Understand the drug metabolism of your compound by identifying which metabolites are formed during in vitro or in vivo studies.
Metabolite profiling and identification is included in Cyprotex's portfolio of ADME services. Cyprotex deliver consistent, high quality data in line with regulatory guidelines, and can adapt protocols based on specific customer requirements.
We encourage the identification of differences in drug metabolism between animals used in nonclinical safety assessments and humans as early as possible during the drug development process. The discovery of disproportionate drug metabolites late in drug development can potentially cause development and marketing delays.
1FDA Guidance for Industry: Safety Testing of Drug Metabolites (February 2008)
Matrices Analyzed | Typically, microsomal incubations, hepatocyte incubations, expressed enzyme incubations, and plasma (others available on request) |
Instruments | AB Sciex TripleTOF® 6600 Waters Xevo® G2-S QTof LabLogic Beta-Ram® (for radiolabelled studies) |
Data Delivery | Various options are available depending on the depth and breadth of metabolite profiling or identification that is required. Typically data is delivered as either:
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A summary of all potential metabolites is provided.
Name | Formula | Mass Difference | m/z Found | Mass Error (ppm) | Identifier | Rt (min) |
---|---|---|---|---|---|---|
Parent | C18H25NO | - | 272.02010 | -1.5 | - | 8.33 |
Dehydration | -H2O | -18.0118 | 254.1896 | -4.9 | M5 | 9.09 |
Demethylation | -CH2 | -14.0159 | 258.1855 | -1.0 | M3 | 8.28 |
Demethylation | -CH2 | -14.0154 | 258.1860 | 0.9 | M1 | 5.72 |
Oxidation | +O | +15.9935 | 288.1949 | -1.4 | M4 | 8.85 |
Oxidation | +O | +15.9946 | 288.1930 | -0.3 | M2 | 6.67 |
Why is understanding the routes of metabolism of a compound important?
There are several reasons why understanding the metabolic profile of a compound is important. Firstly, the FDA Guidance for Safety Testing of Drug Metabolites (2008) recommends in vitro evaluation of interspecies differences in drug metabolism between humans and those animals which are expected to be used in preclinical safety assessments. If a metabolite is formed only in humans and is absent in the animal test species, or if the metabolite is present at disproportionately higher levels in humans than in the animal species, then it may be necessary to assess the preclinical safety of the metabolite in question. Secondly, understanding the metabolic liability of a compound is important in directing chemistry. If a compound is very rapidly cleared, then identifying which functional groups are undergoing metabolism will be valuable in understanding how the chemistry may be altered to reduce compound degradation.
What do you recommend as an appropriate strategy for evaluation of the routes of metabolism of a compound?
Before embarking on an in vitro metabolite profiling study, it is recommended that the compound has previously been investigated in the in vitro drug metabolizing system of choice. This will give an indication of the extent of metabolism and whether sensitivity by mass spectrometry is likely to be a problem.
The decision to progress to metabolite profiling and/or identification, and the level of interpretation which is required is driven by the goal of the study. For example, screening multiple compounds through microsomal clearance may only require the identification of the major metabolites, in order to localize any metabolic hot-spots that can then be designed out. A single compound that is screened through multiple hepatocyte species is likely to be require a more complete interpretation if it is to aid in the decision point for pre-clinical toxicology models (minimize the chances of human specific metabolites). Cyprotex are able to offer services ranging from data acquisition through to full structural elucidation.
What are the advantages of high resolution accurate mass spectrometry instruments for metabolite profiling studies?
By utilizing high resolution, accurate mass spectrometry, specificity is improved without compromising on sensitivity. Greater confidence in structural elucidation can occur as both MS precursor and 'MS/MS' products ions can only be rationalized with a very small number of elemental combinations – therefore increasing the confidence in structural assignment.1FDA Guidance for Industry: Safety Testing of Drug Metabolites (February 2008)
Learn more about drug metabolism in Chapter 3 of our popular Everything you need to know about ADME guide.
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