Benzimidazolyl-pyrazolo[3,4-]pyridinones, Selective Inhibitors of MOLT-4 Leukemia Cell Growth and Sea Urchin Embryo Spiculogenesis: Target Quest.

1,3-Substituted pyrazolo[3,4-]pyridinones - were synthesized by a three-component condensation of Meldrum's acid with aryl aldehydes and 1,3-substituted 5-aminopyrazoles. Their biological activity was evaluated using the phenotypic sea urchin embryo assay and the cytotoxicity screen against human cancer cell lines. In the sea urchin embryo model, 1-benzimidazolyl-pyrazolo[3,4-]pyridinones caused inhibition of hatching and spiculogenesis at sub-micromolar concentrations.

Sea Urchin Embryo Model As a Reliable in Vivo Phenotypic Screen to Characterize Selective Antimitotic Molecules. Comparative evaluation of Combretapyrazoles, -isoxazoles, -1,2,3-triazoles, and -pyrroles as Tubulin-Binding Agents.

A series of both novel and reported combretastatin analogues, including diarylpyrazoles, -isoxazoles, -1,2,3-triazoles, and -pyrroles, were synthesized via improved protocols to evaluate their antimitotic antitubulin activity using in vivo sea urchin embryo assay and a panel of human cancer cells. A systematic comparative structure-activity relationship studies of these compounds were conducted. Pyrazoles 1i and 1p, isoxazole 3a, and triazole 7b were found to be the most potent antimitotics across all tested compounds causing cleavage alteration of the sea urchin embryo at 1, 0.

Synthesis and anti-mitotic activity of 6,7-dihydro-4H-isothiazolo[4,5-b]pyridin-5-ones: In vivo and cell-based studies.

A series of 3,7-diaryl-6,7-dihydroisothiazolo [4,5-b]pyridin-5(4H)-ones 8 and 9 was synthesized by multicomponent condensation of 3-aryl-5-isothiazolecarboxylic acid esters 4a-f with aromatic (or thienyl) aldehydes 7 and Meldrum's acid in an acidic medium. The targeted compounds were evaluated for their antimitotic microtubule destabilizing activity using in vivo phenotypic sea urchin embryo model and in vitro human cancer cell-based assays. Selected dihydroisothiazolopyridinones altered sea urchin egg cleavage in 2-10 nM concentrations together with significant cytotoxicity against cancer cells including chemoresistant cell lines (IC in submicromolar - low nanomolar concentration range).

A facile synthesis and microtubule-destabilizing properties of 4-(1H-benzo[d]imidazol-2-yl)-furazan-3-amines.

A series of 4-(1H-benzo[d]imidazol-2-yl)-furazan-3-amines (BIFAs) were prepared in good yields (60-90% for each reaction step) via a novel procedure from aminofurazanyl hydroximoyl chlorides and o-diaminobenzenes. The synthetic sequence was run under mild reaction conditions, it was robust and did not require extensive purification of intermediates or final products. Furthermore, there was no need for protection of reactive moieties allowing for the parallel synthesis of diverse BIFA derivatives.

Comparative in vivo evaluation of polyalkoxy substituted 4H-chromenes and oxa-podophyllotoxins as microtubule destabilizing agents in the phenotypic sea urchin embryo assay.

A series of polyalkoxy substituted 7-hydroxy- and 7-methoxy-4-aryl-4H-chromenes were evaluated using the sea urchin embryo model to yield several compounds exhibiting potent antimitotic microtubule destabilizing activity. Data obtained by the assay were further confirmed in the NCI60 human cancer cell screen. The replacement of methylenedioxy ring A and lactone ring D in podophyllotoxin analogues by 7-methoxy, 2-NH2, and 3-CN groups in 4-aryl-4H-chromenes resulted in potent antimitotic microtubule destabilizing agents.

3-(5-)-Amino-o-diarylisoxazoles: regioselective synthesis and antitubulin activity.

A regioselective synthesis of both 5-amino- and 3-aminodiarylisoxazoles substituted with polyalkoxyaryl pharmacophores has been validated. Starting materials for the synthetic scheme were easily available from plant extracts. The targeted molecules were further tested in the phenotypic sea urchin embryo assay to identify compounds with antimitotic microtubule destabilizing activity.

Synthesis and antiproliferative activity of conformationally restricted 1,2,3-triazole analogues of combretastatins in the sea urchin embryo model and against human cancer cell lines.

A series of 1,5-diaryl- and 4,5-diaryl-1,2,3-triazole derivatives of combretastatin A4 were synthesized and evaluated as antimitotic microtubule destabilizing agents using the sea urchin embryo model. Structure-activity relationship studies identified compounds substituted with 3,4,5-trimethoxyphenyl and 3,4-methylenedioxy-5-methoxyphenyl ring A and 4-methoxyphenyl ring B as potent antiproliferative agents with high cytotoxicity against a panel of human cancer cell lines including multi-drug resistant cells. 4,5-Diaryl-1,2,3-triazoles (C-C geometry) were found to be considerably more active than the respective 1,5-diaryl-1,2,3-triazoles (N-C geometry).

cis-Restricted 3-aminopyrazole analogues of combretastatins: synthesis from plant polyalkoxybenzenes and biological evaluation in the cytotoxicity and phenotypic sea urchin embryo assays.

We have synthesized a series of novel cis-restricted 4,5-polyalkoxydiaryl-3-aminopyrazole analogues of combretastatins via short synthetic sequences using building blocks isolated from dill and parsley seed extracts. The resulting compounds were tested in vivo in the phenotypic sea urchin embryo assay to reveal their antimitotic and antitubulin effects. The most potent aminopyrazole, 14a, altered embryonic cell division at 10 nM concentration, exhibiting microtubule-destabilizing properties.

Synthesis and comparative evaluation of 4-oxa- and 4-aza-podophyllotoxins as antiproliferative microtubule destabilizing agents.

A series of novel 4-oxa-podophyllotoxin derivatives 7 featuring the intact lactone ring D and various substituents in rings B and E has been synthesized and evaluated in a phenotypic sea urchin embryo assay along with the representative 4-aza-analogs 5 for their antimitotic and microtubule destabilizing activity. The most active compounds exhibited myristicin-derived or a 3',5'-dimethoxy substitution pattern in the ring E and a 6-methoxy moiety replacing the methylenedioxy ring A. Compounds 5xb, 5xe, 5yb, 7xa, 7xb, and 7xc showed potent antiproliferative effects in the NCI60 cytotoxicity screen.

Polyalkoxybenzenes from plants. 5. Parsley seed extract in synthesis of azapodophyllotoxins featuring strong tubulin destabilizing activity in the sea urchin embryo and cell culture assays.

A series of 4-azapodophyllotoxin derivatives with modified rings B and E have been synthesized using allylpolyalkoxybenzenes from parsley seed oil. The targeted molecules were evaluated in vivo in a phenotypic sea urchin embryo assay for antimitotic and tubulin destabilizing activity. The most active compounds identified by the in vivo sea urchin embryo assay featured myristicin-derived ring E.

1,7-Disubstituted oxyindoles are potent and selective EP(3) receptor antagonists.

A series of novel 1,7-disubstituted oxyindoles were shown to be potent and selective EP(3) receptor antagonists. Variation of substitution pattern at the C-3 position of indole enhanced in vitro metabolic stability of the resulting derivatives. Series 27a-c showed >1000-fold selectivity over a panel of prostanoid receptors including IP, FP, EP(1), EP(2) and EP(4).

Novel derivatives of 1,3,4-oxadiazoles are potent mitostatic agents featuring strong microtubule depolymerizing activity in the sea urchin embryo and cell culture assays.

A series of novel 1,3,4-oxadiazole derivatives based on structural and electronic overlap with combretastatins have been designed and synthesized. Initially, we tested all new compounds in vivo using the phenotypic sea urchin embryo assay to yield a number of agents with anti-proliferative, anti-mitotic, and microtubule destabilizing activities. The experimental data led to identification of 1,3,4-oxadiazole derivatives with isothiazole (5-8) and phenyl (9-12) pharmacophores featuring activity profiles comparable to that of combretastatins, podophyllotoxin and nocodazole.

Design of phosphodiesterase 4D (PDE4D) allosteric modulators for enhancing cognition with improved safety.

Phosphodiesterase 4 (PDE4), the primary cAMP-hydrolyzing enzyme in cells, is a promising drug target for a wide range of conditions. Here we present seven co-crystal structures of PDE4 and bound inhibitors that show the regulatory domain closed across the active site, thereby revealing the structural basis of PDE4 regulation. This structural insight, together with supporting mutagenesis and kinetic studies, allowed us to design small-molecule allosteric modulators of PDE4D that do not completely inhibit enzymatic activity (I(max) approximately 80-90%).

Structure-activity relationship studies leading to the identification of (2E)-3-[l-[(2,4-dichlorophenyl)methyl]-5-fluoro-3-methyl-lH-indol-7-yl]-N-[(4,5-dichloro-2-thienyl)sulfonyl]-2-propenamide (DG-041), a potent and selective prostanoid EP3 receptor antagonist, as a novel antiplatelet agent that does not prolong bleeding.

The EP(3) receptor on the platelet mediates prostaglandin E(2) potentiation of thrombogenic coagonists including collagen and adenosine diphosphate (ADP). A pharmacophore driven approach led to the identification of diverse peri-substituted heterocycles as potent and selective EP(3) receptor antagonists. A simultaneous chemical optimization and druglike assessment of prioritized molecules converged on a lead compound 50 (DG-041) that displayed favorable in vitro and functional activities as an inhibitor of human platelet aggregation.

Discovery of 4-[(2S)-2-{[4-(4-chlorophenoxy)phenoxy]methyl}-1-pyrrolidinyl]butanoic acid (DG-051) as a novel leukotriene A4 hydrolase inhibitor of leukotriene B4 biosynthesis.

Both in-house human genetic and literature data have converged on the identification of leukotriene 4 hydrolase (LTA(4)H) as a key target for the treatment of cardiovascular disease. We combined fragment-based crystallography screening with an iterative medicinal chemistry effort to optimize inhibitors of LTA(4)H. Ligand efficiency was followed throughout our structure-activity studies.

Heterocyclic 1,7-disubstituted indole sulfonamides are potent and selective human EP3 receptor antagonists.

We have developed a pharmacophore model for the EP(3) receptor antagonists based on its endogenous ligand PGE(2). This ligand-based design yielded a series of novel peri-substituted [4.3.

Synthesis and structural assignment of two major metabolites of the LTA4H inhibitor DG-051.

The same two major CYP mediated metabolites of DG-051 were produced in the presence of rat, dog, monkey and human liver microsomes. Their respective structures were hypothesized based on mass spectrometry data correlated with the parent structure and confirmed by comparison with authentic synthetic samples. The number of regioisomers synthesized as candidates for metabolite M1 was narrowed down using a metabolic study of a selectively deuterated DG-051 analogue.

Discovery of leukotriene A4 hydrolase inhibitors using metabolomics biased fragment crystallography.

We describe a novel fragment library termed fragments of life (FOL) for structure-based drug discovery. The FOL library includes natural small molecules of life, derivatives thereof, and biaryl protein architecture mimetics. The choice of fragments facilitates the interrogation of protein active sites, allosteric binding sites, and protein-protein interaction surfaces for fragment binding.

3-Acrylamide-4-aryloxyindoles: synthesis, biological evaluation and metabolic stability of potent and selective EP3 receptor antagonists.

A series of potent and selective EP(3) receptor antagonists are described. Utilizing a pharmacophore model developed for the EP(3) receptor, a series of 3,4-disubstituted indoles were found to be efficient ligands for this target. These compounds showed high selectivity over IP, FP and other EP receptors.

Peri-substituted hexahydro-indolones as novel, potent and selective human EP3 receptor antagonists.

A series of peri-substituted [4.3.0] bicyclic non-aromatic heterocycles have been identified as potent and selective hEP(3) receptor antagonists.

3,4-Disubstituted indole acylsulfonamides: a novel series of potent and selective human EP3 receptor antagonists.

A series of potent and selective EP(3) receptor antagonists are described. Utilizing a pharmacophore model developed for the EP(3) receptor, a series of 3,4-disubstituted indoles were shown to be high affinity ligands for this target. These compounds showed high selectivity over IP, FP and other EP receptors and are potent antagonists in functional assays.

Regulators of chemokine receptor activity as promising anticancer therapeutics.

Chemokines are a family of small proteins inducing directed cell migration via specific chemokine receptors, which play important roles in a variety of biological and pathological processes. Their respective ligands act as proinflammatory mediators that primarily control leukocyte migration into selected tissues and upregulation of adhesion receptors, and also have a role in pathological conditions that require neovascularization. Therapeutic strategies based on modulation of chemokine receptor pathways were reported to be promising clinical strategies in the treatment of inflammatory diseases and viral infections.

Novel mitotic targets and their small-molecule inhibitors.

With several successful anticancer drugs on the market and numerous compounds in clinical developments, antimitotic agents represent an important category of anticancer agents. However, clinical utility of the tubulin-binding agents is somewhat limited due to multiple drug resistance (MDR), poor pharmacokinetics and therapeutic index. There is ongoing need for the modulators of other intracellular targets that result in the same anti-mitotic effect without adverse effects of "traditional" tubulin binders.