Development and multicenter validation of an LC–MS-based bioanalytical method for antisense therapeutics: supplementary figures

Background: Many bioanalytical methods for antisense oligonucleotides (ASOs) using LC–MS have been

reported. However, no data have been available on the reproducibility and robustness of a single

bioanalytical method for ASOs. As such, in the current study, we evaluated the reproducibility and

robustness of LC–MS-based bioanalytical methods for ASOs in multiple laboratories. Methods/Results:

Seven independent laboratories were included in this study.Mipomersen was measured by ion-pairing LC–

MS (IP-LC–MS) as a model ASO using different LC–MS. The validation results of calibration curve, accuracy,

precision and selectivity met the criteria of conventional bioanalytical method validation guidelines using

LC/GC–MS for drugs in all laboratories. Meanwhile, carryover (>20%) was detected in three laboratories.

Conclusion: We first demonstrated the multicenter-validated IP-LC–MS bioanalytical method for ASOs.

Our data showed that the method was sensitive, robust and reproducible. However, the occurrence of

carryover should be carefully monitored in its future application.