Microsomes: The Preferred In Vitro Metabolic Model for Nonclinical Drug Research

Microsomes: The Preferred In Vitro Metabolic Model for Nonclinical Drug Research

Microsomes: Definition and Functions

In most cases, microsomes refer to spherical, vesicular membrane structures approximately 100 nm in diameter, formed by the self-fusion of fragmented endoplasmic reticulum (ER) during cell homogenization and differential centrifugation. These are heterogeneous assemblies comprising two fundamental components: ER membranes and ribosomes. Microsomes are primarily enriched with cytochrome P450 (CYP450) oxidase enzymes, which play a pivotal role in oxidative metabolism and are key enzymes in drug metabolism processes.

In vitro, microsomes retain the essential functions of the ER, including protein synthesis, protein glycosylation, and lipid biosynthesis, providing a versatile tool for studying various biochemical and pharmacological processes.

Figure 1: Microsomes (Source: Internet)

1. Key Organs in Drug Metabolism

Drug metabolism refers to the chemical changes a drug undergoes within the body, resulting in structural modifications. This process, also known as biotransformation, primarily occurs in organs such as the liver, kidneys, lungs, stomach, intestines, and skin. Among these, the liver is the principal site of drug metabolism, followed by the kidneys as the second most significant organ.

Within the liver, drug-metabolizing enzymes catalyze structural transformations that can generally be classified into two phases: Phase I metabolism and Phase II metabolism.

• Phase I Metabolism (Phase I Reaction): This phase involves oxidative, reductive, or hydrolytic reactions that generate intermediate products, often including electrophilic groups and oxygen radicals. These reactions may lead to hepatotoxicity.
• Phase II Metabolism (Phase II Reaction): This phase consists of conjugation reactions, which primarily serve to detoxify drugs. After metabolism, most drugs lose their pharmacological activity, though a minority may become active therapeutic agents.

The liver handles approximately 70%-80% of the total drug metabolism, underscoring its central role in biotransformation.

In addition to the liver, the kidneys contribute significantly to drug metabolism, accounting for about 10%-20% of total metabolic activity. The kidneys excrete drugs and their metabolites through filtration and secretion. However, their capacity for drug excretion is limited, which can lead to the accumulation of certain drugs and potential toxicity.

Beyond the liver and kidneys, other organs such as the intestinal enzyme systems and the lungs also play a role in influencing drug absorption, distribution, and metabolism, albeit to a lesser extent.

Figure 2: Reaction Catalyzed by Monooxygenase (Source: Internet)

2. Key Enzymes in Drug Metabolizing Organs

As discussed, drug metabolism primarily depends on the proper functioning of various enzyme systems in the liver, kidneys, gastrointestinal tract, and other organs. Understanding the enzymatic profiles of these organs is essential for a comprehensive study of drug metabolism processes.

Enzymes involved in drug metabolism are generally categorized into two classes: microsomal enzyme systems and non-microsomal enzyme systems.

• Microsomal Enzyme Systems:
  These enzymes are primarily localized in the lipophilic membranes of the endoplasmic reticulum in liver cells and other cells. The most important group of oxidative enzymes in hepatic microsomes is the hepatic microsomal mixed-function oxidase system, also known as monooxygenases (CYP450). These enzymes represent the primary pathway for drug metabolism in the body, capable of catalyzing a wide range of oxidative reactions. The biotransformation process catalyzed by these enzymes requires the involvement of cytochrome P450 (CYP450), coenzyme II, molecular oxygen, Mg²⁺, flavoproteins, non-heme iron proteins, and other cofactors.
  Additionally, UDP-glucuronosyltransferases (UGTs), a key component of Phase II metabolism, are also present on the luminal side of the endoplasmic reticulum, making UGT enzymes part of the microsomal system.
• Non-Microsomal Enzyme Systems:
  Also known as Type II enzymes, these include UGTs, sulfotransferases (SULTs), glutathione-S-transferases (GSTs), N-acetyltransferases (NATs), and amino acid conjugating enzymes. Non-microsomal enzymes primarily facilitate Phase II metabolism.

Beyond its physiological role in maintaining water and electrolyte balance and excreting endogenous and exogenous substances, the kidney is also a key organ for Phase I and Phase II metabolic biotransformations.

• Phase I Metabolism in the Kidney:
  Includes P450 enzymes, dehydrogenases, and various monooxygenases, though their concentrations and activities are significantly lower than those in the liver, making kidney Phase I metabolism less dominant.
• Phase II Metabolism in the Kidney:
  Primarily involves UGTs, SULTs, GSTs, NATs, and amino acid conjugating enzymes, playing a major role in renal drug metabolism.

The intestine, as one of the largest digestive organs, also plays a critical role in drug metabolism. In the intestinal tract, many drugs undergo metabolic reactions that transform them into more excretable and eliminable metabolites. These metabolic processes occur via two pathways:

1. Enzymatic metabolism within intestinal epithelial cells, which involves enzymes such as CYP450, UGTs, and lipases.
2. Microbial-mediated metabolism by intestinal microbiota.

Together, these enzymatic systems in different organs ensure the efficient metabolism and clearance of drugs, highlighting the complexity and integration of drug metabolism pathways in the body.

However, with the continuous advancements in medicine, inhaled drugs have garnered significant attention in recent years due to their rapid absorption, quick onset of action, and ability to bypass first-pass metabolism. Unlike conventional oral drugs, inhaled formulations deliver medication directly to the pulmonary tissues, avoiding hepatic first-pass effects. This highlights the importance of investigating drug metabolism within the lungs for inhaled formulations.

The lungs contain a variety of drug-metabolizing enzymes, including P450 enzymes, hydrolases, conjugation enzymes, monoamine oxidases, and flavin-containing monooxygenases. Among these, pulmonary P450 enzymes play a critical role in the biotransformation of xenobiotics, the inactivation of inhaled chemical carcinogens, and the detoxification of pulmonary toxins.

In summary, drug metabolism within the body is typically a coordinated process involving multiple organs and enzyme systems. Thus, during the early nonclinical drug development phase, selecting appropriate in vitro models is crucial for elucidating metabolic pathways and identifying key metabolizing enzymes.

3. In Vitro Drug Metabolism Models: Microsomes

Compared to in vivo metabolism studies, in vitro studies minimize interference from physiological factors, allowing for the direct observation of interactions between drugs and enzymes. Consequently, in vitro metabolism models have become the preferred choice during early drug development. Common models for in vitro drug metabolism studies include microsomes, S9 fractions, cytosol, tissue homogenates, and primary cells. Given that the liver is the principal site of drug metabolism, hepatic cells and their subcellular components—such as liver microsomes, liver S9 fractions, liver tissue homogenates, and liver cytosol—are the primary models for studying drug metabolism.

Microsomes, specifically, are vesicular membrane structures derived from fragmented endoplasmic reticulum that self-assemble during cell homogenization and differential centrifugation. They are widely distributed in organs such as the liver, kidneys, intestines, and lungs. As microsomes contain Phase I enzymes like cytochrome P450 (CYP450) and Phase II enzymes like UGTs and SULTs, they encompass a wide range of metabolic pathways for various drugs. Thus, selecting tissue-specific microsomes is a critical step in in vitro drug metabolism research.

Furthermore, according to the Technical Guidelines for Nonclinical Pharmacokinetic Studies of Drugs, experimental animals such as mice, rats, rabbits, guinea pigs, dogs, miniature pigs, and monkeys are commonly used. For innovative drugs, at least two species should be employed, with one being a rodent and the other a non-rodent species. Beyond animal species, humanized materials—such as human liver microsomes—are also emphasized as key tools for nonclinical ADME studies. Consequently, selecting microsomes from multiple species, including humans, is a pivotal consideration in drug metabolism research.

In light of this, IPHASE, as a leading provider of in vitro biological reagents, has successfully developed microsome products derived from various tissues of multiple species, including humans, monkeys, dogs, rats, and mice. These products offer a wide range of options for studies on species differences, metabolic stability, P450 inhibition, and metabolic enzyme phenotyping.

With strict quality control measures, IPHASE ensures product reliability, helping clients save time and improve efficiency. IPHASE microsomes are the ideal choice for in vitro nonclinical research.

Advantages of IPHASE Microsome Products:

• Compliance: All tissues used in production are sourced through certified channels with clear traceability.
• Safety: Production tissues are tested for pathogens to ensure product quality and safety.
• High Quality: Products undergo rigorous internal quality control, ensuring large batch sizes with minimal inter-batch variability.
• Customizability: Tailored microsome products from specific species or tissues are available to meet unique client requirements.

Leveraging years of R&D expertise, IPHASE has launched high-end research reagents across multiple fields and categories. These products serve as essential tools for early-stage drug development, offering new materials, methods, and techniques for exploring life sciences. They also provide convenient solutions for genetic toxicity studies in food, pharmaceuticals, and chemicals.

We look forward to supporting researchers with our innovative and reliable products!



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