Lectin microarray technology was applied to compare the glycosylation pattern of the monoclonal antibody MB311 expressed in SP2.0 cells to an antibody-dependent cellular cytotoxic effector function (ADCC)-optimized variant (MB314). MB314 was generated by a plant expression system that uses genetically modified moss protoplasts (Physcomitrella patens) to generate a de-fucosylated version of MB311. In contrast to MB311, no or very low interactions of MB314 with lectins Aspergillus oryzae l-fucose (AOL), Pisum sativum agglutinin (PSA), Lens culinaris agglutinin (LCA), and Aleuria aurantia lectin (AAL) were observed. These lectins are specific for mono-/biantennary N-glycans containing a core fucose residue. Importantly, this fucose indicative lectin-binding pattern correlated with increased MB314 binding to CD16 (FcγRIII; receptor for the constant region of an antibody)—whose affinity is mediated through core fucosylation—and stronger ADCC. In summary, these results demonstrate that lectin microarrays are useful orthogonal methods during antibody development and for characterization.

The detection of KRAS mutations in codons 12 and 13 is critical for anti-EGFR therapy strategies; however, only those methodologies with high sensitivity, specificity, and accuracy as well as the best cost and turnaround balance are suitable for routine daily testing. Here we compared the performance of compact sequencing using the novel hybcell technology with 454 next-generation sequencing (454-NGS), Sanger sequencing, and pyrosequencing, using an evaluation panel of 35 specimens. A total of 32 mutations and 10 wild-type cases were reported using 454-NGS as the reference method. Specificity ranged from 100% for Sanger sequencing to 80% for pyrosequencing. Sanger sequencing and hybcell-based compact sequencing achieved a sensitivity of 96%, whereas pyrosequencing had a sensitivity of 88%. Accuracy was 97% for Sanger sequencing, 85% for pyrosequencing, and 94% for hybcell-based compact sequencing. Quantitative results were obtained for 454-NGS and hybcell-based compact sequencing data, resulting in a significant correlation (r = 0.914). Whereas pyrosequencing and Sanger sequencing were not able to detect multiple mutated cell clones within one tumor specimen, 454-NGS and the hybcell-based compact sequencing detected multiple mutations in two specimens. Our comparison shows that the hybcell-based compact sequencing is a valuable alternative to state-of-the-art methodologies used for detection of clinically relevant point mutations.

The World Health Organization (WHO) defines a similar biotherapeutic product, i.e. a biosimilar as a biotherapeutic product that is similar in terms of quality, safety and efficacy to an already licensed reference product on the market (termed as originator or innovator). To ensure similar efficacy and safety, comparability studies for biosimilars should be carried out at quality, preclinical and clinical level. Glycosylation profiles of biosimilars are getting an additional critical quality attribute. The portfolio required for regulatory comparison of identity and release testing has to be assessed with a large number of analytical tests, which are presented here in detail for the example of adalimumab. Adalimumab, as one of the blockbuster drugs and the best-selling pharmaceutical product world-wide, is analyzed additionally with a competitive field presenting the marketed products or such in development. This competitive field is not surprisingly very crowded. Furthermore, general information about the currently marketed biosimilars including Mode of Action (MoA), clinical indication(s), patent situation, and market situation are shortly summarized for each of the respective originator / biosimilar molecules. As there are not only markets in developed and highly regulated countries, but also in other regions, examples for companies and products in Asia, South and Middle America, and even in Africa are highlighted as well.