Modelling ligand depletion for simultaneous affinity and binding site quantification on cells and tissue.

The quantification of the number of targets in biological systems is an important parameter to assess the suitability of surface markers as targets for drugs, drug delivery and medical imaging. Likewise, quantifying the interaction with the target in terms of affinity and binding kinetics is essential during drug development. Commonly used approaches to quantify membrane antigens on live cells are based on manual saturation techniques that are labour-intensive, require careful calibration of the generated signal and do not quantify the binding rates. Here, we present how measuring interactions in real-time on live cells and tissue under ligand depletion conditions can be used to simultaneously quantify the kinetic binding parameters as well as the number of available binding sites in a biological system. Suitable assay design was explored with simulated data and feasibility of the method verified with experimental data for exemplary low molecular weight peptide and antibody radiotracers as well as fluorescent antibodies. In addition to revealing the number of accessible target sites and improving the accuracy of binding kinetics and affinities, the presented method does not require knowledge about the absolute signal generated per ligand molecule. This enables a simplified workflow for use with both radioligands and fluorescent binders.

Practical considerations for navigating the regulatory landscape of non-clinical studies for clinical translation of radiopharmaceuticals.

BACKGROUND: The development of radiopharmaceuticals requires extensive evaluation before they can be applied in a diagnostic or therapeutic setting in Nuclear Medicine. Chemical, radiochemical, and pharmaceutical parameters must be established and verified to ensure the quality of these novel products. MAIN BODY: To provide supportive evidence for the expected human in vivo behaviour, particularly related to safety and efficacy, additional tests, often referred to as "non-clinical" or "preclinical" are mandatory. This document is an outcome of a Technical Meeting of the International Atomic Energy Agency. It summarises the considerations necessary for non-clinical studies to accommodate the regulatory requirements for clinical translation of radiopharmaceuticals. These considerations include non-clinical pharmacology, radiation exposure and effects, toxicological studies, pharmacokinetic modelling, and imaging studies. Additionally, standardisation of different specific clinical applications is discussed. CONCLUSION: This document is intended as a guide for radiopharmaceutical scientists, Nuclear Medicine specialists, and regulatory professionals to bring innovative diagnostic and therapeutic radiopharmaceuticals into the clinical evaluation process in a safe and effective way.