A real‐time cell‐binding assay reveals dynamic features of STxB–Gb3 cointernalization and STxB‐mediated cargo delivery into cancer cells

João Crispim Encarnação (1, 2) , Valeria Napolitano (3) , Giulia Opassi (4) , U. Helena Danielson (4) , Grzegorz Dubin (5) , Grzegorz Popowicz (6, 7) , Hélène Munier-Lehmann (8) , Jos Buijs (1, 2) , Karl Andersson (1, 2) , Hanna Björkelund (1)

1 Ridgeview Instruments AB [Uppsala, Sueden]
2 IGP - Department of Immunology, Genetics and Pathology [Uppsala, Sueden]
3 UJ - Uniwersytet Jagielloński w Krakowie = Jagiellonian University
4 Department of Chemistry [Uppsala, Sueden]
5 MCB - Malopolska Centre of Biotechnology [Krakow]
6 Helmholtz Zentrum München = German Research Center for Environmental Health
7 CIPSM - Center for Integrated Protein Science
8 Chimie et Biocatalyse

The interaction between the Shiga toxin B-subunit (STxB) and its globotriaosylceramide receptor (Gb3) has a high potential for being exploited for targeted cancer therapy. The primary goal of this study was to evaluate the capacity of STxB to carry small molecules and proteins as cargo into cells. For this purpose, an assay was designed to provide real-time information about the StxB-Gb3 interaction as well as the dynamics and mechanism of the internalization process. The assay revealed the ability to distinguish the process of binding to the cell surface from internalization and presented the importance of receptor and STxB clustering for internalization. The overall setup demonstrated that the binding mechanism is complex, and the concept of affinity is difficult to apply. Hence, time-resolved methods, providing detailed information about the interaction of STxB with cells, are critical for the optimization of intracellular delivery.

Mots clés: cancer; cell surface receptor; real-time cell-binding assays; receptor internalization; Shiga toxin; binding kinetics

Domaines: Sciences du Vivant [q-bio]; Sciences de l'ingénieur [physics]

João Crispim Encarnação, Valeria Napolitano, Giulia Opassi, U. Helena Danielson, Grzegorz Dubin, et al.. A real‐time cell‐binding assay reveals dynamic features of STxB–Gb3 cointernalization and STxB‐mediated cargo delivery into cancer cells. FEBS Letters, 2020, 594 (15), pp.2406-2420. ⟨10.1002/1873-3468.13847⟩. ⟨pasteur-02892876⟩