Handbook of Drug Metabolism, Third Edition

Paul G. Pearson, Larry C. Wienkers, Paul G. Pearson, Larry C. Wienkers

• Publisher: CRC Press
• Year: 2019
• Print ISBN:
  9780367779764
  9781482262032
• eBook ISBN:
  9780429529825
• Edition: 3
• Topic: Medicina
• Subtopic: Teoria, pratica e riferimenti medici

This book continues to be the definitive reference on drug metabolism with an emphasis on new scientific and regulatory developments. It has been updated based on developments that have occurred in the last 5 years, with new chapters on large molecules disposition, stereo-selectivity in drug metabolism, drug transporters and metabolic activation of drugs. Some chapters have been prepared by new authors who have emerged as subject area experts in the decade that has passed since publication of the first edition.

Section II

Factors Which Affect Drug Metabolism

6

Non-CYP Drug Metabolizing Enzymes and Their Reactions

Shuguang Ma, Ryan H. Takahashi, Yong Ma, Sudheer Bobba, Donglu Zhang, and S. Cyrus Khojasteh

CONTENTS

Introduction

Flavin-Containing Monooxygenases

Subcellular Location and Organ Distribution

Multiplicity and Species Differences

Catalytic Mechanism

Substrates and Inhibitors

Reactions and Relevance to Human Drug Metabolism

Monoamine Oxidases

Subcellular Location and Organ Distribution

Catalytic Mechanism

Substrates and Inhibitors

Reactions and Relevance to Human Drug Metabolism

Alcohol Dehydrogenases

Subcellular Location and Organ Distribution

Multiplicity and Species Differences

Catalytic Mechanism

Substrates, Inhibitors, and Inducers

Reactions and Relevance to Human Drug Metabolism

Aldehyde Dehydrogenases

Subcellular Location and Organ Distribution

Multiplicity and Species Differences

Catalytic Mechanism

Substrates, Inhibitors, and Inducers

Reactions and Relevance to Human Drug Metabolism

Peroxidases

Catalytic Mechanism

Reactions and Relevance to Human Drug Metabolism

Aldo-keto Reductases

Subcellular Location and Organ Distribution

Multiplicity and Species Differences

Catalytic Mechanism

Substrates and Inhibitors

Reactions and Relevance to Human Drug Metabolism

Epoxide Hydrolases

Subcellular Location and Organ Distribution

Multiplicity and Species Differences

Catalytic Mechanism

Substrates and Inhibitors

Reactions and Relevance to Human Drug Metabolism

Carboxyl Esterases

Subcellular Location and Organ Distribution

Multiplicity and Species Differences

Catalytic Mechanism

Substrates and Inhibitors

Reactions and Relevance to Human Drug Metabolism

γ-Glutamyl transpeptidase

Reactions and Relevance to Human Drug Metabolism

Cathepsins

Reactions and Relevance to Human Drug Metabolism

Glutathione S-transferases

Subcellular Location and Organ Distribution

Catalytic Mechanism

Substrates and Inhibitors

Reactions and Relevance to Human Drug Metabolism

Sulfotransferases

Subcellular Location and Organ Distribution

Multiplicity and Species Differences

Catalytic Mechanism

Substrates and Inhibitors

Reactions and Relevance to Human Drug Metabolism

N-acetyltransferases

Subcellular Location and Organ Distribution

Catalytic Mechanism

Multiplicity and Species Differences

Substrate, Inhibitors, and Inducers

Reactions and Relevance to Human Drug Metabolism

Conclusions

Acknowledgment

References

Introduction

Metabolism is the major elimination pathway of a drug from the body initiated by drug metabolizing enzymes (DMEs). DMEs are present mainly in the liver, intestine, and blood and facilitate the excretion of xenobiotics from the body by converting lipophilic drugs into hydrophilic compounds. Cytochrome P450 enzymes (CYPs) are responsible for the clearance of a majority of drugs on the market, and, accordingly, much emphasis has been placed on understanding the role of CYPs in metabolism, clearance, drug-drug interactions, and oral bioavailability [1].

The widespread use of metabolic stability screening using microsomal fractions or hepatocytes for oxidative and glucuronic acid conjugative metabolism at the early discovery stage has greatly increased the number of drug candidates with a high level of metabolic stability. This screening strategy, however, has forced new chemical entities (NCEs) into chemical spaces that rely on non-CYP enzymes for clearance, and therefore, the importance of non-CYP enzymes in the elimination of drugs has been increasingly recognized in drug discovery and development [2,3,4]. In a recent survey of a set of 125 small molecule drugs approved by the United States Food and Drug Administration (FDA) from 2006 to 2015, approximately 30% of the metabolism of these drugs is carried out by non-CYP enzymes [5]. These non-CYP enzymes can also modulate efficacy, contribute to detoxification, or produce therapeutically active or reactive/toxic metabolites [6,7]. Here, we discuss the non-CYP enzymes involved in drug metabolism reactions (except for enzymes discussed in Chapters 4 and 5) and their subcellular locations, organ distributions, mechanisms of reactions, and typical substrates and inhibitors.

Flavin-Containing Monooxygenases

The flavin-containing monooxygenase (FMO) (EC 1.14.13.8) enzymes are NADPH-dependent, membrane bound to the endoplasmic reticulum (ER), and have molecular weights of approximately 60 kDa. FMOs metabolize a wide range of substrates by oxidizing nitrogen, sulfur, phosphorous, and selenium atoms. The source of the added oxygen atom is molecular O₂.

The role of FMO is often underestimated in the calculation of total clearance because several of the reactions they carry out are similar to those mediated by CYPs. In addition, CYPs use the same cofactor and are also located on the ER [8]. FMO-mediated reactions can be distinguished from those mediated by CYPs by the following observations:

1. FMO activity is optimal at a pH of >9 while CYP activity is optimal at a pH of 7.4.

2. All FMOs, except for FMO2, are thermally unstable when pre-incubated in the absence of NADPH.

3. 1-Aminobenzotriazole (ABT) inactivates CYP enzymes in a time-dependent manner but does not inhibit FMOs.

4. Inhibitory antibodies to cytochrome P450 reductase will impact CYP-dependent processes, but not those that are FMO-dependent.

5. Detergents (such as Triton-X 100) inhibit CYP enzymes but have little effect on FMO activity.

6. Recombinant enzymes can be used to distinguish between FMO- and CYP-catalyzed reactions.

7. FMOs are rarely induced, but many CYPs are inducible.

Subcellular Location and Organ Distribution

FMOs are distributed differently in organs throughout the body depending on the isoform (Table 6.1) [9]. In humans, the five functional forms of FMO (FMO1 through FMO5) share 50%–55% sequence identity. Even though FMO1 is the major form in many animals, in humans, this isoform is found only in fetal livers and is absent in adults [9]. FMO1 has a shallow substrate binding channel and, therefore, broad substrate specificity. The highest concentration of FMO2 is in lung tissue, followed by the kidney where the concentration is 7-fold less. FMO3 is the major isoform in human liver at a concentration of 46 pmol/mg [10] but is present at much lower amounts in the lung and kidney (<5%). FMO3 has a deep substrate binding channel (8–10 Å) and, therefore, a narrower substrate specificity than FMO1. FMO4 and FMO5 play minor roles in drug metabolism.

In humans, trimethylamine, a foul-smelling chemical derived from dietary sources, such as choline and carnitine, is metabolized by FMO3 to an odorless N-oxide metabolite. FMO3 deficiency leads to trimethylaminuria (“fish-like odor syndrome”) [11].

TABLE 6.1

FMO Distribution in Different Species

Form	Mouse	Rat	Monkey	Human
FMO1	Kidney	Liver, kidney	Kidney	Kidney >> lung, small intestine >> liver
FMO2	Lung	Lung	Lung	Lung >> kidney > liver, small intestine
FMO3	Liver, kidney	Kidney	Liver, kidney	Liver >> lung > kidney >> small intestine

Multiplicity and Species Differences

Pronounced differences in FMO expression exist amongst preclinical species [12–13]. The flavin adenine dinucleotide (FAD) and NADPH binding domains, with 4–32 and 186–213 residues, respectively, are the conserved regions across species.

Young female mice have very high expressions of FMO3 and FMO5, which most closely resembles FMO expression in the adult human liver. The FMO3 activity in young female rats is 5–10 fold higher than that in male rats. Adult rats have relatively low FMO3 and FMO5 expressions and a similar FMO1 expression to humans.

Catalytic Mechanism

The overall reaction of FMO-mediated metabolism involves the use of FAD as a prosthetic group to initiate a two-electron oxidation (Figure 6.1). The steps of this reaction are described below:

1. At resting state, the enzyme is present as a 4α-hydroperoxyflavin (FAD-OOH). The distal oxygen is electrophilic.

2. A nucleophilic substrate (X:) attacks the distal oxygen of FAD-OOH and results in formation of an oxygenated product (O-X) and 4α-hydroxyflavin (FAD-OH).

3. The metabolite is released, and FAD-OH losses a water molecule to form FAD. This is thought to be the rate-limiting step of the reaction.

4. FAD receives an electron from NADPH to form FADH₂ (not shown). This is further oxidized by O₂ to form FAD-OOH. This step returns FMO to its resting state.

FIGURE 6.1 Reaction mechanism for oxidation by flavin-containing monooxygenases.

Substrates and Inhibitors

FIGURE 6.2 Examples of substrates and inhibitors of flavin-containing monooxygenase.

FMOs...

Table of contents

• Cover
• Half Title
• Title Page
• Copyright Page
• Dedication
• Table of Contents
• Preface
• Editors
• Contributors
• Section I Fundamental Aspects of Drug Metabolism
• Section II Factors Which Affect Drug Metabolism
• Section III Technologies to Study Drug Metabolism
• Section IV Applications of Metabolism Studies in Drug Discovery and Development
• Index