Creative Biolabs has worked extensively on a wide variety of metabolism studies. As part of our in vitro metabolism service, we provide metabolic stability assay service for all small molecule formulations to measure in vitro intrinsic clearance or to estimate metabolic half-life.
The aim of metabolic stability study is measuring the disappearance rate of a chemical compound. The half-life, in vitro clearance measured from metabolic stability assay can be used to predict hepatic clearance, and also to evaluate dose, toxicity, oral bioavailability, etc. For instance, if a drug is rapidly metabolized when entering to the body, it may require more daily dosing to maintain its blood or tissue concentrations in order to get an ideal therapeutic effect. On the other hand, if a drug is slowly metabolized, the dose need to be adjusted and more preclinical toxicity tests should be conducted as long-time exposure may cause toxic build-up.
Figure 1. General scheme of preclinical metabolism studies that are currently being performed to increase the chances of selecting candidate drugs that will be successful in the clinical phase of drug development. CYP = cytochrome P450. (Masimirembwa et al. 2003)
Creative Biolabs provides UPLC/HR-MS/MS technology for liver microsomal, hepatocyte, S9, and recombinant enzymes to assess metabolic stability. Multispecies samples are available for testing interspecies differences. Additionally, we provide microsomal binding assay to determine the free chemicals available in the microsomal incubations and metabolite profiling and identification service to elucidate metabolites formed in the stability studies.
Liver Microsomal Stability
The liver is the main organ for drug metabolism in the body. Microsome, the liver subcellular fraction, is an important model for drug metabolism studies. It contains many drug metabolizing enzymes (like P450s, FMOs, and UGTs), easy to prepare, has long storing period, easy to adaptable to HTS, and can make donor pool from multiple donors to minimize the effect of interindividual variability. Creative Biolabs provides liver microsomal stability assay which incubates microsomes with the test compound at 37°C followed by monitoring disappearance of the test compound. The results are characterized by UPLC/HR-MS/MS.
Hepatocyte Stability
Intact hepatocytes contain the cytochrome P450s (CYPs), non-P450 enzymes, and phase II enzymes such as sulfo- and glucuronosyltransferases. Therefore, hepatocyte assays can more broadly assess the overall cellular metabolism of the test compound than liver microsomal assay. Creative Biolabs provides immortalized human liver cell line HepaRG for hepatocyte stability assays.
S9 Stability
S9 fraction (post-mitochondrial supernatant fraction) is another important model for drug metabolism studies. S9 fraction consists of both microsomal and cytosolic enzymes (SULT, GST, XO, ADHs, NATs) and it can be supplemented with cofactors such as UDPGA and PAPS to investigate Phase II metabolic pathways.
Recombinant Enzymes Stability
Creative Biolabs uses mixture of expressed enzymes (e.g. CYP450s and UGTs) to identify which enzyme metabolizing isoforms are responsible for the metabolism of a test compound. Also, it is useful for identifying potential drug-drug interactions. This system is especially useful for studying highly stable compounds.
Creative Biolabs also provide several in vivo models for metabolic stability assay. For more detailed information, please feel free to contact us or directly sent us an inquiry.
Reference
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