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When Analytical Characterization of a Biosimilar Reveals Shifts in Quality Attributes of a Reference Product


The attributes for the N-glycan profile, FcγRIIIa binding activity, and relative antibody-dependent cell-mediated cytotoxicity (ADCC) activity for the reference trastuzumab resulted in reduced ADCC and FcγRIIIa binding activities.

Biosimilar approval relies on a demonstration of the totality of the evidence, including analytical, nonclinical, and clinical similarity of the proposed product and its reference. Analytical similarity is crucial to understanding whether there is the potential for any differences in clinical outcomes between a biosimilar and its reference, and the assessment must be based on analytical data relevant to the reference drug’s critical quality attributes.

As described in a newly published paper, when drug maker Samsung Bioepis was developing its biosimilar trastuzumab, later approved as Ontruzant, the developers analyzed 154 lots of EU-licensed and US-licensed reference trastuzumab and found 2 marked changes in critical quality attributes in the reference Herceptin.1

According to the authors, the attributes for the N-glycan profile, FcγRIIIa binding activity, and relative antibody-dependent cell-mediated cytotoxicity (ADCC) activity showed 2 shifts during monitoring; first, galactose and afucose were decreased and second, the level of high mannose was “remarkably” increased.

Since N-glycan levels correspond to ADCC activity (the mechanism by which trastuzumab attracts immune cells to HER2-overexpressing tumor sites), as well as to FcγRIIIa binding activity (which is linked with effector binding in the Fc region during ADCC), the changes resulted in reduced ADCC and FcγRIIIa binding activities in the reference trastuzumab.

The authors write that, in newer lots of the reference trastuzumab, which have expiry dates after February 2020, the biological profile of the reference product recovered to one that is similar to that of lots previous to these observed quality shifts, suggesting that the changes in biological profile—which have the potential to impact the clinical efficacy of the product—were abnormal events.

In order to establish the target quality ranges for the biosimilar, the developers of Ontruzant had to exclude the affected lots of the reference product from their analysis. According to the authors, the cause of the now-resolved shift in the reference trastuzumab remains unknown.

“The abrupt changes in biological profile also occurred in other marketed products, Rituxan and Enbrel, which showed alterations of N-glycan and ADCC activity as well as the reference product,” add the authors. Such changes in the rituximab and etanercept products may have resulted from manufacturing process changes, and their influence cannot be confirmed without additional clinical studies, they write.

In the case of trastuzumab, however, Samsung Bioepis recently presented data that showed that, among patients with HER2-positive early or locally advanced breast cancer in the neoadjuvant setting who were treated with Ontruzant and patients treated with the reference Herceptin who had not been exposed to the lots with lower ADCC activity, there was no difference in observed event-free survival (EFS) or overall survival (OS) at 3 years of follow-up.2

However, among patients who were treated with the reference Herceptin, exposure was associated with decreased EFS compared with no exposure. There was also a trend of decreased OS, though the difference was not statistically significant.

These findings raise the important questions of whether these 2 shifts in critical quality attributes in the reference product, which had an impact on patient survival, would have come to light had the biosimilar developer not sampled the affected lots in the biosimilar development process, and whether the cause of the transient but marked changes will be made known. Furthermore, they raise the question of whether shifts in attributes for reference rituximab and etanercept may also have a clinical impact.


1. Lee JH, Paek K, Moon JH, Ham S, Song J, Kim S. Biological characterization of SB3, a trastuzumab biosimilar, and the influence of changes in reference product characteristics on the similarity assessment [published online June 12, 2019]. BioDrugs. doi: 0.1007/s40259-019-00362-5.

2. Pivot X, Pegram MD, Cortes J, et al. Evaluation of survival by ADCC status: Subgroup analysis of SB3 (Trastuzumab Biosimilar) and reference trastuzumab in patients with HER2-positive early breast cancer at three-year follow-up. Presented at: the American Society of Clinical Oncology Annual Meeting 2019; May 31-June 4, 2019; Chicago, Illinois. Abstract 580.

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