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International audience; The epidermal growth factor receptor (EGFR) is a cell-surface receptor with a single transmembrane domain and tyrosine kinase activity carried by the intracellular domain. This receptor is one of the four members of the ErbB family including ErbB2, ErbB3, and ErbB4. Ligand binding, like EGF binding, induces a conformational rearrangement of the receptor and induces a homo/hetero dimerization essentially with ErbB family receptors that leads to the phosphorylation of the kinase domain, triggering a signaling cascade. EGFR can also form inactive dimers in a ligand-independent way through interactions between cytoplasmic domains. To date, the conformation of EGFR extracellular domain engaged in these inactive dimers remains unclear. In this study, we describe the successful selection and characterization of llama anti-EGFR nanobodies and their use as innovative conformational sensors. We isolated three different specific anti-EGFR clones binding to three distinct epitopes. Interestingly, the binding of all three nanobodies was found highly sensitive to ligand stimulation. Two nanobodies, D10 and E10, can only bind the ligand-free EGFR conformation characterized by an intramolecular tether between domains II and IV, whereas nanobody G10 binds both ligand-free and ligand activated EGFR, with an 8-fold higher affinity for the extended conformation in the presence of ligand. Here we took advantage of these conformational probes to reveal the existence of tethered EGFR in EGFR/ErbB2 predimers. These biosensors represent important tools allowing the determination of EGFR conformations and should help the design of relevant inhibitors.

International audience; Despite the widespread availability of immunohistochemical and other methodologies for screening and early detection of lung and breast cancer biomarkers, diagnosis of the early stage of cancers can be difficult and prone to error. The identification and validation of early biomarkers specific to lung and breast cancers, which would permit the development of more sensitive methods for detection of early disease onset, is urgently needed. In this paper, ultra-small and bright nanoprobes based on quantum dots (QDs) conjugated to single domain anti-HER2 (human epidermal growth factor receptor 2) antibodies (sdAbs) were applied for immunolabeling of breast and lung cancer cell lines, and their performance was compared to that of anti-HER2 monoclonal antibodies conjugated to conventional organic dyes Alexa Fluor 488 and Alexa Fluor 568. The sdAbs–QD conjugates achieved superior staining in a panel of lung cancer cell lines with differential HER2 expression. This shows their outstanding potential for the development of more sensitive assays for early detection of cancer biomarkers.