Pharmacokinetics and Drug Metabolism

National Center For Advancing Translational Sciences

Investigators

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Abstract

The DMPK lab was initially established within the Therapeutic Development Branch in the Division of Preclinical Innovation (DPI) in 2011. It became the DMPK Core in Oct. 2021. The mission of the DMPK Core is to address issues related to drug absorption, biodistribution and elimination via metabolism or excretion. The DMPK group has supported diverse projects across DPI, contributing significantly to all stages of translational research at NCATS, from early probe development in drug discovery to Phase II clinical trials. Our major capabilities include: 1. In vitro ADME high-throughput screening (Tier I HTS assays) on solubility, permeability and microsomal stability for all small molecule compounds registered at NCATS (3000 compounds/year). 2. Customized in vitro ADME assays (Tier II assays) as required by each projects specific needs. The common Tier II assays include metabolic stability in different species, metabolite identification (MetID), aldehyde oxidase stability in cytosol fraction, plasma stability for prodrugs and biologics, blood/plasma partition, CYP inhibition, and transporter assessments in Caco-2 and MDKC cells. 3. PK studies in lab animals and bioanalytical measurements of drug concentrations in different biological fluids (e.g., blood, plasma, urine, bile) and tissue extracts. 4. UPLC-MS/MS and high-resolution accurate mass spectrometry for quantitation of small molecules and peptides, and structure identification of metabolites. 5. Bioanalytical method development for therapeutic macromolecules, such as recombinant human proteins and engineered proteins. 6. Pharmacokinetic parameter calculation, modeling and simulation. Scientific data generated from our lab have been used for novel target validations, drug discovery and development for COVID-19, and research publications. Examples of the DMPK group contributions to recent projects within the Therapeutic Development Branch include: 1. Development of In Silico ADME Models as a Powerful Translational Research Tool for Drug Discovery: Characterization of in vitro ADME properties of a novel compound is very important in drug discovery research as it will guide structure optimization and lead selection. We have developed high-throughput assays for key ADME properties (Tier I ADME assays), such as aqueous solubility, membrane permeability and hepatic metabolic stability in microsomes. To date, we have collected data for over 30,000 compounds synthesized or registered at DPI/NCATS from Tier I ADME assays. We also collected data on thousands of compounds from Tier II assays (e.g. human CYP450 enzymes). To ensure data quality, we use controls in each plate and monitor the performance of these controls for all plates. We calculate Minimum Significant Ratio (MSR) for controls, a statistical parameter that characterizes the reproducibility of an assay, to evaluate assay performance. These high quality datasets allow us to develop in silico models for these ADME properties. These in silico models are useful tools for medicinal chemists to design new drug-like molecules, which will potentially reduce the number of compounds to be synthesized during drug discovery, save valuable resources, minimize chemical wastes to ultimately help to accelerate the drug discovery process. The assay protocols for ADME Tier I and Tier II assays have been published in PubChem (see list below, as of August 1, 2023), and the corresponding in silico models can be found at NCATS OpenData Portal: - Kinetic Aqueous Solubility; PubChem BioAssay AID: 1645848; https://opendata.ncats.nih.gov/adme/models/solubility - PAMPA Permeability (pH 7.4); PubChem BioAssay AID: 1508612; https://opendata.ncats.nih.gov/adme/models/pampa - PAMPA Permeability (pH 5); PubChem BioAssay AID: 1645871; https://opendata.ncats.nih.gov/adme/models/pampa - Tier I Rat Liver Microsome Stability; PubChem BioAssay AID: 1508591; https://opendata.ncats.nih.gov/adme/models/rlm - Mouse Cytosol Stability; PubChem BioAassay AID: 1508604 - Human Cytosol Stability; PubChem BioAssay AID: 1508603 - Human CYP3A4; PubChem BioAssay AID: 1645841; https://opendata.ncats.nih.gov/adme/models/cyp450 - Human CYP2C9; PubChem BioAssay AID: 1645842; - Human CYP2D6; PubChem BioAssay AID: 1645840; - Human CYP3A7 Since the launch of our website on the In Silico ADME Models in December 2021, we have 4300 registered users from more than 84 countries. In 2023, we have one new publication on the CYP3A7 modeling. 2. Development of An Automated TEER-96 Device for Transporter Studies: P-glycoproteins (Pgp) play important roles in oral absorption and tissue distribution. To study whether a drug candidate is a Pgp substrate or an inhibitor, we use Caco-2 and MDCK cell lines in 96-tranwell plates. During the cell culture, we measure the trans epithelial electrical resistances (TEER) for each transwell to ensure the integrity of the cell monolayer. Most commercial TEER measurement devices are manual ones for 6- or 24-transwells. To increase the throughput rate in compound screening and reduce the variations caused by the manual measurement, NCATS has been working with Applied BioPhysics via a SBIR funding mechanism to develop an automated TEER measurement device for 96-transwells. The DMPK Core scientists actively participate in the validation and modification of this automated device. Currently the device is in a beta phase release, and several pharma companies are interested in testing it for their research. A Poster was presented in this years SLAS conference and a manuscript has been submitted. 3. ADME/PK Studies for Antiviral Drug Candidates The DMPK Core contributes significantly to the drug discovery and development for antiviral drugs. These efforts can be illustrated in two examples: a. Drug discovery research for niclosamide prodrugs: Niclosamide (Nc) is an FDA-approved anthelmintic drug that was recently identified from a drug repurposing screening to possess antiviral activity against SARS-CoV-2. However, due to the low solubility and permeability of Nc, its in vivo efficacy was limited by its poor oral absorption. NCATS medicinal chemists have been working on the prodrug approach to increase the solubility and permeability of Nc. The DMPK Core characterized the physical/chemical and PK properties of these prodrugs. A manuscript has been published recently (PMID:37113753. Yang et al. Use of physiological based pharmacokinetic modeling for cross-species prediction of pharmacokinetic and tissue distribution profiles of a novel niclosamide prodrug). b. Development of GS-441524 as an oral drug for COVID-19 treatment: GS-441524 is the active metabolite of Remdesivir developed by Gilead. Since Remdesivir has to be administered intravenously, it is highly desirable to investigate the feasibility of development of GS-441524 as an oral drug that could have a broader use for patients, especially for those in countries with poor medical facilities. We conducted a series of in vitro ADME and in vivo PK studies for GS-441524. The compound is current in the drug development stage. A publication was released (PMID: 36052127. Wang et. al. Preclinical pharmacokinetics and in vitro properties of GS-441524, A potential oral drug candidate for COVID-19 treatment).