Early Hit-to-Lead ADME screening bundle

In vitro ADME screening during the lead optimization stage of drug discovery positively impacts drug candidate selection with an enhanced probability of success in clinical trials. Since most new drug candidates fail during preclinical and clinical development, and the late stage of the drug development cycle can be a lengthy and costly process, any means of identifying drug candidates with optimized ADME and pharmacokinetics properties in the discovery stage will have a significant impact on the drug discovery process overall.

Focused on Solutions to Address Drug Metabolism and Pharmacokinetics (DMPK) Issues and to Enable the Success of Our Clients

Our scientists provide a range of DMPK services, routinely conducting industry standard in vitro metabolism and DDI-based assays, including highly automated ADME in vitro screens. We can help drive your discovery phase structure activity relationship (SAR) by optimizing for ADME properties, in parallel to your receptor binding potency and selectivity, for more rapid identification of high quality drug candidates.

Metabolic stability, risk assessment for inhibiting key Cytochrome P450 enzymes, and cell permeability are three main early hit-to-lead ADME screening assays that all new chemical entities (NCEs) are tested for in the industry in effort to optimize key ADME properties.

In Vitro ADME Screening Services: Early Hit-to-Lead ADME Screening Bundle

Intrinsic Clearance Assay in Liver Microsomes
Liver microsomes; species selectable Typical turnaround time is ≤10 business days Test article prepared at 10mM in DMSO Substrate and positive control incubated at 0.3µM Incubations performed in HLMs at a protein concentration of 0.25mg/mL	Incubation performed using 1mM co-factor (NADPH) Incubation time points (+NADPH): 0, 5, 15, 30, 45 minutes 45 min negative control (-NADPH) for recovery assessment Samples analysis and quantitation by LC-MS/MS Deliverables: Unscaled intrinsic clearance, recovery Results provided via electronic spreadsheet format

CYP Single Concentration Cocktail Assay (CYP 2C9, 2D6, 3A4)
Human Liver Microsomes (HLM) Typical turnaround time is ≤10 business days Inhibitors prepared at 10mM in DMSO CYP3A4 – Midazolam (5µM), CYP2D6 – Bufuralol (10µM), CYP2C9 – Diclofenac (10µM); substrates incubated as a cocktail Test article and positive control inhibitor incubated at 10µM Incubations performed in HLMs at a protein concentration of 0.05mg/mL Incubation performed using 1mM co-factor (NADPH)	Incubation time: 3 minutes LC/MS-MS analysis using cocktail of heavy labeled internal standard for each metabolite A decrease in the formation of the metabolites compared to vehicle control is used to calculate a percent inhibition value Samples analysis and quantitation by LC/MS-MS Deliverables: Percent inhibition at 10µM Results provided via electronic spreadsheet format

CYP Single Concentration Cocktail Assay (CYP 2C9, 2D6, 3A4)

Human Liver Microsomes (HLM)
Typical turnaround time is ≤10 business days
Inhibitors prepared at 10mM in DMSO
CYP3A4 – Midazolam (5µM), CYP2D6 – Bufuralol (10µM), CYP2C9 – Diclofenac (10µM); substrates incubated as a cocktail
Test article and positive control inhibitor incubated at 10µM
Incubations performed in HLMs at a protein concentration of 0.05mg/mL
Incubation performed using 1mM co-factor (NADPH)

Incubation time: 3 minutes
LC/MS-MS analysis using cocktail of heavy labeled internal standard for each metabolite
A decrease in the formation of the metabolites compared to vehicle control is used to calculate a percent inhibition value
Samples analysis and quantitation by LC/MS-MS
Deliverables: Percent inhibition at 10µM
Results provided via electronic spreadsheet format

MDCK II Bi-directional Permeability Assay
MDCK II cell line (Madin-Darby Canine Kidney cells) – Sigma Aldrich (ECACC) Typical turnaround time is ≤15 business days Test article prepared at 10mM in DMSO Substrate and positive control incubated at 1µM in duplicate Incubations performed in both A:B and B:A in the presence of Cyclosporin A (P-gp inhibitor) Incubations carried out in cell culture incubator at 37°C/5%CO²/95% RH	Samples taken from the A:B plate and B:A plate following a 3 hour incubation A 9-pt concentration curve is prepared for quantitation of samples (2000, 1000, 500, 250, 125, 62.5, 31.3, 15.6, 7.81nM) Dextran Texas Red (DTR) used as monolayer integrity check Sample analysis and quantitation by LC/MS-MS Deliverables: Papp in both A:B and B:A direction, BA:AB ratio, mass balance (using both pre and post dose solutions), percent cell leakage (% DTR) Results provided via electronic spreadsheet format

MDCK II Bi-directional Permeability Assay

MDCK II cell line (Madin-Darby Canine Kidney cells) – Sigma Aldrich (ECACC)
Typical turnaround time is ≤15 business days
Test article prepared at 10mM in DMSO
Substrate and positive control incubated at 1µM in duplicate
Incubations performed in both A:B and B:A in the presence of Cyclosporin A (P-gp inhibitor)
Incubations carried out in cell culture incubator at 37°C/5%CO²/95% RH

Samples taken from the A:B plate and B:A plate following a 3 hour incubation
A 9-pt concentration curve is prepared for quantitation of samples (2000, 1000, 500, 250, 125, 62.5, 31.3, 15.6, 7.81nM)
Dextran Texas Red (DTR) used as monolayer integrity check
Sample analysis and quantitation by LC/MS-MS
Deliverables: Papp in both A:B and B:A direction, BA:AB ratio, mass balance (using both pre and post dose solutions), percent cell leakage (% DTR)
Results provided via electronic spreadsheet format

Liquid chromatography–mass spectrometry (LC-MS) and ImmunoAffinity-LC-MS services

For the quantitative determination of small molecule, peptide and macromolecule therapeutics in support of pharmacokinetic (PK) studies.

In vitro ADME assays and metabolite identification services

including permeability, metabolic stability, metabolite identification (cold & radiolabeled), and in vitro drug-drug interaction risk assessment, ranging from highly automated discovery screening platforms to definitive development studies that enable regulatory submission.

Immunoassay services

Including quantitative determination of large molecule therapeutics using ligand binding assays technologies in support of pharmacokinetic (PK) studies and immunogenicity assays assessments for pre-clinical and clinical studies including tier-based approaches, neutralizing assays, and isotyping assays.

Fit-for-purpose biomarker services

Including quantitative determination of large molecule therapeutics using ligand binding assays technologies in support of pharmacokinetic (PK) studies and immunogenicity assays assessments for pre-clinical and clinical studies including tier based approaches, neutralizing assays, and isotyping assays.