Inhibitor noncompetitive

Compounds that strongly chelate iron have been known for many years to stabilize HIF-la as well as upregulate proteins involved in red blood cell production erythropoietin (EPO), angiogenesis, vascular endothelial growth factor (VEGF), and iron transport. Some, but not all, of the pharmacological actions of iron chelators are produced by inhibition of PHD enzymes resulting in elevation of cellular HIF content. The action of selected iron chelators as they relate to PHD inhibition are briefly summarized here. 

In a recent patent application, mice treated with a related iron chelator, deferasirox (DFS), showed reduced body weight while on a high fat diet compared to untreated controls. Additionally, DFS was claimed to improve whole body metabolism and energy expenditure as measured by increased 02 consumption and C02 production as well as a reduction in white adipose and visceral fat, despite little difference between food intake in the control and treated animal groups [25]. 

4-Dihydroxybenzoic acid (DHB) is also a commonly used tool to measure the pharmacological effects of HIF-la stabilization via PHD inhibition. Recently, it was shown that mice pretreated with DHB (100 mg/kg, i.p.) showed a marked resistance to the neurotoxic effects of l-methyl-4-phenyl-l,2,3,6-tetrahydropyridine (MPTP) via protection of dopaminergic cell loss and striatal denervation. Importantly, this protection was seen to coincide with HIF-la stabilization, and the prevention of the MPTP-induced loss of ferroportin and striatal iron. Additionally, in these studies, DHB was also observed to block MPTP-induced reduction in mitochondrial pyruvate dehydrogenase, at both the mRNA level and through the measurement of enzyme activity in midbrain substantia nigra [26]. 

It is well known that 1,10-phenanthrolines are highly active ironchelating agents. The parent compound itself has recently been shown to increase HIF-la levels in ocular tissue and to suppress 02-mediated epithelial cell proliferation when administered to mice [29]. A quantitative assay was developed to measure transcriptional potency of certain HIF stabilizers via an HRE-mediated (3-lactamase production in which the EC50 of 1,10-phenanthroline was measured to be approximately 8 pM. In addition, VEGF was dose-dependently produced in mouse embryonic fibroblasts by 1,10-phenanthroline with an EC50 of 

---

Other stmctural variations in both series are the stereochemistry at C3 and the degree of oxidation on the nucleus and side chains. Cardiac steroids probably exert their inotropic effects by acting as specific, noncompetitive inhibitors of — ATPases, known as sodium pumps, and thus... 

Unlike a competitive inhibitor, a noncompetitive inhibitor does not compete with the substrate for the binding site, siace the inhibitor and substrate can biad to the enzyme either iadepeadeatiy or smi A. 2in.eou y.]S1oncompetitive inhibition... 

Like a noncompetitive inhibitor, an uncompetitive inhibitor does not compete with the substrate since it binds to the enzyme—substrate complex but not to the free enzyme. Uncompetitive inhibition... 

Both threo- (14) and eo f >"4-fluoro-DL-glutamic acid (/5) are noncompetitive inhibitors of glutamine synthase, an enzyme that catalyzes the synthesis of glutamine from L-glutamic acid and ammonia. This mhibibon may explain the... 

Noncompetitive inhibitors interact with both E and ES (or with S and ES, but this is a rare and specialized case). Obviously, then, the inhibitor is not binding to the same site as S, and the inhibition cannot be overcome by raising [S]. There are two types of noncompetitive inhibition pure and mixed. 

Anti-gout Drugs. Figure 1 Xanthine oxidase-catalyzed reactions. Xanthine oxidase converts hypoxanthine to xanthine and xanthine to uric acid, respectively. Hypoxanthine and xanthine are more soluble than uric acid. Xanthine oxidase also converts the uricostatic drug allopurinol to alloxanthine. Allopurinol and hypoxanthine are isomers that differ from each other in the substitution of positions 7 and 8 of the purine ring system. Although allopurinol is converted to alloxanthine by xanthine oxidase, allopurinol is also a xanthine oxidase inhibitor. Specifically, at low concentrations, allopurinol acts as a competitive inhibitor, and at high concentrations it acts as a noncompetitive inhibitor. Alloxanthine is a noncompetitive xanthine oxidase inhibitor. XOD xanthine oxidase. 

Allopurinol is an analog of hypoxanthine and is converted to alloxanthine by XOD. Both allopurinol and hypoxanthine inhibit XOD (Fig. 1). Alloxanthine is a noncompetitive inhibitor of XOD as is allopurinol at high concentrations. At low concentrations, allopurinol is a competitive inhibitor of XOD. As a result of XOD inhibition, the formation of the poorly soluble... 

A noncompetitive inhibitor is one that binds to both E and E S. If both dissociation constants are the same, the Michaelis-Menten equation is... 

NMR line shape analysis, 260-263 Negative activation enthalpy, 162 Noncompetitive inhibitor, 93 Nucleophilic catalysis, 237-238 Numerical simulations, 112-119... 

Love RA, Parge HE, Yu X, Hickey MJ, Diehl W, Gao J, Wriggers H, Ekker A, Wang L, Thomson JA, Dragovich PS, Fuhrman SA (2003) Crystallographic identification of a noncompetitive inhibitor binding site on the hepatitis C vims NS5B RNA polymerase enzyme. J Virol 77 7575-7581... 

Double reciprocal plots distinguish between competitive and noncompetitive inhibitors and simpbfy evaluation of inhibition constants Aj. v, is determined at several substrate concentrations both in the presence and in the absence of inhibitor. For classic competitive inhibition, the lines that connect the experimental data points meet at they axis (Figure 8-9). Since they intercept is equal to IIV, this pattern indicates that wben 1/[S] approaches 0, Vj is independent of the presence of inhibitor. Note, however, that the intercept on the X axis does vary with inhibitor concentration—and that since is smaller than HK, (the apparent... 

The effects of competitive inhibitors, which typically resemble substrates, are overcome by raising the concentration of the substrate. Noncompetitive inhibitors lower but do not affect K. ... 

Nonnucleoside reverse transcriptase inhibitor (NNRTI) A noncompetitive inhibitor of the viral reverse transcriptase enzyme that binds to the active site of the enzyme itself, rather than by terminating the enzymatic product. NNRTIs are only active against human immunodeficiency virus-1. 

A noncompetitive inhibitor is one that displays binding affinity for both the free enzyme and the enzyme-substrate complex or subsequent species. In this situation the binding affinity cannot be defined by a single equilibrium dissociation constant ... 

Because noncompetitive inhibitors bind to both the free enzyme and the ES complex, or subsequent species in the reaction pathway, we would expect these molecules to exert a kinetic effect on the E + S —> ES" process, thus effecting the apparent values of both VmdX/KM (influenced by both the K and al, terms) and Vmax (influenced by the aK term). This is reflected in the velocity equation for noncompetitive inhibition ... 

Referring back to Equation (3.2), we see that the effect of a noncompetitive inhibitor on the kinetic constants is to lower the apparent value of Vmax and to increase, decrease, or leave unaffected the apparent value of Ku, depending on whether a is >1, <1 or =1, respectively (see Table 3.3). These effects are apparent in plots... 

The nonnucleoside reverse transcriptase inhibitors (NNRTIs), used in the treatment of AIDS, provide interesting examples of clinically relevant noncompetitive inhibitors. The causative agent of AIDS, HIV, belongs to a virus family that relies on an RNA-based genetic system. Replication of the vims requires reverse transcription of the viral genomic RNA into DNA, which is then incorporated into the genome of the infected host cell. Reverse transcription is catalyzed by a virally encoded nucleic acid polymerase, known as reverse transcriptase (RT). This enzyme is critical for viral replication inhibition of HIV RT is therefore an effective mechanism for abrogating infection in patients. 

Figure 3.6 Substrate titration of steady state velocity for an enzyme in the presence of a noncompetitive inhibitor (a = 1) at varying concentrations. (A) Untransformed data (B) data as in (A) plotted on a semilog scale (C) data as in (A) plotted in double reciprocal form. For all three plots the data are fit to Equation (3.2).

Yonetani-Theorell analysis can be quite useful in determining whether chemically distinct noncompetitive inhibitors are likely to share a common binding pocket on a target enzyme. This information can be very valuable in defining strategies for parallel SAR studies on two or more chemical series of inhibitiors. 

In this chapter we described the thermodynamics of enzyme-inhibitor interactions and defined three potential modes of reversible binding of inhibitors to enzyme molecules. Competitive inhibitors bind to the free enzyme form in direct competition with substrate molecules. Noncompetitive inhibitors bind to both the free enzyme and to the ES complex or subsequent enzyme forms that are populated during catalysis. Uncompetitive inhibitors bind exclusively to the ES complex or to subsequent enzyme forms. We saw that one can distinguish among these inhibition modes by their effects on the apparent values of the steady state kinetic parameters Umax, Km, and VmdX/KM. We further saw that for bisubstrate reactions, the inhibition modality depends on the reaction mechanism used by the enzyme. Finally, we described how one may use the dissociation constant for inhibition (Kh o.K or both) to best evaluate the relative affinity of different inhibitors for ones target enzyme, and thus drive compound optimization through medicinal chemistry efforts. 

Figure 7.8 Plot of IC50 as a function of substrate concentration (plotted as the ratio [SI/ Tm on the x-axis) for tight binding noncompetitive inhibitors when a = 5 (closed circles) and when a = 0.2 (open circles).

Photoaffinity labeling can be particularly useful when dealing with noncompetitive inhibitors, where the site of binding cannot be inferred from competition with specific substrate or cofactor molecules. 

A much more useful classification of inhibitors can be made on the basis of the mechanisms by which they act. Competitive inhibitors combine, with the enzyme at the same site as the substrate does, thus blocking the first step in the sequence. Noncompetitive inhibitors combine with the enzyme at some other site to give a complex that can still combine with the substrate, but the resultant ternary complex is unreactive. Uncompetitive inhibition results when the inhibitor and substrate combine with enzyme forms as in the following mechanism. 

A retro-l,3-dipolar cycloaddition followed by an 1,3-dipolar cycloaddition was used for a highly efficient total synthesis of (-)-histrionicotoxin (4-354) (HTX) by Holmes and coworkers [123]. HTX is a spiropiperidine-containing alkaloid which was isolated by Doly, Witkop and coworkers [124] from the brightly colored poison-arrow frog Dendrobates histrionicus. It is of great pharmacological interest as a noncompetitive inhibitor of acetylcholine receptors. 

---