Objective: To optimize the tumor cell killing activity assay method of recombinant human GM-CSF herpes simplex virus injection (OrienX010) against michigan cancer foundation-7 cells (MCF-7). Methods: Different concentrations of reactive oxygen species (ROS) inhibitor/ROS scavenger were added at different times to determine the condition that had the least influence on the experiment and can effectively remove ROS. The precision and applicability of the optimized method were verified. Tumor cell killing activity of the same batch of OrienX010 samples was tested 3 times by the assay methods before and after optimization, to determine whether there was a statistic difference before and after optimization. Different generations of MCF-7 cell were tested to determine the cell generation that could be used. Results: About 30 min before adding CCK-8 reagent, 10 μL of 50 times diluted ROS inhibitor/ROS scavenger was added to each well as the best condition. The tumor cell killing activity (MOI ED50) of OrienX010 was tested 6 times on the same day by the same analyst, and the results showed that RSD was 15.1%, meeting acceptable standard. On this basis, the same experiment was tested by another analyst on another day, and the results showed that the RSD of 2 analysts was 20.5%, meeting acceptable standard. Multiple batches of OrienX010 were tested and the results met quality standard. Tumor cell killing activity of the same batch number OrienX010 samples was tested three times by the assay methods before and after optimization. t Stat>t two-tailed critical value, indicating significant differences in the mean values of the two groups of data before and after optimization. MCF-7 cell was tested to P 39, and the results were qualified and stable. Conclusion: The optimized method is effective to eliminate the effect of ROS on tumor cell killing activity detection, and the method has good precision and applicability. The optimized method can be used for the tumor cell killing activity detection of recombinant human GM-CSF herpes simplex virus injection (OrienX010). There is statistic difference before and after optimization. MCF-7 cell can be used up to P 39.
SUN Dong-mei, WANG Yan-hui, WANG Li-qian, ZHAO Feng-ying, MA Yan-qiu, WANG Ning
. Optimization of the assay method for tumor cell killing activity of recombinant human GM-CSF herpes simplex virus injection (OrienX010)[J]. Chinese Journal of Pharmaceutical Analysis, 2022
, 42(12)
: 2101
-2109
.
DOI: 10.16155/j.0254-1793.2022.12.05
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