Project Portugal 2030

Summary

We propose to develop and preclinically validate an anti-HER2 Nb-based-tool that crosses the brain blood barrier (BBB) and can simultaneously diagnose and treat HER2+ BrM. The ultimate goal of this project will be developing a new theranostic agent for early detection and treatment of BrM in BC patients. This project benefits from the high specificity and affinity (low nM range) of Nbs for this particular target, the advances of molecular imaging technologies and the emerging radiopharmaceutical-based therapies tools. This proposal represents a promising strategy to improve the prognosis of HER2+ BC patients with BrM that perfectly fits the sustainable development goal of the 2030 Agenda of Promoting Health and Well-being. To achieve this, we have four main objectives to address, following this logical order: 1.Optimize the process of radiolabeling Nbs with the theranostic pair (61Cu/67Cu) to achieve a formulation safe for human administration without impacting its target affinity. First, the choice for the most suitable chelator is necessary to grant both purity and stability. Following, the establishment of the GMP synthesis process of copper-61 will be carried out and similarly implemented for copper-67 counterpart. Finally, the optimization of the ideal formulation and in vitro affinity studies will be performed to guarantee simultaneously high stability in vivo and the target affinity of the Nbs. 2.Demonstrate the affinity of for HER2-expressing cells and its imaging diagnostic potential for early detection of BrM by PET/MRI; The in vivo affinity studies of Nbs for HER2-expressing cells will be evaluated in already established brain metastatic mouse model using PET/MRI to demonstrate that 61Cu-labeled Nbs are able to early detect of HER2+ BrM. Pharmacokinetic and pharmacodynamic parameters will be also assessed. Additionally, dose-activity curves will be used to estimate the activity dose in each organ and BrM, and to predict the therapeutic dose with 67Cu-Nbs. 3.Demonstrate the therapeutic safety and efficacy of 67Cu-Nbs against HER2+ BrM; The maximum tolerated dose of 67Cu-Nbs will be determined, which corresponds to the dose that does not causes loss of body weight or death of the animals. Different therapeutic doses will be tested in the brain metastatic model to assess the inhibitory effect on metastasis growth and reduction in lesions size as a readout of therapeutic efficacy. Kaplan–Meier plots and log-rank tests will be performed for survival analysis. 4.Confirm the diagnostic potential of radiolabeled Nbs in patient-specific tumor microenvironment. Organotypic slice cultures of metastatic tissues from HER2+ BC patients will be used to confirm the biological affinity and binding properties of 61Cu-Nbs. Through this project, we will develop an innovative, more specific, and non-invasive imaging tool allowing the detection HER2 receptors in BC patients with BrM. This will facilitate the screening of potential patients who may benefit from HER2-targeted therapy. Subsequently, we will also develop a therapeutic radiopharmaceutical using 67Cu to treat the identified patients. This theranostic approach with diagnostic and therapeutic counterparts represents a unique alternative to the current clinical management of these patients. Although preclinical, these results have great translational potential since they will be settled using non-immunogenic synthetic Nbs and radionuclides that are currently used in clinical practice.