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Principal Investigator: Xin Xu
Organization: NATIONAL CENTER FOR ADVANCING TRANSLATIONAL SCIENCES
Fiscal Year: 2024
Award: $5,131,109
Funding agency: National Center for Advancing Translational Sciences
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 Core has supported projects across NCATS intramural research labs, contributing 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 kinetic aqueous solubility, membrane permeability and microsomal stability for all small molecule compounds synthesized by NCATS MedChem Scientists (3000 compounds/year).
2. Conducting customized in vitro ADME assays (Tier II assays) as required by each project’s 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, plasma protein binding, CYP inhibition, and transporter assessments in Caco-2 and MDKC cells.
3. Performing 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. Developing sensitive bioanalytical methods with UPLC-MS/MS and high-resolution accurate mass spectrometry for quantitation of small molecules (including peptides) in biologic fluids and elucidation of metabolite structures.
5. Bioanalytical method development for therapeutic macromolecules, such as recombinant human proteins, engineered proteins and gene therapies.
6. Pharmacokinetic parameter calculation, modeling and simulation.
Scientific data generated from our lab have been used for novel target validations, drug discovery and development, in silico ADME model development and other research publications. Examples of the DMPK Core contributions to recent NCATS Intramural research projects include:
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 important in drug discovery as it guides structure optimization and lead selection. We developed high-throughput assays for key ADME properties (Tier I ADME assays), such as aqueous solubility, PAMPA membrane permeability and hepatic metabolic stability in microsomes. We have collected data for 30,000 compounds synthesized or registered at 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 the quality of the datasets, 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 by using Machine Learning or other AI modeling approaches. 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 and minimize chemical wastes. Ultimately it could help to accelerate the drug discovery process. The assay protocols for ADME Tier I and Tier II assays have been published in PubChem (as of July 29, 2024), and the corresponding in silico models can be found at NCATS OpenData Portal.
Kinetic Aqueous Solubility; PubChem AID 1645848
PAMPA Permeability (pH 7.4); AID 1508612
PAMPA Permeability (pH 5); AID 1645871
PAMPA-BBB; AID: 1845228
Rat Liver Microsome Stability; AID: 1508591
Human Liver Microsome Stability
Mouse Cytosol Stability; AID: 1508604
Human Cytosol Stability; AID: 1508603
Human CYP3A4; AID: 1645841
Human CYP2C9; AID: 1645842
Human CYPY2D6; AID: 1645840
Human CYP3A7; AID: 196359
Since the launch of our website on the In Silico ADME Models in December 2021, we have 5900 registered users from more than 84 countries. In addition, two manuscripts were published on our new models (PMID: 37676606; 38108064).
2. Development of An Automated TEER-96 Device for Transporter Studies:
In collaboration with the Applied BioPhysics, an automated transepithelial electrical resistance (TEER) measurement device for a 96-transwell plate was developed with the NCATS support through SBIR grant (HHSN271201800635P). The system is able to measure TEER of a transwell system in a continuous way under incubation conditions, which was not previously possible. To showcase this new avenue of evaluation of biological membranes, we conducted a study with orally available toxicants from the NCATS TOX21 Library on Caco-2 cells (a cell line of human intestinal origin) to evaluate if and how TEER values correlate with the membrane permeability, cell viability, and tight-junction gene expression. This work is conducted with an NCATS Opportunities Committee Grant and in collaboration with TOX21 and the National Center for Toxicological Research (NCTR). The work resulted in a manuscript that was published in SLAS Technology (PMID: 37923083).
3. ADME/PK Studies for Drug Discovery and Development
The DMPK lab contributes significantly to the drug discovery and development of NCATS project portfolio. In FY2024, we successfully filed two Investigational New Drug (IND) application with US FDA:
1. PTH-IA (parathyroid hormone inverse agonist)
A Phase 1/2 Open-label First-in-Human Dose-Escalating Safety and Efficacy Study Evaluating Subcutaneous Administration of PTH-IA in Adults and Children with Jansen’s Metaphyseal Chondrodysplasia (JMC)
2. GS-441542 (an active metabolite of Remdesivir)
First-in-Human Study of Orally Administered GS-441524 for COVID-19
In both projects, we developed sensitive bioanalytical methods to measure PTH-IA and GS-441524 in plasma samples. We also developed anti-drug antibody (ADA) assay for PTH-IA to assess immunogenicity of this peptide. These assays were used in the GLP animal toxicity studies and will be used in the planned human clinical trial. We conducted both in vitro and in vivo studies to fully characterize ADME and PK properties of these new drug candidates. Our data were presented in national conferences:
1) Wang AQ, et al., Ultra-Performance Liquid Chromatography-Tandem Mass Spectrometry Method Development and Pharmacokinetics of PTH-IA in Mice. AAPS PharmSci 360, Orlando, FL, Oct 22 - 25, 2023.
2) Nduwumwami A, et al., Thiol-based Conjugation for the Generation of ADA Bridging Assay Reagents to Detect Antibodies to PTH-IA in Human Plasma. ASC Fall, Denver, CO, Aug 18-22, 2024.
3) Wang AQ, et al., UPLC-MS/MS method development for GS-441524 and preclinical pharmacokinetic studies. HPLC 2022: 50th International symposium and exposition on HPLC and related techniques, San Diego, CA, Jun 18 - 23, 2022.
FY24 is also marked by a successful FDA approval of Emflaza, a treatment for Duchenne muscular dystrophy (DMD), in that the NCATS DMPK lab contributed extensively to the in vitro ADME and preclinical pharmacokinetic studies. We presented our research on the drug metabolism of Emflaza (VBP15) in a national conference: Xu X, et al., In vitro and in vivo metabolism of VBP15 in laboratory animals and human hepatocytes. AAPS Annual Meeting, San Diego, CA, Nov 2-6, 2014.
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