Drug Metabolism in Drug Discovery and Preclinical Development

*Benjamin Mann, Roger Melton and David Thompson*

## **Abstract**

Drug metabolism or more generally, xenobiotic metabolism, is the biotransformation of exogenous compounds by living organisms, usually through specialized enzymatic systems. The metabolism of experimental therapeutics is an important aspect of pharmacology and translational medicine as the rate and the interindividual variability of drug metabolism can determine the duration and/or efficacy of a drug's pharmacologic action. Since the introduction of metabolites in safety testing guidance by the Food and Drug Administration, major changes have occurred in the experimental methods for the identification and quantification of metabolites, evaluation of metabolites, and the timing of critical nonclinical studies to generate this information.

**Keywords:** Drug Metabolism, Drug Discovery, Preclinical Development, Lead Selection, Lead Optimization, Drug Candidate Selection, Metabolite Safety Testing, CYP450 Inhibition, CYP450 Induction, Soft Spot Analysis, Metabolite Profiling, Membrane Drug Transporters

### **1. Introduction**

Drug metabolism is a drug-clearing event from systemic circulation influencing efficacy and toxicity in humans and preclinical species. The primary endpoints of enzymatic metabolism studies in both the drug discovery and preclinical stages are to resolve metabolic stability, identify and quantify primary metabolites, identify metabolic routes, and measure the potential for drug–drug interactions (DDI) [1]. The majority of *in vivo* and *in vitro* drug metabolism assessments center around hepatic models as the liver is the main metabolizing organ; however, drug metabolism does occur in other organs and tissues (e.g., lungs, kidneys, and intestine). As a result, drug discovery and preclinical development investigations include evaluation of drug metabolism in both hepatic and extra-hepatic models.

Over the past several decades *in vitro* and *in vivo* methodologies to quantitatively measure the absorption, distribution, metabolism, and excretion (ADME) properties of a chemical entity have matured to the point to where these properties can be reliably modeled and simulated in order to predict the general disposition of the chemical entity (or class of molecules) across multiple species and into humans [2]. As a result, approximations of oral bioavailability (F), total systemic clearance (CL), volume of distribution (Vd), and half-life (t1/2) can be predicted and subsequently evaluated. These intrinsic properties of the chemical entity with

### *Drug Metabolism*

consideration of the physiological processes can be used to assess the disposition of single and multiple dose escalation studies, identify potentially saturable hepatic pathways, and assess the formation of metabolites. From this information, potential DDIs can be predicted and investigated.

In order to effectively utilize the drug metabolism and pharmacokinetic (DMPK) information generated from *in vitro* and *in vivo* studies, it is important to have a proper understanding of when in the development timeline the data should be generated. Additionally, a thorough understanding of the clearance mechanism of a chemical entity can help lessen the need for some downstream *in vivo* studies potentially saving the pharmaceutical organization hundreds of thousands of dollars. While the timing of specific investigations may change due to the nature of the chemical entity being tested, the following general DMPK chronology and endpoints should be considered during discovery and development [3, 4]. The bullets relating to drug metabolism are bolded for emphasis and will be expanded in greater detail in the following pages.

	- Plasma protein binding, red blood cell partitioning, and intestinal permeability (Caco-2) assessments
	- *In vitro* **metabolic stability screening using hepatocytes, microsomes, and S9 fractions**
	- *In vivo* rodent PK studies
	- Investigate the *in vitro*:*in vivo* correlation (IVIVC) of CL and t1/2 and identify the series with problematic kinetics (e.g., high CL or low t1/2)
	- Identification of the relevant tissues where the chemical entity and metabolites may be sequestered
	- Pharmacokinetic studies in rodent and nonrodent species
	- **Identification of clinically relevant DDIs (***In vitro* **assessment of P450 induction and P450 inhibition)**
	- **Screening Cocktail DDI Study**
	- *In vitro* **soft spot analysis/metabolic identification to identify metabolically liable sites on investigated chemical entities**
	- **Drug transporter and tissue sequestration assessments**
	- Pharmacokinetic-Pharmacodynamic (PK-PD; exposure-effect) modeling
	- **Mass balance study in rodents and nonrodents in order to support nonrodent toxicology species selection**

4.Metabolite Safety Testing (Post FIH and concurrent with Phase 2 trials)

• **Human specific metabolites or metabolites with exposures at higher levels in humans than in any of the animal test species are assessed in preclinical species.**
