TOOLBOX: Strep‑Tagged nano‑assemblies of antibody‑drug‑conjugates (ADC) for modular and conditional cancer drugging

Herein, we describe TOOLBOX, a 3‑step modular nano‑assembly targeting system that permits the combinatorial exchange of antibody specificities and toxic payloads, introducing modularity in antibody‑drug conjugate (ADC) manufacturing. TOOLBOX integrates 3 building blocks: i) a recombinant antibody fragment (that in the selected setting targets the proto‑oncogene ERBB2) genetically fused to an 8 amino acid Strep‑Tag^®; ii) a multivalent protein adapter, called Strep‑Tactin^®; iii) two anticancer agents, e.g. DNA nanobinders and the maytansinoid DM1, both carrying a chemically attached Strep‑Tag that reversibly turns them into inactive prodrugs. Stoichiometrically optimized complexes of Strep‑Tagged antibody fragments and drugs, bridged by Strep‑Tactin, were specifically uptaken by breast cancer cells expressing ERBB2, and this unexpectedly resulted in conditional prodrug reactivation. A promoter‑reporter system showed that TOOLBOX inhibited downstream ERBB2 signaling not only in ERBB2‑overexpressing/‑amplified SK‑BR‑3 cells grown in vitro, but also in ERBB2‑low/non‑amplified BRC230 triple‑negative breast carcinoma cells xenotransplanted in nude mice. Thus, TOOLBOX is a modular ADC‑like nano‑assembly platform for precision oncology.