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This is an RSS file. You can use it to subscribe to this data in your favourite RSS reader or to display this data on your own website or blog.Irreversible Enzyme Inhibition Kinetics and Drug–Drug Interactions
This chapter describes the types of irreversible inhibition of drug-metabolizing enzymes and the methods commonly employed to quantify the irreversible inhibition and subsequently predict the extent and time course of clinically important drug–drug interactions. (Source: Springer protocols feed by Pharmacology/Toxicology)
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Multienzyme Kinetics and Sequential Metabolism
Enzymes are the catalysts of biological systems and are extremely efficient. A typical enzyme accelerates the rate of a reaction by factors of at least a million compared to the rate of the same reaction in the absence of the enzyme. In contrast to traditional catalytic enzymes, the family of cytochrome P450 (CYP) enzymes are catalytically promiscuous, and thus they possess remarkable versatility in substrates. The great diversity of reactions catalyzed by CYP enzymes appears to be based on two unique properties of these heme proteins, the ability of their iron to exist under multiple oxidation states with different reacti...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Consideration of the Unbound Drug Concentration in Enzyme Kinetics
The study of enzyme kinetics in drug metabolism involves assessment of rates of metabolism and inhibitory potencies over a suitable concentration range. In all but the very simplest in vitro system, these drug concentrations can be influenced by a variety of nonspecific binding reservoirs that can reduce the available concentration to the enzyme system under investigation. As a consequence, the apparent kinetic parameters that are derived, such as K
m or K
i, can deviate from the true values. There are a number of sources of these nonspecific binding depots or barriers, including membrane pe...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Enzyme Kinetics of Conjugating Enzymes: PAPS Sulfotransferase
The sulfotransferase (SULT) enzymes catalyze the formation of sulfate esters or sulfamates from substrates that contain hydroxy or amine groups, utilizing 3′-phosphoadenosyl-5′-phosphosulfate (PAPS) as the donor of the sulfonic group. The rate of product formation depends on the concentrations of PAPS and substrate as well as the sulfotransferase enzyme; thus, if PAPS is held constant while varying substrate concentration (or vice versa), the kinetic constants derived are apparent constants. When studied over a narrow range of substrate concentrations, classic Michaelis–Menten kinetics can be observed wit...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Enzyme Kinetics of Uridine Diphosphate Glucuronosyltransferases (UGTs)
Glucuronidation, catalyzed by uridine diphosphate glucuronosyltransferases (UGTs), is an important process for the metabolism and clearance of many lipophilic chemicals, including drugs, environmental chemicals, and endogenous compounds. Glucuronidation is a bi-substrate reaction that requires the aglycone and a cofactor, UDPGA. Accumulating evidence suggests that the bi-substrate reaction follows a compulsory-order ternary mechanism. To simplify the kinetic modelling of glucuronidation reactions in vitro, UDPGA is usually added to incubations in large excess. Many factors have been shown to influence UGT activity and kine...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Principles and Experimental Considerations for In Vitro Transporter Interaction Assays
Drug transporters are now universally acknowledged as important determinants of the absorption, distribution, metabolism and excretion of both endogenous and exogenous compounds. Altered transporter function, whether due to genetic polymorphism, DDIs, disease, or environmental factors such as dietary constituents, can result in changes in drug efficacy and/or toxicity due to changes in circulating or tissue levels of either drugs or endogenous substrates. (Source: Springer protocols feed by Pharmacology/Toxicology)
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Enzyme Kinetics of Oxidative Metabolism: Cytochromes P450
The cytochrome P450 enzymes (CYPs) are the most important enzymes in the oxidative metabolism of hydrophobic drugs and other foreign compounds (xenobiotics). The versatility of these enzymes results in some unusual kinetic properties, stemming from the simultaneous interaction of multiple substrates with the CYP active site. Often, the CYPs display kinetics that deviate from standard hyperbolic saturation or inhibition kinetics. Non-Michaelis–Menten or “atypical” saturation kinetics include sigmoidal, biphasic, and substrate inhibition kinetics (see
Chapter 3
...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Enzyme Kinetics, Inhibition, and Regioselectivity of Aldehyde Oxidase
The aldehyde oxidase (AO) enzyme family plays an increasing role in drug development. However, a number of compounds that are AO substrates have failed in the clinic because the clearance or toxicity is underestimated by preclinical species. Human AO is much more active than rodent AO, and dogs do not have functional AO. While AOs normally make non-reactive metabolites such as lactams, the metabolic products often have much lower solubility that can lead to renal failure. While an endogenous substrate for the oxidation reaction is not known, electron acceptors for the reductive part of the reaction include oxygen and nitri...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Rationalizing Underprediction of Drug Clearance from Enzyme and Transporter Kinetic Data: From In Vitro Tools to Mechanistic Modeling
Over the years, there has been an increase in the number and quality of available in vitro tools for the assessment of clearance. Complexity of data analysis and modelling of corresponding in vitro data has increased in an analogous manner, in particular for the simultaneous characterization of transporter and metabolism kinetics, together with intracellular binding and passive diffusion. In the current chapter, the impact of different factors on the in vitro–in vivo extrapolation of clearance will be addressed in a stepwise manner, from the selection of the most adequate in vitro system and experimental design/condi...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
A Structural Model for the Mass Action Kinetic Analysis of P-gp Mediated Transport Through Confluent Cell Monolayers
The structural model for P-gp mediated transport across confluent cell monolayers uses the generally accepted mass action reactions for P-gp binding and efflux, together with the known structural parameters for P-gp (large substrate binding site accessible from the membrane) and the apical plasma membrane in which it resides (lipid bilayer partition coefficient of substrate and volume of apical plasma membrane allow estimation of substrate concentration at binding site). The model considers binding of substrate to P-gp from within the inner leaflet of the apical membrane, with an on rate constant, k
1 (M&mi...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Systems Biology Approaches to Enzyme Kinetics: Analyzing Network Models of Drug Metabolism
Intracellular drug metabolism involves transport, bioactivation, conjugation, and other biochemical steps. The dynamics of these steps are each dependent on a number of other cellular factors that can ultimately lead to unexpected behavior. In this review, we discuss the confounding processes and coupled reactions within bioactivation networks that require a systems-level perspective in order to fully understand the time-varying behavior. When converting known in vitro characteristics of drug–enzyme interactions into descriptions of cellular systems, features such as substrate availability, cell-to-cell variability, ...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Variability in Human In Vitro Enzyme Kinetics
There are many factors which are known to cause variability in human in vitro enzyme kinetic data. Factors such as the source of enzyme and how it was prepared, the genetics and background of the donor, how the in vitro studies are designed, and how the data are analyzed contribute to variability in the resulting kinetic parameters. It is important to consider not only the factors which cause variability within an experiment, such as selection of a probe substrate, but also those that cause variability when comparing kinetic data across studies and laboratories. For example, the artificial nature of the microsomal lipid me...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Sources of Interindividual Variability
The efficacy, safety, and tolerability of drugs are dependent on numerous factors that influence their disposition. A dose that is efficacious and safe for one individual may result in sub-therapeutic or toxic blood concentrations in other individuals. A major source of this variability in drug response is drug metabolism, where differences in pre-systemic and systemic biotransformation efficiency result in variable degrees of systemic exposure (e.g., AUC, C
max, and/or C
min) following administration of a fixed dose. (Source: Springer protocols feed by Pharmacology/Toxicology)
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Case Study 1. Practical Considerations with Experimental Design and Interpretation
At some point, anyone with knowledge of drug metabolism and enzyme kinetics started out knowing little about these topics. This chapter was specifically written with the novice in mind. Regardless of the enzyme one is working with or the goal of the experiment itself, there are fundamental components and concepts of every experiment using drug metabolism enzymes. The following case studies provide practical tips, techniques, and answers to questions that may arise in the course of conducting such experiments. Issues ranging from assay design and development to data interpretation are addressed. The goal of this section is ...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
Case Study 2. Practical Analytical Considerations for Conducting In Vitro Enzyme Kinetic Studies
The objective of this case study is to answer the following questions:
(a)
Why is it necessary to determine lower and upper limits of quantification (LLOQ and ULOQ, respectively) of a bioanalytical assay specifically for enzyme kinetic assays? How do you utilize LLOQ and ULOQ to correctly interpret your kinetic data?
(b)
Why should one use a linear fit and not a quadratic fit for standard curves?
...
Source: Springer protocols feed by Pharmacology/Toxicology - February 14, 2014 Category: Drugs & Pharmacology Source Type: news
