A Highly Potent Non-Nucleoside Adenosine Deaminase Inhibitor:  Efficient Drug Discovery by Intentional Lead Hybridization

Terasaka, Tadashi; Kinoshita, Takayoshi; Kuno, Miyuki; Nakanishi, Isao

doi:10.1021/ja038606l

Cited by 57 publications

(56 citation statements)

References 17 publications

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“…In 7 the two phenyl rings are connected through an amide linker while in 8 the linker is an ureic residue. Both these functions are present in other potent ADA inhibitors (7,12). As summarized in Table 1, neither 7 nor 8 showed any appreciable inhibitory activity, proving that in the pyrazolo [1,5-a]pyrimidin-7-one series the preferred substitution pattern is represented by the flexible alkyl chain.…”

Section: Resultsmentioning

confidence: 96%

“…A rational drug design is further complicated by the fact that in response to different inhibitors, ADA can assume either an open or a closed conformation by changing the position of the H3 α-helix (Thr57-Ala73). The open conformation corresponds to the apo-form (PDB ID code 3iar) and is preferred when a nonnucleoside inhibitor is bound (12,13). In this case, the active site presents a hydrophilic subsite S0 and three hydrophobic subsites F0, F1, and F2 ( Fig.…”

mentioning

confidence: 99%

“…In fact, experimental evidences suggest that the n-hexyl group of EHNA interacts with the narrow hydrophobic entrance of the ADA active site destabilizing the closed form. Another example of the difficulty of predicting a ligand binding model comes from the inhibitor FR221647, which has also been found to bind to the open conformation in spite of having been designed as a binder of the closed form (12).…”

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confidence: 99%

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Sampling protein motion and solvent effect during ligand binding

Limongelli

Marinelli

Cosconati³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

110

105

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An exhaustive description of the molecular recognition mechanism between a ligand and its biological target is of great value because it provides the opportunity for an exogenous control of the related process. Very often this aim can be pursued using high resolution structures of the complex in combination with inexpensive computational protocols such as docking algorithms. Unfortunately, in many other cases a number of factors, like protein flexibility or solvent effects, increase the degree of complexity of ligand/protein interaction and these standard techniques are no longer sufficient to describe the binding event. We have experienced and tested these limits in the present study in which we have developed and revealed the mechanism of binding of a new series of potent inhibitors of Adenosine Deaminase. We have first performed a large number of docking calculations, which unfortunately failed to yield reliable results due to the dynamical character of the enzyme and the complex role of the solvent. Thus, we have stepped up the computational strategy using a protocol based on metadynamics. Our approach has allowed dealing with protein motion and solvation during ligand binding and finally identifying the lowest energy binding modes of the most potent compound of the series, 4-decyl-pyrazolo[1,5-a]pyrimidin-7-one.ADA | well-tempered metadynamics | ligand/protein docking | path collective variables | reweighting algorithm A denosine Deaminase (ADA) regulates the purine metabolism by catalyzing the irreversible hydrolysis of adenosine to inosine and 2′-deoxyadenosine to 2′-deoxyinosine. Thus, this enzyme plays a crucial role in many pathologies such as inflammation, some types of cancer, and others which are strictly connected to the physiological level of these nucleosides (1-5). Despite great efforts in developing ADA inhibitors, only Pentostatin is currently in clinical use (I in Fig. S1) (3). However, recent progress has been reported by Terasaka et al. and by some of us who have developed a new generation of nonnucleoside ADA inhibitors (II, III, and IV in Fig. S1) (6-11). Unfortunately, the understanding at molecular level of the ligand/ADA interaction is hampered by the pronounced ability of the active site to accommodate different inhibitors and by the crucial role played by water molecules during ligand binding. A rational drug design is further complicated by the fact that in response to different inhibitors, ADA can assume either an open or a closed conformation by changing the position of the H3 α-helix (Thr57-Ala73). The open conformation corresponds to the apo-form (PDB ID code 3iar) and is preferred when a nonnucleoside inhibitor is bound (12, 13). In this case, the active site presents a hydrophilic subsite S0 and three hydrophobic subsites F0, F1, and F2 (Fig. 1A) (13). The S0 subsite is defined by the structural gate formed by a β-strand (Leu182-Asp185) and two leucine side chains attached to the H3 α-helix while the F0 site is formed by the hydrophobic side chains of the H3 α-helix. In the op...

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Section: Resultsmentioning

confidence: 96%

mentioning

confidence: 99%

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confidence: 99%

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Sampling protein motion and solvent effect during ligand binding

Limongelli

Marinelli

Cosconati³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

110

105

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“…Indeed, ADA inhibitors reported to date are largely nucleoside analogues such as EHNA. [27] MAC-0038732 and other non-nucleoside inhibitors [28] represent fresh approaches to this emerging target in inflammation and immunity.…”

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confidence: 99%

Small‐Molecule Screening Made Simple for a Difficult Target with a Signaling Nucleic Acid Aptamer that Reports on Deaminase Activity

et al. 2006

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Conventional approaches to small-molecule screening are currently being challenged by the availability of vast amounts of genomic sequence information and a plethora of potential targets. This challenge has precipitated a need for new screening paradigms that emphasize simplicity, capacity, and parallelization. Herein we report the application of signaling DNA-aptamer technology for the development and execution of a high-throughput screen for an otherwise problematic target, adenosine deaminase (ADA). The approach employed a signaling DNA aptamer that reports on adenosine concentration over the course of the enzymatic reaction. The assay was extremely robust in a screen of more than 44 000 molecules and revealed a new competitive inhibitor of the deaminase. Nucleic acid aptamers have proven worthy as a routinely selectable species for a wide variety of small molecules and so this proof-of-principle work has broad

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“…X-ray crystallography [31,32] and molecular dynamics [33] were analyzed at the molecular level for the ADA-inhibitor complex. Binding sites for some of inhibitors and the effect of their structure on the enzyme were investigated.…”

Section: Introductionmentioning

confidence: 99%

Kinetic, thermodynamic and statistical studies on the inhibition of adenosine deaminase by aspirin and diclofenac

Ajloo

Saboury

Haghi-Asli

et al. 2007

Journal of Enzyme Inhibition and Medicinal Chemistry

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The kinetic and thermodynamic effects of aspirin and diclofenac on the activity of adenosine deaminase (ADA) were studied in 50 mM phosphate buffer pH ¼ 7.5 at 27 and 378C, using UV-Vis spectrophotometry and isothermal titration calorimetry (ITC). Aspirin exhibits competitive inhibition at 27 and 378C and the inhibition constants are 42.8 and 96.8 mM respectively, using spectrophotometry. Diclofenac shows competitive behavior at 278C and uncompetitive at 378C with inhibition constants of 56.4 and 30.0 mM, at respectively. The binding constant and enthalpy of binding, at 278C are 45 mM, 2 64.5 kJ/mol and 61 mM, 2 34.5 kJ/mol for aspirin and diclofenac. Thermodynamic data revealed that the binding process for these ADA inhibitors is enthalpy driven. QSAR studies by principal component analysis implemented in SPSS show that the large, polar, planar, and aromatic nucleoside and small, aromatic and polar non-nucleoside molecules have lower inhibition constants.

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A Highly Potent Non-Nucleoside Adenosine Deaminase Inhibitor: Efficient Drug Discovery by Intentional Lead Hybridization

Cited by 57 publications

References 17 publications

Sampling protein motion and solvent effect during ligand binding

Sampling protein motion and solvent effect during ligand binding

Small‐Molecule Screening Made Simple for a Difficult Target with a Signaling Nucleic Acid Aptamer that Reports on Deaminase Activity

Kinetic, thermodynamic and statistical studies on the inhibition of adenosine deaminase by aspirin and diclofenac

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