Site-directed Mutagenesis of Key Residues Unveiled a Novel Allosteric Site on Human Adenosine Kinase for Pyrrolobenzoxa(thia)zepinone Non-Nucleoside Inhibitors.

Most nucleoside kinases, besides the catalytic domain, feature an allosteric domain which modulates their activity. Generally, non-substrate analogs, interacting with allosteric sites, represent a major opportunity for developing more selective and safer therapeutics. We recently developed a series of non-nucleoside non-competitive inhibitors of human adenosine kinase (hAK), based on a pyrrolobenzoxa(thia)zepinone scaffold.

Discovery of Multitarget Antivirals Acting on Both the Dengue Virus NS5-NS3 Interaction and the Host Src/Fyn Kinases.

This study describes the discovery of novel dengue virus inhibitors targeting both a crucial viral protein-protein interaction and an essential host cell factor as a strategy to reduce the emergence of drug resistance. Starting from known c-Src inhibitors, a virtual screening was performed to identify molecules able to interact with a recently discovered allosteric pocket on the dengue virus NS5 polymerase. The selection of cheap-to-produce scaffolds and the exploration of the biologically relevant chemical space around them suggested promising candidates for chemical synthesis.

Studies on the ATP Binding Site of Fyn Kinase for the Identification of New Inhibitors and Their Evaluation as Potential Agents against Tauopathies and Tumors.

Fyn is a member of the Src-family of nonreceptor protein-tyrosine kinases. Its abnormal activity has been shown to be related to various human cancers as well as to severe pathologies, such as Alzheimer's and Parkinson's diseases. Herein, a structure-based drug design protocol was employed aimed at identifying novel Fyn inhibitors.

Super-telomeres in transformed human fibroblasts.

Telomere length maintenance is critical for organisms' long-term survival and cancer cell proliferation. Telomeres are kept within species-specific length ranges by the interplay between telomerase activity and telomeric chromatin organization. In this paper, we exploited telomerase immortalized human fibroblasts (cen3tel) that gradually underwent neoplastic transformation during culture propagation to study telomere composition and length regulation during the transformation process.

Clostridium difficile DNA polymerase IIIC: basis for activity of antibacterial compounds.

Based on the finding that aerobic Gram-positive antibacterials that inhibit DNA polymerase IIIC (pol IIIC) were potent inhibitors of the growth of anaerobic Clostridium difficile (CD) strains, we chose to clone and express the gene for pol IIIC from this organism. The properties of the recombinant enzyme are similar to those of related pol IIICs from Gram-positive aerobes, e.g.

N-Phenyl-9-(hydroxyalkyl)guanines and related compounds are substrates for Herpes simplex virus thymidine kinases.

Herpes simplex virus (HSV) types 1 and 2 thymidine kinases (TK) are responsible for phosphorylation of antiherpes acyclonucleosides such as acyclovir (ACV) and 9-(4-hydroxybutyl)guanine (HBG). Related compounds, the N-phenyl-9-(hydroxyalkyl)guanines, are devoid of direct antiviral activity, but potently inhibit the viral TKs and block viral reactivation from latency . The similarity in structure between the acyclonucleosides and TK inhibitors raised the question of the relevance of phosphorylation of certain of the latter analogs in their mechanisms of action.

Enantioselective binding of second generation pyrrolobenzoxazepinones to the catalytic ternary complex of HIV-1 RT wild-type and L100I and K103N drug resistant mutants.

We investigated some pyrrolobenzoxazepinone (PBOs, 3e-i) analogues of early described effective non-nucleoside inhibitors of HIV-1 reverse transcriptase (RT). Enzymological studies of 3e-i enantiomers, with wild type (wt) RT and some drug-resistant mutants, revealed a stereoselective mode of action and selectivity for RT ternary complex. Unexpectedly (+)-3g was found more potent towards the L100I mutant than towards the wt RT, whereas (+)-3h inhibited the K103N mutant and RT wt with comparable potency.

Non-nucleoside inhibitors of human adenosine kinase: synthesis, molecular modeling, and biological studies.

Adenosine kinase (AK) catalyzes the phosphorylation of adenosine (Ado) to AMP by means of a kinetic mechanism in which the two substrates Ado and ATP bind the enzyme in a binary and/or ternary complex, with distinct protein conformations. Most of the described inhibitors have Ado-like structural motifs and are nonselective, and some of them (e.g.

Sensitivity of monkey B virus (Cercopithecine herpesvirus 1) to antiviral drugs: role of thymidine kinase in antiviral activities of substrate analogs and acyclonucleosides.

Herpes B virus (B virus [BV]) is a macaque herpesvirus that is occasionally transmitted to humans where it can cause rapidly ascending encephalitis that is often fatal. To understand the low susceptibility of BV to the acyclonucleosides, we have cloned, expressed, and characterized the BV thymidine kinase (TK), an enzyme that is expected to "activate" nucleoside analogs. This enzyme is similar in sequence and properties to the TK of herpes simplex virus (HSV), i.

Simplified analogues of immucillin-G retain potent human purine nucleoside phosphorylase inhibitory activity.

A set of deazaguanine derivatives 1-3 targeting human purine nucleoside phosphorylase (hPNP) have been designed and synthesized. The new compounds are characterized by the presence of a structurally simplified "azasugar" motif to be more easily accessible by chemical synthesis than previous inhibitors. In the enzymatic assays, some of the new derivatives proved to be able to inhibit hPNP at low nanomolar concentration, thereby showing the same inhibitory potency in vitro as immucillin-H (IMH).

Inhibition of herpes simplex virus thymidine kinases by 2-phenylamino-6-oxopurines and related compounds: structure-activity relationships and antiherpetic activity in vivo.

Derivatives of the herpes simplex thymidine kinase inhibitor HBPG [2-phenylamino-9-(4-hydroxybutyl)-6-oxopurine] have been synthesized and tested for inhibitory activity against recombinant enzymes (TK) from herpes simplex types 1 and 2 (HSV-1, HSV-2). The compounds inhibited phosphorylation of [3H]thymidine by both enzymes, but potencies differed quantitatively from those of HBPG and were generally greater for HSV-2 than HSV-1 TKs. Changes in inhibitory potency were generally consistent with the inhibitor/substrate binding site structure based on published X-ray structures of HSV-1 TK.

Microwave-assisted synthesis and biological evaluation of novel uracil derivatives inhibiting human thymidine phosphorylase.

New 5-chloro-6-substituted-uracil derivatives have been prepared by microwave assisted-synthesis and tested in vitro as thymidine phosphorylase inhibitors. One of these compounds showed potent inhibitory activity, with an IC50 value in the submicromolar range. The biological activity of the new compounds is discussed in terms of structure-activity relationship.

Enantioselectivity of ribonucleotide reductase: a first study using stereoisomers of pyrimidine 2'-azido-2'-deoxynucleosides.

In this paper, the enantioselectivity of ribonucleotide reductase (RNR, EC 1.17.4.