding affinity and specificity for phenolic bis-styrylbenzenes, monostyrylbenzenes, and alkyne controls were determined by fluorescence titration with β-amyloid peptide Aβ(1-40) and a fluorescence assay using APP/PS1 transgenic mouse brain sections. Bis-styrylbenzene SAR is derived largely from work on symmetrical compounds. This study is the first to describe Aβ binding data for bis-styrylbenzenes unsymmetrical in the outer rings. With one exception, binding affinity and specificity were decreased by adding and/or changing the substitution pattern of phenol functional groups, changing the orientation about the central phenyl ring, replacing the alkene with alkyne bonds, or eliminating the central phenyl ring. The only compound with an Aβ binding affinity and specificity comparable to 4 was its 3-hydroxy regioisomer 8. Like 4, 8 crossed the blood-brain barrier and bound to Aβ plaques in vivo. By use of a DPPH assay, phenol functional groups with para orientations seem to be a necessary, but insufficient, criterion for good free radical scavenging properties in these compounds.

Estradiol (E2), the most important estrogen in humans, is involved in the initiation and progression of estrogen-dependent diseases such as breast cancer and endometriosis. Its local production in the target cell is regulated by 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1), which catalyzes E2-formation by reduction of the weak estrogen estrone (E1). Because the enzyme is expressed in the diseased tissues, inhibition of 17β-HSD1 is considered as a promising therapy for the treatment of estrogen-dependent diseases. For the development of novel inhibitors, a structure- and ligand-based design strategy was applied, resulting in bicyclic substituted hydroxyphenylmethanones. In vitro testing revealed high inhibitory potencies toward human placental 17β-HSD1. Compounds were further evaluated with regard to selectivity (17β-HSD2, estrogen receptors ERα and ERβ), intracellular activity (T47D cells), and metabolic stability. The most promising compounds, 14 and 15, showed IC(50) values in the low nanomolar range in the cell-free and cellular assays (8-27 nM), more than 30-fold selectivity toward 17β-HSD2 and no affinity toward the ERs. The data obtained make these inhibitors interesting candidates for further preclinical evaluation.

A series of N-acyl O-amino derivatives of seco-CBI-indole(2) are reported and examined as prototypical members of a unique class of reductively activated (cleaved) prodrugs of the duocarmycin and CC-1065 family of antitumor agents. These prodrugs were designed to be potentially preferentially activated in hypoxic tumor environments which carry an intrinsically higher concentration of "reducing" nucleophiles (e.g., thiols) capable of activating such derivatives by nucleophilic cleavage of a weak N-O bond. A remarkable range of stabilities and a resulting direct correlation with in vitro/in vivo biological potencies was observed for these prodrugs, even enlisting subtle variations in the electronic and steric environment around the weak N-O bond. An in vivo evaluation of several of the prodrugs demonstrates that some approach the potency and exceed the efficacy of the free drug itself (CBI-indole(2)), suggesting the prodrugs may offer an additional advantage related to a controlled or targeted release.

An array of analogues of the wasp toxin philanthotoxin-433, in which the asymmetric polyamine moiety was exchanged for spermine and the headgroup replaced with a variety of structurally diverse moieties, was prepared using parallel solid-phase synthesis approaches. In three analogues, the spermine moiety was extended with an amino acid tail, six compounds contained an N-acylated cyclohexylalanine, and four analogues were based on a novel diamino acid design with systematically changed spacer length between N-cyclohexylcarbonyl and N-phenylacetyl substituents. The analogues were studied using two-electrode voltage-clamp electrophysiology employing Xenopus laevis oocytes expressing GluA1(i) AMPA or GluN1/2A NMDA receptors. Several of the analogues showed significantly increased inhibition of the GluN1/2A NMDA receptor. Thus, an analogue containing N-(1-naphtyl)acetyl group showed an IC(50) value of 47 nM. For the diamino acid-based analogues, the optimal spacer length between two N-acyl groups was determined, resulting in an analogue with an IC(50) value of 106 nM.

Inhibitors of the chaperone Hsp90 are potentially useful as chemotherapeutic agents in cancer. This paper describes an application of fragment screening to Hsp90 using a combination of NMR and high throughput X-ray crystallography. The screening identified an aminopyrimidine with affinity in the high micromolar range and subsequent structure-based design allowed its optimization into a low nanomolar series with good ligand efficiency. A phenolic chemotype was also identified in fragment screening and was found to bind with affinity close to 1 mM. This fragment was optimized using structure based design into a resorcinol lead which has subnanomolar affinity for Hsp90, excellent cell potency, and good ligand efficiency. This fragment to lead campaign improved affinity for Hsp90 by over 1,000,000-fold with the addition of only six heavy atoms. The companion paper (DOI: 10.1021/jm100060b) describes how the resorcinol lead was optimized into a compound that is now in clinical trials for the treatment of cancer.

The 14 kDa homodimeric N1L protein is a potent vaccinia and variola (smallpox) virulence factor. It is not essential for viral replication, but it causes a strong attenuation of viral production in culture when deleted. The N1L protein is predicted to contain the BH3-like binding domain characteristic of Bcl-2 family proteins, and it is able to bind the BH3 peptides. Its overexpression has been reported to prevent infected cells from committing apoptosis. Therefore, interfering with the N1L apoptotic blockade may be a legitimate therapeutic strategy affecting the viral growth. By using in silico ligand docking and an array of in vitro assays, we have identified submicromolar (600 nM) N1L antagonists belonging to the family of polyphenols. Their affinity is comparable to that of the BH3 peptides (70-1000 nM). We have also identified the natural polyphenol resveratrol as a moderate N1L inhibitor. Finally, we show that our ligands efficiently inhibit growth of vaccinia virus.

17beta-Hydroxysteroid dehydrogenase type 1 (17beta-HSD1) is responsible for the catalytic reduction of weakly active E1 to highly potent E2. E2 stimulates the proliferation of hormone-dependent diseases via activation of the estrogen receptor alpha (ERalpha). Because of the overexpression of 17beta-HSD1 in mammary tumors, this enzyme should be an attractive target for the treatment of estrogen-dependent pathologies. Recently, we have reported on a series of potent 17beta-HSD1 inhibitors: bis(hydroxyphenyl) azoles, thiophenes, and benzenes. In this paper, different substituents are introduced into the core structure and the biological properties of the corresponding inhibitors are evaluated. Computational methods and analysis of different X-rays of 17beta-HSD1 lead to identification of two different binding modes for these inhibitors. The fluorine compound 23 exhibits an IC(50) of 8 nM and is the most potent nonsteroidal inhibitor described so far. It also shows a high selectivity (17beta-HSD2, ERalpha) and excellent pharmacokinetic properties after peroral application to rats.

We have previously shown that modification of polyphenols with a ferrocenyl group can dramatically enhance their cytotoxicity. We now present two new [3]ferrocenophane compounds, one of which has an antiproliferative effect seven times stronger than the corresponding noncyclic species, with IC50 values of 90 and 94 nM on hormone-independent MDA-MB-231 breast and PC-3 prostate cancer cell lines, respectively. Solubility studies in water using methylated beta-cyclodextrin and electron transfer studies are also presented.

Bearing in mind the pharmacophoric requirements of both (-)-trans-Delta(9)-tetrahydrocannabinol (THC) and anandamide (AEA), we designed a novel pharmacophore consisting of both a rigid aromatic backbone and a flexible chain with the aim to develop a series of stable and potent ligands of cannabinoid receptors. In this paper we report the synthesis, docking studies, and structure-activity relationships of new resorcinol-anandamide "hybrids" differing in the side chain group. Compounds bearing a 2-methyloctan-2-yl group at position 5 showed a significantly higher affinity for cannabinoid (CB) receptors, in particular when an alkyloxy chain of 7 or 10 carbon atoms was also present at position 1. Derivative 32 was a potent CB(1) and CB(2) ligand, with K(i) values similar to that of WIN 55-212 and potent antinociceptive activity in vivo. Moreover, derivative 38, although less potent, proved to be the most selective ligand for CB(2) receptor (K(i)(CB(1)) = 1 muM, K(i)(CB(2)) = 35 nM).

Salicylaldoximes possess a hydrogen-bonded pseudocyclic A' ring in place of the typical phenolic A ring that is characteristic of most estrogen receptor (ER) ligands. Monoaryl-substituted salicylaldoximes were obtained by replacing the phenol moiety (ring A) of the ERbeta pharmacophore with the pseudocycle A' ring, which has previously been shown to behave as a bioequivalent of phenols in nonselective ER ligands. In this series, small substituents (CH 3, CN, Cl) were introduced into the central phenyl scaffold. An efficient sequential halogen-selective double cross-coupling reaction was developed for the synthesis of the methyl-substituted ER ligand. The measured ERbeta affinity proved to be very sensitive to the effect of central core substituents. The binding affinities of the compounds herein reported were in good agreement with the results of computational docking analysis. The chloro-substituted derivative showed the highest beta affinity and selectivity, and it also proved to be an ERbeta partial agonist with an EC 50 of 11 nM.

In studies aimed toward identifying effective and safe inhibitors of kinase signaling cascades that underlie ischemia/reperfusion (I/R) injury, we synthesized a series of pteridines and pyridopyrazines. The design strategy was inspired by the examination of naturally occurring PI3K inhibitors such as wortmannin and quercetin, and building a pharmacophore-based model used for optimization. Structural modifications led to hybrid molecules which incorporated aminopyrimidine and aminopyridine moieties with ATP mimetic characteristics into the pharmacophore motifs to modulate kinase affinity and selectivity. Elaborations involving substitutions of the 2 and 4 positions of the pyrimidine or pyridine ring and the 6 and 7 positions of the central pyrazine ring resulted in in vivo activity profiles which identified potent inhibitors of vascular endothelial growth factor (VEGF) induced vascular leakage. Pathway analysis identified a diaminopteridine-diphenol as a potent and selective phosphatidylinositol-3-kinase (PI3K) inhibitor. The structure-activity relationship studies of various analogues of diaminopteridine-diphenol-based on biochemical assays resulted in potent inhibitors of PI3K.

A series of phenolic hydrazones were synthesized and evaluated for their inhibition of macrophage migration inhibitory factor (MIF) tautomerase activity. Compound 7 emerged as a potent inhibitor of MIF with an IC50 of 130 nM. Compound 7 dose-dependently suppressed TNFalpha secretion from lipopolysaccharide stimulated macrophages. The therapeutic importance of the MIF inhibition by 7 is demonstrated by the significant protection from the lethality of sepsis when administration of the compound was initiated in a clinically relevant time frame.

A series of C5-substituted 1,2,4-triaryl-1H-imidazoles was synthesized. Their gene-activating properties were determined on estrogen receptor alpha positive MCF-7 breast cancer cells, stably transfected with the plasmid EREwtcluc (MCF-7-2a cells). The influence of 4-OH and 2-Cl substituents on the phenyl rings as well as the significance of a methyl, ethyl, or phenyl group at C5 on the estrogen receptor binding and the resulting gene activation in MCF-7-2a cells was studied. The alkyl and aryl groups at C5 of 1,2,4-tris(4-hydroxyphenyl)-1H-imidazole 1 increased the transactivation, while chlorine atoms on the phenyl rings diminished this effect. 5-Ethyl-1,2,4-tris(4-hydroxyphenyl)-1H-imidazole 9 was identified as the most active compound. Its excellent transcriptional activity did not only depend on the C5 ethyl group, but also on the three hydroxyl groups of the phenyl rings. Compounds (11-14) with a reduced number of hydroxyl groups displayed distinctly lower gene activation.

This report describes the first results of a rational hit-finding strategy to design novel small molecule antiinflammatory drugs targeting selectins, a family of three cellular adhesion molecules. Based on recent progress in understanding of molecular interaction between selectins and their natural ligands as well as progress in clinical development of synthetic antagonists like 1 (bimosiamose, TBC1269), this study was initiated to discover small molecule selectin antagonists with improved pharmacological properties. Considering 1 as template structure, a ligand-based approach followed by focused chemical synthesis has been applied to yield novel synthetic small molecules (MWr < 500) with a trihydroxybenzene motif, bearing neither peptidic nor glycosidic components, with nanomolar in vitro activity. Biological evaluation involves two kinds of in vitro assays, a static molecular binding assay, and a dynamic HL-60 cell attachment assay. As compared to controls, the novel compounds showed improved biological in vitro activity both under static and dynamic conditions.

The synthesis and study of the antioxidant and vasodilator properties of a new class of phenols able to release nitric oxide are described. The products were designed through a symbiotic approach using selected phenols and selected nitrooxy and furoxan NO-donors as reference models. The antioxidant activities of the hybrid products were assessed by detecting the 2-thiobarbituric acid reactive substances (TBARS) produced in the ferrous salt/ascorbate-induced autoxidation of lipids present in microsomial membranes of rat hepatocytes. The vasodilator activity was assessed on rat aortic strips precontracted with phenylephrine. Some of the products (13, 35, 37, 60-62, 64) behave principally as vasodilators and others as antioxidants (24, 32, 72), and the two properties are relatively balanced in 19, 41, and 68. Further in vivo studies should clarify whether some of these products may become preclinical candidates for the treatment of cardiovascular disease underpinned by atheroma.

Estrogen receptor beta (ERbeta), a less active ER subtype that appears to have a restraining effect on the more active ERalpha, could be a factor that determines the level of estrogen action in certain estrogen target tissues. ERbeta is found in breast cancer, and its levels relative to ERalpha decline with disease progression. Thus, the independent quantification of ERalpha and ERbeta levels in breast cancer by imaging might be predictive of responses to different hormone therapies. To develop an imaging agent for ERbeta, we synthesized a fluoroethyl analogue of DPN (2,3-bis(4-hydroxyphenyl)propanonitrile), a known ERbeta-selective ligand. This analogue, FEDPN (5-fluoro-(2R,3S)-2,3-bis(4-hydroxyphenyl)pentanenitrile), has an 8.3-fold absolute affinity preference for ERbeta. [18F]Fluoride-labeled FEDPN was prepared from a toluenesulfonate precursor, which provided [18F]FEDPN with a specific activity greater than 3100 Ci/mmol after HPLC purification. Biodistribution studies in immature female rats using estradiol as a blocking agent revealed specific uptake of [18F]FEDPN in the uterus and ovaries. Experiments using ERalpha- and ERbeta-knockout mice demonstrated the expected ERalpha-subtype dependence in the tissue uptake of the known 16alpha-[18F]fluoro-17beta-estradiol ([18F]FES), which has a 6.3-fold preference for ERalpha. The tissue uptake of [18F]FEDPN in the ER knockout mice showed some evidence of mediation by ERbeta, but the levels of specific uptake of this agent were relatively modest. Based on our results, imaging of ERalpha can be done effectively with [18F]FES, but imaging of ERbeta will likely require agents with more optimized ERbeta binding affinity and selectivity than [18F]FEDNP.

Candidate estrogen receptor (ER) ligands with two phenolic residues on a three-dimensional hydrophobic core structure (carborane, bicyclo[2.2.2]octene, or adamantane) were synthesized and biologically evaluated. The biological properties of the ligands were markedly dependent on the nature of the hydrophobic core structure. Bis(4-hydroxyphenyl)-o-carborane (6) was a partial agonist/antagonist for ER. 1,2-Bis(4-hydroxyphenyl)bicyclo[2.2.2]octene (10) exhibited potent agonist activity for ER, even though the two phenolic groups are located similarly to those of 6.

We report here the synthesis and the strong and unexpected antiproliferative effect of the organometallic diphenolic compound 1,1-bis(4'-hydroxyphenyl)-2-ferrocenyl-but-1-ene (4) on both hormone-dependent (MCF7) and -independent (MDA-MB231) breast cancer cells (IC(50) = 0.7 and 0.6 microM). Surprisingly, 6 [1,2-bis(4'-hydroxyphenyl)-2-ferrocenyl-but-1-ene], the regioisomer of 4, shows only a modest effect on these cell lines. This pertinent organometallic modification seems to trigger an intracellular oxidation of the structurally favorable compound 4, leading to the generation of a potent cytotoxic compound.

The first small molecule agonists of the estrogen-related receptors have been identified. GSK4716 (3) and GSK9089 (4) show binding to ERRgamma with remarkable selectivity over the classical estrogen receptors. Notably, in cell-based reporter assays, 3 mimics the protein ligand PGC-1alpha in activation of human ERRbeta and ERRgamma.

Benzazepines 1 and 2 (SCH 23390 and SCH 39166, respectively) are two classical benzazepine D1/D5 antagonists, with Ki values 1.4 and 1.2 nM, respectively. Compound 2 has been in human clinical trials for a variety of diseases, including schizophrenia, cocaine addition, and obesity. Both 1 and 2 displayed low plasma levels and poor oral bioavailability, due to rapid first-pass metabolism of the phenol moieties. Several heterocyclic systems containing an N-H hydrogen bond donor were synthesized and evaluated as phenol isosteres. The preference orientation of the hydrogen bond was established by comparison of analogues containing different NH vectors. Replacement of the phenol group of 2 with an indole ring generated the first potent D1/D5 antagonist 11b. Further optimization led to the synthesis of very potent benzimidazolones 19, 20 and benzothiazolone analogues 28, 29. These compounds have excellent selectivity over D2-D4 receptors, alpha2a receptor, and the 5-HT transporter. Compared to 2, these heterocyclic phenol isosteres showed much better pharmacokinetic profiles as demonstrated by rat plasma levels. In sharp contrast, similar phenolic replacements in 1 decreased the binding affinity dramatically, presumably due to a conformational change of the pendant phenyl group. However, one indazole compound 33 was identified as a potent D1/D5 ligand in this series.

Basic side chains determine the pharmacology of selective estrogen receptor modulators such as tamoxifen or raloxifene. In this study we tried to turn the hormonal profile of (4R,5S)/(4S,5R)-4,5-bis(4-hydroxyphenyl)-2-imidazolines from agonistic to antagonistic by introduction of a dimethylaminoethane, a piperidin-1-ylethane, or a pyrrolidin-1-ylethane side chain into one of the 4-hydroxyphenyl rings. The compounds were tested for agonistic and antagonistic activity on hormone sensitive, ERalpha-positive MCF7-2a cells, stably transfected with the plasmid ERE(wtc)luc and on U-2 OS cells transiently transfected with plasmids encoding for ERalpha (pSG5-ERalpha) or ERbeta (pSG5-ERbeta FL) as well as the reporter plasmid (ERE)(2)luc(+). Despite the presence of a basic side chain, the majority of the 4,5-diaryl-2-imidazolines showed agonistic effects. The most active compound, (4R,5S)/(4S,5R)-4-(2-chloro-4-(2-piperidin-1-ylethoxy)phenyl)-5-(2,6-dichloro-4-hydroxyphenyl)-2-imidazoline (5a), achieved at ERalpha an EC(50) value of 0.085 microM and at ERbeta an EC(50) = 0.40 microM. High antagonistic properties only possessed the C2 ethyl substituted compounds 2a and 4a. (4R,5S)/(4S,5R)-2-Ethyl-4-(4-hydroxyphenyl)-5-(4-(2-piperidin-1-ylethoxy)phenyl)-2-imidazoline (2a) reduced the effect of estradiol at ERalpha strongly with IC(50) = 0.038 microM, while its antagonistic properties at ERbeta were distinctly lower (IC(50) = 9.00 microM), probably due to the partial agonistic effects (EC(50) = 0.50 microM).

Pathway-selective ligands for the estrogen receptor (ER) inhibit NF-kappaB-mediated inflammatory gene expression causing a reduction of cytokines, chemokines, adhesion molecules, and inflammatory enzymes. SAR development of a series of 4-(indazol-3-yl)phenols has led to the identification of WAY-169916 an orally active nonsteroidal ligand with the potential use in the treatment of rheumatoid arthritis without the classical proliferative effects associated with estrogens.

Vascular cell adhesion molecule-1 (VCAM-1) mediates recruitment of leukocytes to endothelial cells and is implicated in many inflammatory conditions. Since part of the signal transduction pathway that regulates the activation of VCAM-1 expression is redox-sensitive, compounds with antioxidant properties may have inhibitory effects on VCAM-1 expression. Novel phenolic compounds have been designed and synthesized starting from probucol (1). Many of these compounds demonstrated potent inhibitory effects on cytokine-induced VCAM-1 expression and displayed potent antioxidant effects in vitro. Some of these derivatives (4o, 4p, 4w, and 4x) inhibited lipopolysaccharide (LPS)-induced secretion of pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), and IL-6 from human peripheral blood mononuclear cells (hPBMCs) in a concentration-dependent manner in vitro and showed antiinflammatory effects in an animal model. Compounds 4ad and 4ae are currently in clinical trials for the treatment of rheumatoid arthritis (RA) and prevention of chronic organ transplant rejection, respectively.

New diphenolic azoles as highly selective estrogen receptor-beta agonists are reported. The more potent and selective analogues of these series have comparable binding affinities for ERbeta as the natural ligand 17beta-estradiol but are >100-fold selective over ERalpha. Our design strategy not only followed a traditional SAR approach but also was supported by X-ray structures of ERbeta cocrystallized with various ligands as well as molecular modeling studies. These strategies enabled us to take advantage of a single conservative residue substitution in the ligand-binding pocket, ERalpha Met(421) --> ERbeta Ile(373), to optimize ERbeta selectivity. The 7-position-substituted benzoxazoles (Table 5) were the most selective ligands of both azole series, with ERB-041 (117) being >200-fold selective for ERbeta. The majority of ERbeta selective agonists tested that were at least approximately 50-fold selective displayed a consistent in vivo profile: they were inactive in several models of classic estrogen action (uterotrophic, osteopenia, and vasomotor instability models) and yet were active in the HLA-B27 transgenic rat model of inflammatory bowel disease. These data suggest that ERbeta-selective agonists are devoid of classic estrogenic effects and may offer a novel therapy to treat certain inflammatory conditions.

[1-(3,5-Difluoro-4-hydroxyphenyl)-1H-pyrrol-3-yl]phenylmethanone (6) was synthesized as a putative bioisostere of the known aldose reductase (AR) inhibitor (3-benzoylpyrrol-1-yl)acetic acid (I). It was found that 6 is approximately 5 times more potent as an in vitro inhibitor of AR than I, with an IC(50) value in the submicromolar range. Furthermore, 6 showed considerable activity in an in vitro experimental glycation model of diabetes mellitus. Our results support the notion that 6 might become a useful lead structure.

Several phenolic acids-caffeic and gallic acid derivatives-were synthesized and screened for their potential antiproliferative and cytotoxic properties, in different human cancer cell lines: mammary gland and cervix adenocarcinomas and lymphoblastic leukemia. The selected phenols were structurally related, which allowed us to gather important information regarding the structure-activity relationships underlying the biological activity of such compounds. This is proposed to be due to a balance between the antioxidant and pro-oxidant properties of this kind of agent. Distinct effects were found for different cell lines, which points to a significant specificity of action of the drugs tested. It was verified, for the types of cancer investigated, that the trihydroxylated derivatives yielded better results than the dihydroxylated ones. Tests in noncancerous cells, human lung fibroblasts, were also undertaken, in view of determining the toxic side effects of the compounds studied.

4,4'-dihydroxybenzophenone-2,4-dinitrophenylhydrazone (A-007) has recently completed a phase I clinical trial in advanced cancer with minimal toxicity, and impressive objective responses were noted. A-007 possesses three moieties that appear to have an influence on its anticancer activities: diphenylmethane, hydrazone, and dinitrophenyl. The goals of this study were to modify A-007's chemical moieties with the ultimate goal of maximizing its anticancer activity through increased planarity and introduction of functional groups. Thirty-five phenylhydrazone analogues of A-007 were synthesized and evaluated in vitro in a human primary cancer explant assay. Anticancer activities for selected analogues were also assayed for activity vs established human/murine cell lines. One-hundred-eighty-six fresh human solid tumors were used to screen for anticancer activity. Selected analogues were assayed for therapeutic indices (vs GM-CFC from bone marrow) in preparation for preclinical studies. Several polyaryl phenylhydrazones demonstrated improved cytotoxic activities by factors of 10(2)-10(3) when compared with A-007. However, the polyaryl quinone moieties of the latter analogues introduced potential toxic properties (cardiac, hematological) that do not exist with A-007.

C2-Alkyl substituted derivatives of the 1,1,2-tris(4-hydroxyphenyl)ethene 3a (alkyl = Me (3b), Et (3c), Prop (3d), But (3e)) were synthesized by reaction of 1,2-bis(4-methoxyphenyl)ethanone with the appropriate alkyl halide, followed by a Grignard reaction with 4-methoxyphenylmagnesium bromide, dehydration with phosphoric acid or hydrobromic acid, and ether cleavage with BBr(3). The compounds were tested for estrogen receptor (ER) binding affinity in a competition experiment with radio labeled estradiol ([(3)H]-E2) and for gene activation on the ER-positive MCF-7-2a cell line. All compounds showed high receptor binding affinity (RBA-value: 3b (52.1%) > 3a (45.5%) > 3c (29.6%) > 3d (4.03%) > 3e (0.95%)). The tests on hormone dependent MCF-7-2a breast cancer cells, stably transfected with the plasmid ERE(wtc)luc, revealed that all 1,1,2-tris(4-hydroxyphenyl)ethenes antagonized the effect of 1 nM estradiol (E2). The compounds 3b (IC(50) = 15 nM) and 3c (IC(50) = 10 nM) were equal in their effects to 4-hydroxytamoxifen (4OHT) (IC(50) = 7 nM). Agonistic effects were low. Only 3a and 3b activated the luciferase expression (relative activation at 1 microM: 3a 60%; 3b 35%). Despite their highly antagonistic potency, the 1,1,2-tris(4-hydroxyphenyl)ethenes showed only low cytotoxic properties on the hormone sensitive MCF-7 cell line.

Five nonnatural beta-C-nucleoside 5'-triphosphates bearing a 3,4-dihydroxyphenyl (1TP), a 2-hydroxyphenyl (2TP), a 3-hydroxyphenyl (3TP), a 4-hydroxyphenyl (4TP), or a phenyl (5TP) group were synthesized, and their structure-activity relationships were examined for a series of DNA polymerase reactions in vitro under typical polymerase chain reaction conditions. We found that the 5'-triphosphates (1TP-5TP) are not incorporated into DNA strands but inhibit the DNA polymerase reactions in the presence of natural nucleoside 5'-triphosphates (dNTPs). 1TP having two phenolic hydroxy groups at the nucleobase moiety showed the most potent inhibitory effect against DNA synthesis by Ex Taq polymerase (IC(50) = 30 microM). The competition assay indicated that 1TP and dNTPs are most likely to affect DNA polymerase reactions competitively. This finding may raise the appealing possibility that artificial nucleoside 5'-triphosphates having phenolic hydroxy groups could exhibit potent inhibitory activity against DNA-directed enzymatic reactions.

A series of conformationally restricted analogues were synthesized in order to elucidate the possible effects of different amide conformations of lavendustin A derivatives on cytotoxicity in cancer cell cultures and on inhibition of tubulin polymerization. The conformationally restricted analogues were based on the oxazinedione and isoindolone ring systems. In addition, the amide bond was replaced by both cis and trans alkene moieties. Surprisingly, the results indicated very little effect of conformational restriction on biological activity. Because all of the compounds synthesized had similar cytotoxicities and potencies as tubulin polymerization inhibitors, the side chain present on the aniline ring system does not appear to be important in the biological effects of the lavendustins. The hydroquinone ring of lavendustin A may be a more important determinant of the biological activity than the structure surrounding the aniline ring.

A variety of nonsteroidal systems can function as ligands for the estrogen receptor (ER), in some cases showing selectivity for one of the two ER subtypes, ER alpha or ER beta. We have prepared a series of heterocycle-based (furans, thiophenes, and pyrroles) ligands for the estrogen receptor and assessed their behavior as ER ligands. An aldehyde enone conjugate addition approach and an enolate alkylation approach were developed to prepare the 1,4-dione systems that were precursors to the trisubstituted and tetrasubstituted systems, respectively. All of the diones were easily converted into the corresponding furans, but formation of the thiophenes and pyrroles from the more highly substituted 1,4-diones was problematical. Of the systems investigated, the tetrasubstituted furans proved to be most interesting. They were ER alpha binding- and potency-selective agents, with the triphenolic 3-alkyl-2,4,5-tris(4-hydroxyphenyl)furans (15a-d) displaying generally higher subtype binding selectivity than the bisphenolic analogues (15f-i). Binding selectivity for ER alpha was as high as 50-70-fold, and transcriptional activation studies showed that several members of this series were ER alpha selective agonists, with the best compound [3-ethyl-2,4,5-tris(4-hydroxyphenyl)furan, 15b] having full transcriptional activity on ER alpha while being inactive on ER beta. Comparative binding affinity analysis and molecular modeling were used to investigate the preferred binding mode adopted by the furan ligands, which appears to have the C(2) phenol mimicking the important role of the A-ring of estradiol. These ligands should be useful in studying the biological roles of both ER alpha and ER beta, and they might form the basis for the development of novel estrogen pharmaceuticals.

In the search for a novel water-soluble general anesthetic agent the activity of an alpha-amino acid phenolic ester lead, identified from patent literature, was markedly improved. In addition to improving in vivo activity in mice, good in vitro activity at GABA(A) receptors was also conferred. Within the series of compounds good enantioselectivity for both in vitro and in vivo activity was found, supporting a protein-mediated mechanism of action for anesthesia involving allosteric modulation of GABA(A) receptors. alpha-Amino acid phenolic ester 19, as the hydrobromide salt Org 25435, was selected for clinical evaluation since it retained the best overall anesthetic profile coupled with improved stability and water solubility. In the clinic it proved to be an effective intravenous anesthetic in man with rapid onset of and recovery from anesthesia at doses of 3 and 4 mg/kg.

A series of N-alkylamide analogues of the lavendustin A pharmacophore were synthesized and tested for inhibition of the epidermal growth factor receptor (EGFR) protein tyrosine kinase and the nonreceptor protein tyrosine kinase Syk. Although several compounds in the series were effective inhibitors of both kinases, it seemed questionable whether their inhibitory effects on these kinases were responsible for the cytotoxic properties observed in a variety of human cancer cell cultures. Accordingly, a COMPARE analysis of the cytotoxicity profile of the most cytotoxic member of the series was performed, and the results indicated that its cytotoxicity profile was similar to that of antitubulin agents. This mechanism of action was supported by demonstrating that most compounds in the series were moderately effective as inhibitors of tubulin polymerization. This suggests that the lavendustin A analogues reported here, as well as some of the previously reported lavendustin A analogues, may be acting as cytotoxic agents by a mechanism involving the inhibition of tubulin polymerization.

We have found that certain tetrasubstituted pyrazoles are high-affinity ligands for the estrogen receptor (ER) (Fink et al. Chem. Biol. 1999, 6, 205-219) and that one pyrazole is considerably more potent as an agonist on the ERalpha than on the ERbeta subtype (Sun et al. Endocrinology 1999, 140, 800-804). To investigate what substituent pattern provides optimal ER binding affinity and the greatest enhancement of potency as an ERalpha-selective agonist, we prepared a number of tetrasubstituted pyrazole analogues with defined variations at certain substituent positions. Analysis of their binding affinity pattern shows that a C(4)-propyl substituent is optimal and that a p-hydroxyl group on the N(1)-phenyl group also enhances affinity and selectivity for ERalpha. The best compound in this series, a propylpyrazole triol (PPT, compound 4g), binds to ERalpha with high affinity (ca. 50% that of estradiol), and it has a 410-fold binding affinity preference for ERalpha. It also activates gene transcription only through ERalpha. Thus, this compound represents the first ERalpha-specific agonist. We investigated the molecular basis for the exceptional ERalpha binding affinity and potency selectivity of pyrazole 4g by a further study of structure-affinity relationships in this series and by molecular modeling. These investigations suggest that the pyrazole triols prefer to bind to ERalpha with their C(3)-phenol in the estradiol A-ring binding pocket and that binding selectivity results from differences in the interaction of the pyrazole core and C(4)-propyl group with portions of the receptor where ERalpha has a smaller residue than ERbeta. These ER subtype-specific interactions and the ER subtype-selective ligands that can be derived from them should prove useful in defining those biological activities in estrogen target cells that can be selectively activated through ERalpha.

The activity of new anti-HSV-1 chemical structures, designed by virtual combinatorial chemical synthesis and selected by a computational screening, is determined by an in vitro assay. A virtual library of phenol esters and anilides was formed from two databases of building blocks: one with carbonyl fragments and the other containing both substituted phenoxy and phenylamino fragments. The library of virtually assembled compounds was computationally screened, and those compounds which were selected by our mathematical model as active ones were finally synthesized and tested. Our antiviral activity model is a "tandem" of four linear functions of topological graph-theoretical descriptors. A given chemical structure was selected as active if it satisfies every discriminant equation in that model. The final result was that five new structures were selected, synthesized, and tested: all of them demonstrated activity, and three showed appreciable anti-HSV-1 activity, with IC(50) values of 0.9 microM. The same model, applied to a database of known compounds, has identified the anti-herpes activity of the following compounds: 3,5-dimethyl-4-nitroisoxazole, nitrofurantoin, 1-(pyrrolidinocarbonylmethyl)piperazine, nebularine, cordycepin, adipic acid, thymidine, alpha-thymidine, inosine, 2, 4-diamino-6-(hydroxymethyl)pteridine, 7-(carboxymethoxy)-4-methylcoumarin, 5-methylcytidine, and others that showed less activity.

CP-060 (1), 2-(3, 5-di-tert-butyl-4-hydroxyphenyl)-3-[3-[N-methyl-N-[2-[3, 4-(methylenedioxy)phenoxy]ethyl]amino]propyl]-1,3-thiazolidin-4-on e, is a novel type of Ca(2+) antagonist possessing both Ca(2+) overload inhibition and antioxidant activity. The structure-activity relationships for this series of compounds were studied by synthesizing the analogues and evaluating these three kinds of activity. Ca(2+) antagonistic activity was largely determined by the lipophilicity of the phenyl group at the 2-position and the length of the alkyl chains. As for the antioxidant activity, it was demonstrated that the phenolic hydroxyl group is an essential structural element. Compounds with potent activity were evaluated for their effect on the coronary blood flow in vivo. Among these compounds, compound 1 was shown to be the most potent. Furthermore, the enantiomers of 1 were resolved by high-performance liquid chromatography with a chiral column. Compound (-)-1 showed about 10 times higher Ca(2+) antagonistic activity than (+)-1, though both enantiomers had similar potency in Ca(2+) overload inhibition and antioxidant activity. An X-ray crystal structure determination of (-)-1 hydrogen fumarate identified (-)-1 as having S configuration at the 2-position.

A search of our compound library for compounds with structural similarity to ifenprodil (5) and haloperidol (7) followed by in vitro screening revealed that 4-benzyl-1-(4-phenyl-3-butynyl)piperidine (8) was a moderately potent and selective antagonist of the NR1A/2B subtype of NMDA receptors. Substitution on the benzyl group of 8 did not significantly affect NR1A/2B potency, while addition of hydrogen bond donors in the para position of the phenyl group enhanced NR1A/2B potency. Addition of a hydroxyl moiety to the 4-position of the piperidine group slightly reduced NR1A/2B potency while reducing alpha-1 adrenergic and dopamine D2 receptor binding affinities substantially, resulting in improved overall selectivity for NR1A/2B receptors. Finally, the butynyl linker was replaced with propynyl or pentynyl. When the phenyl was para substituted with amine or acetamide groups, the NR1A/2B potency order was butynyl > pentynyl >> propynyl. For the para methanesulfonamide or hydroxyl groups, the order was butynyl approximately propynyl > pentynyl. The hydroxyl propyne (48) and butyne (23) were among the most potent NR1A/2B antagonists from this study. They both potentiated the effects of L-DOPA in the 6-hydroxydopamine-lesioned rat, a model of Parkinson's disease, dosed at 10 mg/kg ip, but 48 was not active at 30 mg/kg po.