LAUSR

Higher-order structural (HOS) biosimilarity assessment is a regulatory requirement for intended biosimilars, and manufacturers are required to demonstrate the biosimilarity of their product in comparison to the reference product concerning both safety and efficacy. NMR has recently emerged as a powerful technique for complex biologics such as mAbs that offers holistic Higher-order structure (HOS) assessment. In this paper, a proof-of-concept for similarity assessment for marketed biosimilars of trastuzumab has been presented using both 1Dimensional (1H NMR) and 2Dimensional 1H-13C-methyl correlated NMR techniques. Samples were prepared without sample buffer exchange in the presence of excipients and assessed at natural abundance. Data were recorded using a 750 MHz NMR spectrometer equipped with a room temperature probe. Both visual and statistical assessment was performed to test biosimilarity. 1H- NMR was useful for the assessment of overall comparability with sensitivity towards changes in the formulation components, whereas 13C- 1H methyl correlation-based assessment indicated structural biosimilarity. All marketed biosimilars were found to be within the range established by originator batches used in this study. Using 1D and 2D NMR techniques, we aim to provide a perspective on the requirements and challenges of HOS biosimilarity assessments to be conducted for the drug product as a whole, inclusive of excipients as opposed to drug substance alone in buffer exchanged conditions.