The reliability of sample detection is one of the key points in the determination of biological sample concentration. There are many aspects of the evaluation of this parameter, such as the accuracy and precision of the method, the sample stability test, etc., in the process of sample analysis. The reproducibility of the method and the reliability of sample detection were evaluated by reanalysis of the test samples.
In the method validation process, most of the validation samples are obtained by preparing the standard in the biological matrix. These samples have a theoretical concentration value. After the concentration result is measured, the measured result and the theoretical concentration can be compared and calculated. the accuracy of the analysis.
In sample analysis, a standard curve and quality control samples are included in each analysis batch to monitor the accuracy of the analytical method, but the concentration of the sample to be tested is unknown. Although it is possible to estimate the sample concentration through the mode of administration, drug half-life, etc., due to the complex metabolic environment in the body, coupled with individual differences, protein binding, the influence of known and unknown metabolites and other factors, it is impossible to accurately judge the test sample results. accuracy. Therefore, the test sample reanalysis test has become a way to evaluate the accuracy of the real sample measurement.
Test sample reanalysis test, ISR (Incurred Sample Reanalysis), is a test method to prove test reproducibility by repeatedly analyzing test samples. Although the name is Reanalysis (reanalysis), the actual test is the reproducibility of the sample results (Reproducibility).
The FDA Bioanalytical Method Validation-Guidance for Industry requires that all studies that submit NDAs, BLAs, or ANDAs for critical data for product approval or labeling should be subject to ISR. For example, human BE studies for ANDA applications, or pivotal PK, PD and biomarker studies for NDA or BLA applications. For nonclinical safety studies, at least one ISR was performed for each method and species.
It is clear that an ISR should be performed under the following circumstances:
– Toxicokinetic tests, once per species;
– all pivotal bioequivalence studies;
– first-in-human drug trials;
– First-time drug trials in patients;
– First drug trial in patients with hepatic or renal insufficiency.
– For animal experiments, re-analysis of actual samples may only be required in early pivotal experiments, such as experiments designed to dose and measured concentration relationships.
ICH M10 requires that ISR should be performed at least in the following situations:
-• For preclinical studies, ISR is generally required for each species’ primary nonclinical TK study. It is also acceptable to conduct ISR in a PK study but not in a TK study, as long as the corresponding study is pivotal and will be used for regulatory decision-making.
– All pivotal comparative BA/BE studies
– Clinical trials in humans for the first time;
– Pivotal early clinical trials in patients (one per patient population)
– First or pivotal clinical trials in patients with hepatic and/or renal insufficiency.
ISR analysis should follow the following principles:
-Select 10% of the analysis sample size for testing, if the analysis sample size exceeds 1000, the part exceeding 1000 will be calculated as 5%.
– It is recommended to select samples near Cmax and during elimination, including multiple time points.
– Do not mix test samples to avoid abnormal results.
– Re-analyses should be performed in batches on different dates than the initial analysis.
Acceptance Criteria for ISR:
For at least 2/3 of the ISR samples, the difference between the concentrations measured in the primary and re-analyses should be within ±30% of the mean of both.
In order to obtain good ISR results, the following aspects can be noted:
– Perform a reanalysis using the same batch of standard curve and QC samples as the initial analysis.
– Ensure that the re-analyzed samples are within the validated stability range.
-Pick samples for re-analysis that were diluted between 3x LLOQ and 80% ULOQ at the time of initial analysis.
– Perform a reanalysis using the same dilution as the initial analysis.
– For longer duration clinical projects, ISR is recommended in time periods.
If the ISR results fail to meet the acceptance criteria, the cause of the failure needs to be investigated, otherwise the reliability of the overall project testing will be called into question. The investigation can start from the analysis batch, calculation, sample uniformity, stability and so on. After the cause is identified, further tests are taken to test.
Note: ISR is a test performed to evaluate method reproducibility, and reanalysis results are not reported as final sample concentrations.
Bioanalytical Service – PK Studies