ORIGINAL RESEARCH article
Front. Plant Sci.
Sec. Plant Proteomics and Protein Structural Biology
Volume 16 - 2025 | doi: 10.3389/fpls.2025.1607403
This article is part of the Research TopicPlant Glycobiology - A Sweet World of Glycans, Glycoproteins, Glycolipids, and Carbohydrate-Binding Proteins, Volume IIView all articles
Plant-related Quality Attributes Affecting FcγRIIIa Binding: Affinity Chromatography Analysis of Rituximab Glycovariants from Nicotiana benthamiana
Provisionally accepted
Caterina Temporini1*
Sara Tengattini1
Aurora Tini1
Carla Marusic2
Francesca Rinaldi1
Isabella Senini1
Enrica Calleri1Virginia Perez3Claudio Pisano3
Marcello Donini2*- 1University of Pavia, Pavia, Italy
- 2Italian National Agency for New Technologies, Energy and Sustainable Economic Development (ENEA), Rome, Lazio, Italy
- 3Special Product's Line, Anagni, Italy
Select one of your emails
You have multiple emails registered with Frontiers:
Notify me on publication
Please enter your email address:
If you already have an account, please login
You don't have a Frontiers account ? You can register here
Monoclonal antibodies (mAb) produced in plants, known as plantibodies, represent a cost-effective alternative to conventional mammalian cell cultures. Glycoengineering processes are needed to alter the N-glycosylation avoiding plant-typical glycans and enabling, for anti-cancer mAbs, the production of biobetters with improved antibody-dependent cell-mediated cytotoxicity (ADCC). In this study, glycovariants of the mouse/human chimeric anti-CD20 antibody Rituximab were produced in Nicotiana benthamiana plants by transient expression using vacuum-agroinfiltration technology and LED lighting. To modify the glycosylation profile the treatment with kifunensine mannosidase I Inhibitor (K) was used, as well as ΔXF N. benthamiana plants optimized by 'genome editing'. The produced plantibodies were characterized to assess their structural properties, including primary sequence and glycosylation profile. Binding to the FcγRIIIa receptor was investigated by affinity chromatography to explore plantibody ADCC. The influence of the glycosylation on FcγRIIIa receptor affinity was evaluated, as well as the impact of post-translational modifications (PTMs). Both glycoengineering strategies showed to produce mAbs with comparable, or improved, affinity for FcγRIIIa receptor. For the first time, different in vivo glycoengineering approaches have been compared through the characterization of the resulting mAbs and their affinity for FcγRIIIa receptor. This insight into the correlation among expression system, plantibody glycoprofile, and predicted ADCC of individual glycoforms has not been previously reported and provides valuable support for the development of plant-based biosimilars.
Keywords: Plantibodies, monoclonal antibodies, rituximab, Glycosylation, Glycoengineering, FCγRIIIa receptor, Antibody-Dependent Cell Cytotoxicity, affinity chromatography
Received: 07 Apr 2025; Accepted: 13 May 2025.
Copyright: © 2025 Temporini, Tengattini, Tini, Marusic, Rinaldi, Senini, Calleri, Perez, Pisano and Donini. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence:
Caterina Temporini, University of Pavia, Pavia, Italy
Marcello Donini, Italian National Agency for New Technologies, Energy and Sustainable Economic Development (ENEA), Rome, 00196, Lazio, Italy
Disclaimer: All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher.