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Vancomycin analogues bearing an A-ring trimethylammonium salt modification were synthesized and their antimicrobial activity against vancomycin-resistant Enterococci (VRE) was evaluated. The modification increased antimicrobial potency and provided the capability to induce bacteria cell membrane permeabilization, but both properties were weaker than that found with our earlier reported similar C-terminus modification. The results provide further insights on the additive effect and generalizability of the structural and site-specific nature of a peripheral quaternary trimethylammonium salt modification of vancomycin. Image 1 [ABSTRACT FROM AUTHOR]

In recent efforts, several C20′ urea vinblastine analogues were discovered that displayed remarkable potency against vinblastine-sensitive tumor cell lines (IC 50 50–75 pM), being roughly 100-fold more potent than vinblastine, and that exhibited decreased susceptibility to Pgp efflux-derived resistance in a vinblastine-resistant cell line. Their extraordinary activity indicate that it is not likely or even possible that their cellular functional activity is derived from stoichiometric occupancy of the intracellular tubulin binding sites. Rather, their potency indicates sub-stoichiometric or even catalytic occupancy of candidate binding sites may be sufficient to disrupt tubulin dynamics or microtubule assembly during mitosis. We detail efforts to delineate the underlying behavior responsible for the increased potency and show that the ultra-potent extended C20′ ureas retain the mechanistic behavior of vinblastine, display enhanced affinity for tubulin and on-target activity approximately 100-fold both in vitro and in HeLa cells, but do not show evidence of catalytic disassembly of microtubulin. We also use the analogues to show that, in live interphase cells, the effects of the vinblastine class of drugs do not display a catastrophic effect on the microtubule skeleton, but rather a subtler insult to its dynamicity, acting as sub-stoichiometric drugs that inhibit normal microtubulin maturation and dynamics. [ABSTRACT FROM AUTHOR]

New treatment modalities for glioblastoma multiforme (GBM) are urgently needed. Proton therapy is considered one of the most effective forms of radiation therapy for GBM. DNA alkylating agents such as temozolomide (TMZ) are known to increase the radiosensitivity of GBM to photon radiation. TMZ is a fairly impotent agent, while duocarmycin SA (DSA) is an extremely potent cytotoxic agent capable of inducing a sequence-selective alkylation of duplex DNA. Here, the effects of sub-nM concentrations of DSA on the radiosensitivity of a human GBM cell line (U-138) to proton irradiation were examined. Radiation sensitivity was determined by viability, apoptosis, necrosis and clonogenic assays. DSA concentrations as low as 0.001 nM significantly sensitized U-138 cells to proton irradiation. DSA demonstrates synergistic cytotoxicity against GBM cells treated with proton radiation in vitro, which may represent a novel therapeutic alternative for the treatment of GBM. [ABSTRACT FROM AUTHOR]

The total synthesis of (−)-kopsinine and its unnatural enantiomer is detailed, enlisting a late-stage SmI 2 -mediated transannular free radical conjugate addition reaction for construction of the core bicyclo[2.2.2]octane ring system with strategic C21–C2 bond formation. Key to the approach is assemblage of the underlying skeleton by an intramolecular [4+2]/[3+2] cycloaddition cascade of a 1,3,4-oxadiazole that provided the precursor C21 functionalized pentacyclic ring system 1 in a single step in which the C3 methyl ester found in the natural product served as a key 1,3,4-oxadiazole substituent, activating it for participation in the initiating Diels–Alder reaction and stabilizing the intermediate 1,3-dipole. [ABSTRACT FROM AUTHOR]

Clinical association studies have implicated high expression of class III β-tubulin as a predictive factor for lower response rates and reduced overall survival in patients receiving tubulin binding drugs, most notably the taxanes. Because of the implications, we examined a series of key vinblastine analogs that emerged from our studies in functional cell growth inhibition assays for their sensitivity to high expression of class III β-tubulin (human non-small cell lung cancer cell line A549 vs taxol-resistant A549-T24). Unlike taxol, vinblastine and a set of key analogs 3 – 10 did not exhibit any loss in sensitivity toward A549-T24. The results suggest that vinblastine and related analogs are not likely prone to resistance derived from high expression of class III β-tubulin unlike the taxanes. Most significant are the results with 4 – 6 , a subset of 20′ amide vinblastine analogs. They match or exceed the potency of vinblastine and they display more potent activity against taxol-resistant A549-T24 than even wild type A549 cells (1.2–2-fold), complementing our prior observations that they also display no sensitivity to overexpression of Pgp (HCT116/VM46 vs HCT116) and are not subject to resistance derived from Pgp efflux. [ABSTRACT FROM AUTHOR]

Abstract: An efficient and convergent solution-phase synthesis of the cyclic peptide of the natural product chlorofusin, a reported inhibitor of the MDM2–p53 interaction, is detailed. [Copyright &y& Elsevier]

The synthesis of the 15-membered biaryl ring system constituting an appropriately functionalized AB ring system of RP 66453 is detailed. [Copyright &y& Elsevier]

A short personal perspective on the development of an approach to the solution-phase synthesis of combinatorial libraries for modulating cellular signaling by inhibiting, promoting, or mimicking protein–protein or protein–DNA interactions is provided. [Copyright &y& Elsevier]

A total synthesis of (+)-camptothecin is described. The approach is based on a room temperature LUMOdiene-controlled Diels–Alder cycloaddition of the electron deficient N-sulfonyl-1-aza-1,3-butadiene 6 with the electron rich dienophile 1,1,3,3-tetraethoxypropene (7) for assembly of a pyridine precursor to the D-ring pyridone. The 20(S) tertiary alcohol was installed through a Sharpless asymmetric dihydroxylation reaction on the methyl vinyl ether 12, using the 3,4,5-trimethoxyphenyl-derived pyrimidine DHQ dimer ligand 16. In a single reaction vessel, the C and E rings were closed using an acid-catalyzed deprotection of the benzylic ethers to afford the corresponding benzylic bromides (18) which underwent intramolecular nucleophilic displacement by the carboxylate and pyridone nitrogen to furnish (+)-camptothecin. [Copyright &y& Elsevier]

The rates of DNA alkylation were established for the reaction of (+)-duocarmycin SA (1) with the native duplex d(G1TCAATTAGTC11)⋅d(G12ACTAATTGAC22), an 11 bp deoxyoligonucleotide that contains a single high-affinity alkylation site that has been structurally characterized at exquisite resolution, and modified duplexes in which the four backbone phosphates proximal to the C4 carbonyl of bound 1 were replaced with methylphosphonates. All were found to react at comparable rates establishing that these backbone phosphates do not participate in catalysis of the DNA alkylation reaction. [Copyright &y& Elsevier]

Abstract: A vancomycin aglycon analogue that possesses a reduced C-ring and an intact E-ring chloride was prepared and its antimicrobial activity towards Staphylococcus aureus and binding affinity to model cell wall ligands were established. Comparison of the derivative with a series of vancomycin aglycon analogues that possess and lack the chloro substituents on the aryl C- and E-rings defines the impact and further refines the role the C-ring chloride plays in promoting both target binding affinity and binding selectivity for d-Ala-d-Ala and its impact on antimicrobial activity. [Copyright &y& Elsevier]

The preparation, characterization, and examination of the CBI-based 5-membered lactone 5 capable of serving as a prodrug or protein (antibody) conjugation reagent are disclosed along with its incorporation into the corresponding CC-1065 and duocarmycin analog 6 , and the establishment of their properties. [ABSTRACT FROM AUTHOR]

Abstract: A series of α-ketooxazoles containing heteroatoms embedded within conformational constraints in the C2 acyl side chain of 2 (OL-135) were synthesized and evaluated as inhibitors of fatty acid amide hydrolase (FAAH). The studies reveal that the installation of a heteroatom (O) in the conformational constraint is achievable, although the potency of these novel derivatives is reduced slightly relative to 2 and the analogous 1,2,3,4-tetrahydronaphthalene series. Interestingly, both enantiomers (R and S) of the candidate inhibitors bearing a chiral center adjacent to the electrophilic carbonyl were found to effectively inhibit FAAH. [Copyright &y& Elsevier]

Abstract: A summary of the discovery and advancement of inhibitors of fatty acid amide hydrolase (FAAH) is presented. [Copyright &y& Elsevier]

Abstract: Key studies leading to the discovery and definition of the role of endogenous fatty acid amide signaling molecules are summarized. [Copyright &y& Elsevier]

Abstract: Determining the sequence specifity of DNA binding molecules is a non-trivial task. Here we describe the development of a platform for assaying the sequence specificity of DNA ligands using label free detection on high density DNA microarrays. This is achieved by combining Cognate Site Identification (CSI) with Fluorescence Intercalation Displacement (FID) to create CSI–FID. We use the well-studied small molecule DNA ligand netropsin to develop this high throughput platform. Analysis of the DNA binding properties of protein- and small molecule-based libraries with CSI–FID will advance the development of genome-anchored molecules for therapeutic purposes. [Copyright &y& Elsevier]

The binding affinities of several triplex forming oligonucleotides were determined using a fluorescent intercalator displacement (FID) assay. [Copyright &y& Elsevier]

Four hairpin polyamides bearing subtle N- and C-terminal substitutions were examined in a fluorescent intercalator displacement (FID) assay enlisting a library of 512 DNA hairpins that contain all possible five base pair sequences in a challenging probe of its capabilities for establishing DNA binding sequence selectivity. Not only did the assay define the global sequence selectivity expected based on known structural interactions and Dervan''s pairing rules establishing the utility of the method for characterizing such polyamides, but previously unappreciated subtle substituent effects on global sequence selectivity were also revealed. Thus, we report the discovery of a novel five base pair high affinity binding site of the form 5′-WWCWW (vs 5′-WGWWW) for the polyamide ImPyPy-γ-PyPyPy-β-Dp and its structural basis. [Copyright &y& Elsevier]

A concise total synthesis of (−)-4-desacetoxy-1-oxovindoline is disclosed, bearing a single heteroatom exchange in the core structure of the natural product 4-desacetoxyvindoline. Central to the synthesis is powerful oxadiazole intramolecular [4 + 2]/[3 + 2] cycloaddition cascade that formed four C–C bonds, created three new rings, and established five contiguous stereocenters about the new formed central 6-membered ring. [Display omitted] [ABSTRACT FROM AUTHOR]

Protein titration displacement of ethidium bromide bound to hairpin deoxyoligonucleotides containing any sequence of interest provides a well-defined titration curve (measuring the loss of fluorescence derived from the DNA bound ethidium bromide) that provides both absolute binding constants (Ka) and stoichiometry of binding. This use of a fluorescent intercalator displacement (FID) assay for establishing protein DNA binding affinity and selectivity is demonstrated with the examination of the LEF-1 HMG domain binding to hairpin deoxyoligonucleotides containing its commonly accepted consensus sequence 5′-CTTTGWW (W=A or T) and those modified (5′-CTNTGWW) to examine sequences implicated in early studies (5′-CTNTG). The effectiveness of the FID assay coupled with its technically non-demanding experimental use makes it an attractive alternative or complement to selection screening, footprinting or affinity cleavage, and electrophoretic mobility shift assays for detecting, characterizing, and quantitating protein DNA binding affinity and selectivity. [Copyright &y& Elsevier]

Graphical abstract Abstract A series of N -acyl pyrazoles was examined as candidate serine hydrolase inhibitors in which the active site acylating reactivity and the leaving group ability of the pyrazole could be tuned not only through the nature of the acyl group (reactivity: amide > carbamate > urea), but also through pyrazole C4 substitution with electron-withdrawing or electron-donating substituents. Their impact on enzyme inhibitory activity displayed pronounced effects with the activity improving substantially as one alters both the nature of the reacting carbonyl group (urea > carbamate > amide) and the pyrazole C4 substituent (CN > H > Me). It was further demonstrated that the acyl chain of the N -acyl pyrazole ureas can be used to tailor the potency and selectivity of the inhibitor class to a targeted serine hydrolase. Thus, elaboration of the acyl chain of pyrazole-based ureas provided remarkably potent, irreversible inhibitors of fatty acid amide hydrolase (FAAH, apparent K i = 100–200 pM), dual inhibitors of FAAH and monoacylglycerol hydrolase (MGLL), or selective inhibitors of MGLL (IC 50 = 10–20 nM) while simultaneously minimizing off-target activity (e.g., ABHD6 and KIAA1363). [ABSTRACT FROM AUTHOR]

A key series of vinblastine analogs 7 – 13 , which contain modifications to the C20′ ethyl group, was prepared with use of two distinct synthetic approaches that provide modifications of the C20′ side chain containing linear and cyclized alkyl groups or added functionalized substituents. Their examination revealed the unique nature of the improved properties of the synthetic vinblastine 6 , offers insights into the origins of its increased tubulin binding affinity and 10-fold improved cell growth inhibition potency, and served to probe a small hydrophobic pocket anchoring the binding of vinblastine with tubulin. Especially noteworthy were the trends observed with substitution of the terminal carbon of the ethyl group that, with the exception of 9 (R = F vs H, equipotent), led to remarkably substantial reductions in activity (>10-fold): R = F (equipotent with H) > N 3 , CN (10-fold) > Me (50-fold) > Et (100-fold) > OH (inactive). This is in sharp contrast to the maintained ( 7 ) or enhanced activity ( 6 ) observed with its incorporation into a cyclic C20′/C15′-fused six-membered ring. [ABSTRACT FROM AUTHOR]

The design, synthesis, and evaluation of methyl 1,2,8,8a-tetrahydrocyclopropa[ c ]imidazolo[4,5- e ]indol-4-one-6-carboxylate (CImI) derivatives are detailed representing analogs of duocarmycin SA and yatakemycin containing an imidazole replacement for the fused pyrrole found in the DNA alkylation subunit. [ABSTRACT FROM AUTHOR]

Modifications to the enzymatic glycosylation of vancomycin and its residue 4 thioamide analogue are detailed that significantly reduce the enzyme loading and amount of glycosyl donor needed for each glycosylation reaction, provide a streamlined synthesis and replacement for the synthetic UDP-vancosamine glycosyl donor to improve both access and storage stability, and permit a single-pot, two-step conversion of the aglycons to the fully glycosylated synthetic glycopeptides now conducted at higher concentrations. The improvements are exemplified with the two-step, one-pot glycosylation of [Ψ[C(=S)NH]Tpg4]vancomycin aglycon (92%) conducted on a 400 mg scale (2 mg–1 g scales) and vancomycin aglycon itself (5 mg scale, 84%). [Display omitted] [ABSTRACT FROM AUTHOR]

The total syntheses of dihydrolysergic acid and dihydrolysergol are detailed based on a Pd(0)-catalyzed intramolecular Larock indole cyclization for the preparation of the embedded tricyclic indole (ABC ring system) and a subsequent powerful inverse electron demand Diels–Alder reaction of 5-carbomethoxy-1,2,3-triazine with a ketone-derived enamine for the introduction of a functionalized pyridine, serving as the precursor for a remarkably diastereoselective reduction to the N -methylpiperidine D-ring. By design, the use of the same ketone-derived enamine and a set of related complementary heterocyclic azadiene [4+2] cycloaddition reactions permitted the late stage divergent preparation of a series of alternative heterocyclic derivatives not readily accessible by more conventional approaches. [ABSTRACT FROM AUTHOR]

Abstract: Protein citrullination is just one of more than 200 known PTMs. This modification, catalyzed by the protein arginine deiminases (PADs 1–4 and PAD6 in humans), converts the positively charged guanidinium group of an arginine residue into a neutral ureido-group. Given the strong links between dysregulated PAD activity and human disease, we initiated a program to develop PAD inhibitors as potential therapeutics for these and other diseases in which the PADs are thought to play a role. Streptonigrin which possesses both anti-tumor and anti-bacterial activity was later identified as a highly potent PAD4 inhibitor. In an effort to understand why streptonigrin is such a potent and selective PAD4 inhibitor, we explored its structure–activity relationships by examining the inhibitory effects of several analogues that mimic the A, B, C, and/or D rings of streptonigrin. We report the identification of the 7-amino-quinoline-5,8-dione core of streptonigrin as a highly potent pharmacophore that acts as a pan-PAD inhibitor. [Copyright &y& Elsevier]

Abstract: The evaluation of a comprehensive α-helix mimetic library for binding the gp41 NHR hydrophobic pocket recognizing an intramolecular CHR α-helix provided a detailed depiction of structural features required for binding and led to the discovery of small molecule inhibitors (K i 0.6–1.3μM) that not only match or exceed the potency of those disclosed over the past decade, but that also exhibit effective activity in a cell–cell fusion assay (IC50 5–8μM). [Copyright &y& Elsevier]

Arenaviruses are responsible for acute hemorrhagic fevers worldwide and are recognized to pose significant threats to public health and biodefense. Small molecule compounds have recently been discovered that inhibit arenavirus entry and protect against lethal infection in animal models. These chemically distinct inhibitors act on the tripartite envelope glycoprotein (GPC) through its unusual stable signal peptide subunit to stabilize the complex against pH-induced activation of membrane fusion in the endosome. Here, we report the production and characterization of the intact transmembrane GPC complex of Junín arenavirus and its interaction with these inhibitors. The solubilized GPC is antigenically indistinguishable from the native protein and forms a homogeneous trimer in solution. When reconstituted into a lipid bilayer, the purified complex interacts specifically with its cell-surface receptor transferrin receptor-1. We show that small molecule entry inhibitors specific to New World or Old World arenaviruses bind to the membrane-associated GPC complex in accordance with their respective species selectivities and with dissociation constants comparable with concentrations that inhibit GPC-mediated membrane fusion. Furthermore, competitive binding studies reveal that these chemically distinct inhibitors share a common binding pocket on GPC. In conjunction with previous genetic studies, these findings identify the pH-sensing interface of GPC as a highly vulnerable target for antiviral intervention. This work expands our mechanistic understanding of arenavirus entry and provides a foundation to guide the development of small molecule compounds for the treatment of arenavirus hemorrhagic fevers. [ABSTRACT FROM AUTHOR]

Abstract: Nucleocytoplasmic transport of macromolecules is a fundamental process of eukaryotic cells. Translocation of proteins and many RNAs between the nucleus and cytoplasm is carried out by shuttling receptors of the β-karyopherin family, also called importins and exportins. Leptomycin B, a small molecule inhibitor of the exportin CRM1, has proved to be an invaluable tool for cell biologists, but up to now no small molecule inhibitors of nuclear import have been described. We devised a microtiter plate based permeabilized cell screen for small molecule inhibitors of the importin α/β pathway. By analyzing peptidomimetic libraries, we identified β-turn and α-helix peptidomimetic compounds that selectively inhibit nuclear import by importin α/β but not by transportin. Structure–activity relationship analysis showed that large aromatic residues and/or a histidine side chain are required for effective import inhibition by these compounds. Our validated inhibitors can be useful for in vitro studies of nuclear import, and can also provide a framework for synthesis of higher potency nuclear import inhibitors. [Copyright &y& Elsevier]

Abstract: The synthesis and evaluation of a key series of analogs of duocarmycin SA, bearing a single substituent at the C5′ position of the DNA binding subunit, are described. [Copyright &y& Elsevier]

Abstract: The design, synthesis and evaluation of methyl 1,2,8,8a-tetrahydrocyclopropa[c]oxazolo[2,3-e]indol-4-one-6-carboxylate (COI) derivatives are detailed representing analogs of duocarmycin SA containing an oxazole replacement for the fused pyrrole found in the alkylation subunit. [Copyright &y& Elsevier]

Abstract: The design, synthesis, and preliminary evaluation of methyl 1,2,8,8a-tetrahydrocyclopropa[c]thieno[3,2-e]indol-4-one-6-carboxylate (CTI) derivatives are detailed representing a single atom change (N to S) embedded in the duocarmycin SA alkylation subunit. [Copyright &y& Elsevier]

Abstract: The preparation and evaluation of a series of inhibitors of Myc/Max dimerization and Myc-induced cell transformation are described providing mycmycin-1 (3) and mycmycin-2 (4). [Copyright &y& Elsevier]

Abstract: Efficient syntheses and a preliminary evaluation of 1,2,11,11a-tetrahydrocyclopropa[c]naphtho[2,3-e]-indole (CNI) and 1,2,11,11a-tetrahydrocyclopropa[c]naphtho[1,2-e]indole (iso-CNI), and their derivatives containing an anthracene and phenanthrene variant of the CC-1065 or duocarmycin alkylation subunit are detailed. [Copyright &y& Elsevier]

Abstract: The comparative characterization of a series of 4-acyl-1,6-dialkylpiperazin-2-ones as potent cell entry inhibitors of the hemorrhagic fever arenavirus Lassa (LASV) is disclosed. The resolution and examination of the individual enantiomers of the prototypical LASV cell entry inhibitor 3 (16G8) is reported and the more potent (–)-enantiomer was found to be 15-fold more active than the corresponding (+)-enantiomer. The absolute configuration of (–)-3 was established by asymmetric synthesis of the active inhibitor (–)-(S)-3 (lassamycin-1). A limited deletion scan of lassamycin-1 defined key structural features required of the prototypical inhibitors. [Copyright &y& Elsevier]

Abstract: The melting curves of fatty acid amide hydrolase (FAAH) in the presence of 29 reversible inhibitors were measured using a thiol-reactive fluorophore. The thermal stability (Tm) of the FAAH/inhibitor complex varied significantly depending on the chemical characteristics of the inhibitors, notably variations in the head group. Two separate distributions were observed when Tm was plotted against K i. The majority of the inhibitors showed a positive correlation between binding affinity and Tm, however inhibitors with a pyridine carboxylic acid moiety in the head group fell in a distinct and uncorrelated distribution when tail groups were varied. [Copyright &y& Elsevier]

Abstract: A series of C4 substituted α-ketooxazoles were examined as inhibitors of the serine hydrolase fatty acid amide hydrolase in efforts that further define and generalize a fundamental substituent effect on enzyme inhibitory potency. Thus, a plot of the Hammett σ m versus −log K i provided a linear correlation (R 2 =0.90) with a slope of 3.37 (ρ =3.37), that is of a magnitude that indicates that of the electron-withdrawing character of the substituent dominates its effects (a one unit change in σ m provides a >1000-fold change in K i). [Copyright &y& Elsevier]

Viral hemorrhagic fevers caused by the arenaviruses Lassa virus in Africa and Machupo, Guanarito, Junin, and Sabia virus in South America are among the most devastating emerging human diseases with fatality rates of 15-35% and a limited antiviral therapeutic repertoire available. Here we used high throughput screening of synthetic combinatorial small molecule libraries to identity" inhibitors of arenavirus infection using pseudotyped virion particles bearing the glycoproteins (GPs) of highly pathogenic arenaviruses. Our screening efforts resulted in the discovery of a series of novel small molecule inhibitors of viral entry that are highly active against both Old World and New World hemorrhagic arenaviruses. We observed potent inhibition of infection of human and primate cells with live hemorrhagic arenaviruses (IC50 = 500 - 800 nM). Investigations of the mechanism of action revealed that the candidate compounds efficiently block pH-dependent fusion by the arenavirus GPs (IC50 of 200-350 nM). Although our lead compounds were potent against phylogenetically distant arenaviruses, they did not show activity, against other enveloped viruses with class 1 viral fusion proteins, indicating specificity for arenavirus GP-mediated membrane fusion. [ABSTRACT FROM AUTHOR]

The inosine monophosphate cyclohydrolase (IMPCH) component (residues 1-199) of the bifunctional enzyme amino-imidazole-4-carboxamide ribonucleotide transformylase (AICAR Tfase, residues 200 -593)/IMPCH (ATIC) catalyzes the final step in the de novo purine biosynthesis pathway that produces IMP. As a potential target for antineoplastic intervention, we designed IMPCH inhibitors, 1,5-dihydroimidazo[4,5-c] [1,2,6]thiadiazin-4(311)-one 2,2-dioxide (heterocycle, 1), the corresponding nucleoside (2), and the nucleoside monophosphate (nucleotide) (3), as mimics of the tetrahedral intermediate in the cyclization reaction. All compounds are competitive inhibitors against IMPCH (Ki values = 0.13-0.23 µM) with the simple heterocycle 1 exhibiting the most potent inhibition (Ki = 0.13 µM). Crystal structures of bifunctional ATIC in complex with nucleoside 2 and nucleotide 3 revealed IMPCH binding modes similar to that of the IMPCH feedback inhibitor, xanthosine 5′-monophosphate. Surprisingly, the simpler heterocycle 1 had a completely different IMPCH binding mode and was relocated to the phosphate binding pocket that was identified from previous xanthosine 5′-monophosphate structures, The aromatic imidazole ring interacts with a helix dipole, similar to the interaction with the phosphate moiety of 3. The crystal structures not only revealed the mechanism of inhibition of these compounds, but they now serve as a platform for future inhibitor improvements. Importantly, the nucleoside-complexed structure supports the notion that inhibitors lacking a negatively charged phosphate can still inhibit IMPCH activity with comparable potency to phosphate-containing inhibitors. Provocatively, the nucleotide inhibitor 3 also binds to the AICAR Tfase domain of ATIC, which now provides a lead compound for the design of inhibitors that simultaneously target both active sites of this bifunctional enzyme. [ABSTRACT FROM AUTHOR]

Abstract: The discovery of a new class of aminoimidazole carboxamide ribonucleotide transformylase (AICAR Tfase) inhibitors through screening peptidomimetic libraries (>40,000 compounds) that act by inhibiting requisite enzyme dimerization is disclosed. In addition to defining key structural features of the lead compounds responsible for the activity, kinetic analysis of the remarkably small inhibitors established that they act as noncompetitive, dissociative inhibitors of AICAR Tfase with the prototypical lead (A1B3, Cappsin 1) exhibiting a K i of 3.1±0.3μM. Thus, the studies define a unique approach to selectively targeting AICAR Tfase over all other folate-dependent enzymes, and it represents only one of a few enzymes for which inhibition achieved by disrupting requisite enzyme dimerization has emerged from screening unbiased combinatorial libraries. [Copyright &y& Elsevier]

Abstract: The synthesis and evaluation of N-{4-[5-(2,4-diamino-6-oxo-1,6-dihydropyrimidin-5-yl)-2-(2,2,2-trifluoroacetyl)pentyl]benzoyl}-l-glutamic acid (2) as an inhibitor of glycinamide ribonucleotide transformylase (GAR Tfase) and aminoimidazole carboxamide ribonucleotide transformylase (AICAR Tfase) are reported. The inhibitor 2 was prepared in a convergent synthesis involving C-alkylation of methyl 4-(4,4,4-trifluoro-3-dimethylhydrazonobutyl)benzoate with 1-chloro-3-iodopropane followed by construction of the pyrimidinone ring. Compound 2 was found to be an effective inhibitor of recombinant human GAR Tfase (Ki=0.50μM), whereas it was inactive (Ki>100μM) against E. coli GAR Tfase as well as recombinant human AICAR Tfase. Compound 2 exhibited modest, purine-sensitive growth inhibitory activity against the CCRF-CEM cell line (IC50=6.0μM). [Copyright &y& Elsevier]

Abstract: Structurally-related, but non-polyglutamylatable, derivatives of 10-CF3CO-DDACTHF (1), which incorporate l-glutamine (2) and l-isoglutamine (3) in place of l-glutamate, were prepared and evaluated as inhibitors of recombinant human (rh) GAR Tfase. While the l-glutamate α-carboxamide derivative 3 was much less effective as a rhGAR Tfase inhibitor (Ki=4.8μM) and inactive in cellular functional assays, the γ-carboxamide derivative 2 was found to be a potent and selective rhGAR Tfase inhibitor (Ki=0.056μM) being only 4-fold less potent than 1 (Ki=0.015μM). Moreover, 2 was effective in cellular functional assays exhibiting purine sensitive cytotoxic activity (IC50=300nM, CCRF-CEM) only 20-fold less potent than 1 (IC50=16nM), consistent with inhibition of de novo purine biosynthesis via selective inhibition of GAR Tfase. Like 1, 2 is transported into the cell by the reduced folate carrier. Unlike 1, the functional activity of 2 is not dependent upon FPGS polyglutamylation. [Copyright &y& Elsevier]

Abstract: The synthesis and evaluation of 10-methanesulfonyl-DDACTHF (1), 10-methanesulfonyl-5-DACTHF (2), and 10-methylthio-DDACTHF (3) as potential inhibitors of glycinamide ribonucleotide transformylase (GAR Tfase) and aminoimidazole carboxamide ribonucleotide transformylase (AICAR Tfase) are reported. The compounds 10-methanesulfonyl-DDACTHF (1, Ki=0.23μM), 10-methanesulfonyl-5-DACTHF (2, Ki=0.58μM), and 10-methylthio-DDACTHF (3, Ki=0.25μM) were found to be selective and potent inhibitors of recombinant human GAR Tfase. Of these, 3 exhibited exceptionally potent, purine sensitive growth inhibition activity (3, IC50=100nM) against the CCRF–CEM cell line being 3-fold more potent than Lometrexol and 30-fold more potent than the parent, unsubstituted DDACTHF, whereas 1 and 2 exhibited more modest growth inhibition activity (1, IC50=1.0μM and 2, IC50=2.0μM). [Copyright &y& Elsevier]

Abstract: The concurrent implementation of a proteome-wide serine hydrolase selectivity screen with traditional efforts to optimize fatty acid amide hydrolase (FAAH) inhibition potency led to the expedited discovery of a new class of exceptionally potent (Ki<300pM) and unusually selective (>100-fold selective) inhibitors. The iterative inhibitor design and evaluation with assistance of the selectivity screen served to differentiate otherwise indistinguishable inhibitors permitting the simultaneous optimization of potency and selectivity. Significantly, the simultaneous assessment of all potential competitive enzymes with the selectivity screen does not require the use of expressed or purified enzymes or a competitive substrate, no modification of the inhibitors is required, and the relative potency for competitive enzymes can be quantified (IC50’s) including those that lack known substrates or function. [Copyright &y& Elsevier]