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Biosimilars are biological products that are highly similar to existing products approved by health authorities. Demonstration of similarity starts with the comprehensive analysis of the reference product and its proposed biosimilar at the physicochemical and functional levels. Here, we report the results of a comparative analysis of a proposed biosimilar adalimumab MSB11022 and its reference product, Humira®. Three batches of MSB11022 and up to 23 batches of Humira® were analyzed by a set of state-of-the-art orthogonal methods. Primary and higher order structure analysis included N/C-terminal modifications, molecular weight of heavy and light chains, C-terminal lysine truncation, disulfide bridges, secondary and tertiary structures, and thermal stability. Purity ranged from 98.4%–98.8% for MSB11022 batches (N = 3) and from 98.4%–99.6% for Humira® batches (N = 19). Isoform analysis showed 5 isoform clusters within the pI range of 7.94–9.14 and 100% glycan site occupancy for both MSB11022 and Humira®. Functional analysis included Fab-dependent inhibition of tumor necrosis factor (TNF)-induced cytotoxicity in L929-A9 cell line and affinity to soluble and transmembrane forms of TNF, as well as Fc-dependent binding to Fcγ and neonatal Fc receptors and C1q complement proteins. All tested physicochemical and functional parameters demonstrated high similarity of MSB11022 and Humira®, with lower variability between MSB11022 and Humira® batches compared with variability within individual batches of Humira®. Based on these results, MSB11022 is anticipated to have safety and efficacy comparable to those of Humira®.

We analyzed the protective mechanisms induced against respiratory syncytial virus subgroup A (RSV-A

CXCL12α has been shown to be selectively processed at the N- and C-termini in blood and plasma in vitro. In order to study the processing in vivo, several versions of CXCL12α were expressed and purified. The protein was administered either iv or sc to mice, and at different time points postadministration plasma was collected and analyzed. To detect modifications of the CXCL12α molecule in crude plasma a SELDI TOF-MS-based method was developed. Anti-CXCL12 antibodies were immobilized on the SELDI chip and CXCL12α binding to the antibodies was detected by SELDI-TOF-MS. The protein was found to be processed both at the C- and N-termini. The same processed CXCL12α forms as detected in vitro were found; however, in addition further processing was detected at the N-terminus, where altogether seven amino acids were removed. At the C-terminus the lysine was removed as has been seen in vitro, and no further processing was detected. The full-length CXCL12α disappeared within minutes after administration, whereas the processed forms of the protein were detectable for up to 6-8 h postadministration. The same processed forms appeared after iv and sc administration, only the kinetics was different. When the shortest processed form detected in plasma, 7ΔN1ΔC-CXCL12α, was administered directly, no further processed forms were detected. Interestingly, a version of CXCL12α containing a N-terminal methionine was protected against N-terminal processing in plasma in vitro; however, in vivo no protection was seen, the protein was processed in the same way as full-length CXCL12α.