Comparative biophysical and functional analysis of TCZ-UFRJ, a potential biosimilar to Actemra

Biosimilar antibodies have become increasingly significant in the pharmaceutical market, driven by the expiration of patents on many reference products. This study presents a comparative analysis between the originator tocilizumab, used to treat inflammatory diseases such as rheumatoid arthritis, and the biosimilar candidate TCZ-UFRJ, produced in HEK293 cells. The investigation focused on various aspects, including primary, secondary, and tertiary structures, intact mass analysis, glycosylation pattern, and functional testing using the IL-6-sensitive THP-1 cell line and ligand binding assay. LC-MS peptide mapping achieved amino acid full sequence coverage for both TCZ-UFRJ and Actemra, confirming the identity of the biosimilar candidate. Both antibodies exhibited a similar secondary structure with a characteristic beta-sheet spectrum, as determined by circular dichroism. The intrinsic tryptophan emission fluorescence profile confirmed a correctly folded tertiary structure for both mAbs. Intact mass analysis demonstrated similar molecular mass and glycoform profiles. Glycosylation analysis revealed similar glycosylation sites and the presence of major N-glycans in both Actemra and TCZ-UFRJ. Dynamic light scattering analysis indicated a monodisperse sample without the presence of oligomers for both. An LSPR ligand binding assay confirmed the interaction between TCZ-UFRJ and the IL-6 receptor, demonstrating specific binding affinity. Subsequent functional testing in the IL-6-sensitive THP-1 cell line validated the biological activity of TCZ-UFRJ, supporting its potential as a biosimilar candidate. These preliminary findings suggest that TCZ-UFRJ holds promise as a biosimilar candidate to Actemra, but further comprehensive studies, including non-clinical and clinical trials, are essential to establish its safety, efficacy, and overall similarity to the originator drug.