Substituted tetrazoles as multipurpose screening compounds.

Tetrazoles are small functional heterocycles that are suited to serve simultaneously as aromatic platform for diversity and as functional interaction motif. Furthermore, the tetrazole ring and its deprotonated tetrazolate counterpart are metal ion complexing ligands that possess a rich variety of binding and bridging modes. We recently demonstrated that fragments containing the tetrazole moiety and a metal chelating hydrazide group are well suited to discover selective screening hits with high ligand efficiency for a given protein target. Here, we report the synthesis and characterization of new polydentate tetrazole-containing screening compounds and their synthetic precursors as well as their deposition in a multipurpose screening library in the frame of the EU-OPENSCREEN network. The pure and well-characterized screening compounds could be useful to aid drug discovery programs for multiple or hitherto undruggable targets by enclosure of under-represented tetrazole derivatives.

Tetrazolylhydrazides as Selective Fragment-Like Inhibitors of the JumonjiC-Domain-Containing Histone Demethylase KDM4A.

The JumonjiC-domain-containing histone demethylase 2A (JMJD2A, KDM4A) is a key player in the epigenetic regulation of gene expression. Previous publications have shown that both elevated and lowered enzyme levels are associated with certain types of cancer, and therefore the definite role of KDM4A in oncogenesis remains elusive. To identify a novel molecular starting point with favorable physicochemical properties for the investigation of the physiological role of KDM4A, we screened a number of molecules bearing an iron-chelating moiety by using two independent assays. In this way, we were able to identify 2-(1H-tetrazol-5-yl)acetohydrazide as a novel fragment-like lead structure with low relative molecular mass (Mr =142 Da), low complexity, and an IC50 value of 46.6 µm in a formaldehyde dehydrogenase (FDH)-coupled assay and 2.4 µm in an antibody-based assay. Despite its small size, relative selectivity against two other demethylases could be demonstrated for this compound. This is the first example of a tetrazole group as a warhead in JMJD demethylases.

Microwave-assisted synthesis and evaluation of acylhydrazones as potential inhibitors of bovine glutathione peroxidase.

Glutathione peroxidases (GPx) were the first selenocysteine enzymes identified. They play critical roles in cellular defense to excessH(2)O(2) and lipid peroxides, with GPx1 contributing the most cellular activity. In the treatment of cancer, for example, in lymphomas and leukaemias, evidence has been accumulating that up-regulation of the GPx system may serve to protect cancer cells from oxidative stress caused by anticancer drugs. We hypothesize that small molecules which block GPx1 could help overcome acquired resistance to anticancer drugs by raising the level of oxidative stress in cancer cells. Our previous efforts identified an acylhydrazone as a lead structure for the inhibition of GPx1. Now we report the microwave-supported synthesis and inhibitory screening of a series of 78 analogs. The special conformational isomerism resulting of the acylhydrazone functionality was investigated by the analysis of distinct NMR data and crystal structures, indicating that conformers at the C(O)-N hydrazide bond are responsible for this phenomena. Though some of the analogs showed poor aqueous solubility and could not be tested in the enzyme assay, the combinatorial approach led to the identification of a closely related isomer of the lead compound with increased inhibitory activity: N'-[1-(4-hydroxyphenyl)ethylidene]-2-(1H-imidazol-1-yl)acetohydrazide. This success supports the idea that novel GPx1 inhibitors can be developed by drug-design methods and paves the way for a new class of GPx1 inhibitors.

Identification of a glutathione peroxidase inhibitor that reverses resistance to anticancer drugs in human B-cell lymphoma cell lines.

Cancer cells isolated from two patients with malignant non-Hodgkin B-cell lymphomas that became resistant to chemotherapy during clinical treatment were made ≥fourfold resistant in culture to anticancer drugs, that is cisplatin, etoposide, methotrexate and bortezomib. Because most resistant lines showed significantly increased expression of the anti-oxidative enzyme glutathione peroxidase 1 (GPx1), GPx1 was investigated as a target for inhibitor development. Virtual screening of a library of diverse structures by docking them to the active site of the X-ray crystal structure of bovine GPx1 uncovered compounds that might block the enzyme. An enzyme assay confirmed an acylhydrazone heterocycle (3) with GPx inhibitory activity. Combinations of 3 with the anticancer drugs listed above led to reversal of resistance in the lymphoma cell lines.

Synthesis of a hydrolytically stable, fluorescent-labeled ATP analog as a tool for probing adenylyl cyclases.

(2R,3S,4R,5R)-5-(6-Amino-9H-purin-9-yl)-3-hydroxy-4-[2-(methylamino)benzamido]tetrahydrofuran-2-yl-methoxy[(hydroxy)phosphoryloxy][(hydroxy)phosphoryl]dichloromethylphosphonic acid was synthesized as a chemically and metabolically stable analog of ATP substituted with a fluorescent methylanthranoyl (MANT) residue. The compound is intended for studying the binding site and function of adenylyl cyclases (ACs), which was exemplified by studying its interaction with Bacillus anthracis edema factor (EF) AC exotoxin.

Inhibitors of adenosine consuming parasites through polymer-assisted solution phase synthesis of lipophilic 5'-amido-5'-deoxyadenosine derivatives.

Given the more or less global spread of multidrug-resistant plasmodia, structurally diverse starting points for the development of chemotherapeutic agents for the treatment of malaria are urgently needed. Thus, a series of 20 adenosine derivatives with a large lipophilic substituent in N(6)-position were prepared in order to evaluate their potential to inhibit the chloroquine resistant Plasmodium falciparum strain K1 in vitro. The rationale for synthesis of these structures was the high probability of interactions with multiple adenosine associated targets and the assumption that a large hydrophobic N(6)-(4-phenoxy)benzyl substitution should allow the molecules to diffuse across parasite membranes. Starting from readily available inosine, the new compounds were prepared as single isomers using a polymer-assisted acylation protocol enabling the straightforward isolation of the target compounds in pure form. Heterocyclic ring systems were synthesized on-bead on Kenner's safety-catch linker prior to acylation of the scaffold in solution. Most of the highly pure compounds displayed anti-plasmodial activity in the low micromolar or even submicromolar concentration range.