LAUSR

DETERMINATION of the optimal therapeutic dose is a key challenge in drug development. Receptor occupancy assays (RO assays) measure the binding of biotherapeutics to their cellular targets and are one of the tools used to determine starting doses in clinical and preclinical studies. Flow cytometry-based RO assays have been developed for heterogeneous cell populations in suspension and are well suited for evaluating therapeutics targeting peripheral blood cells. Mass cytometry would further expand the possibilities to measure receptor occupancy of complex cell types as metallabeled antibodies and minimal overlap between channels allow for more than 40 markers to be detected simultaneously without loss of sensitivity. Bringeland et al. (1) have developed a mass cytometry-based RO assay, recently described in a technical note. With a backbone of 15 lineage markers, they identified eight cell types of interest and obtained receptor occupancy data for each cell population. Receptor occupancy assays can prevent life-threatening side effects in clinical trials, exemplified by the disastrous TGN1412 first-in-human clinical trial in 2006 (2). TGN1412 is an anti-CD28 superagonistic monoclonal antibody that stimulates T cells, intended for treatment of B-cell chronic lymphocytic leukemia and rheumatoid arthritis. In preclinical trials, cynomolgus monkeys were used to determine toxicity, and starting doses in the first human clinical trial was calculated based on the no-adverse-effect level (NOAEL) in these animals (3). However, despite using doses 500 times lower than the dose found safe in animal models, the human volunteers involved in the trial suffered severe cytokine release syndrome and multiple organ failure after receiving the drug (2). Follow-up studies revealed that biological differences between humans and monkeys, in this case different expression levels of CD28 on their respective T cells, resulted in 90% receptor occupancy and severe cytokine release syndrome with a dose that was expected to lead to only 10% receptor occupancy (4). A simple RO assay could have revealed these differences, and as a consequence of this trial, the procedures for first-in-human clinical trials were changed (5). Today, RO assays play an increasing role in drug development (3,4) and have been used in testing of therapeutic antibodies like anti-PD-1 (6) and anti-PD-L1 (7). Receptor occupancy data can contribute to determining the minimal anticipated biological effect level (MABEL) which is often better suited than NOAEL to predict safe starting doses in first-in-human clinical trials. In addition to revealing safety hazards, RO assays can limit the extent of preclinical testing, which could reduce costs for the pharmaceutical industry and benefit patients by introducing potentially life-saving drugs earlier. During later-phase trials, receptor occupancy data can monitor therapeutic response and guide optimal dosing for individual patients. Bringeland et al. (1) and others (8) have investigated the use of RO assays for dosing of natalizumab, an effective therapy for multiple sclerosis patients. Multiple sclerosis is a chronic inflammatory autoimmune