PD-1 Receptor Occupancy Assay for Mass Cytometry

Pembrolizumab, a therapeutic antibody targeting PD-1, has shown great potential in treating various cancers, even giving durable responses for some patients. However, only a fraction of patients respond to this treatment. Today, the expression of PD-L1 on tumor cells is the most frequently used biomarker to predict response but it is still insufficient to use alone for most cancers. We hypothesized that the occupation by pembrolizumab on PD-1 could identify responders and non-responders. The single-cell analysis technology mass cytometry would enable investigation of occupation in complex cell types as it provides high sensitivity and detection of over 40 parameters. Therefore, we aimed to develop a PD-1 receptor occupancy assay for mass cytometry. A mass cytometry panel of antibodies was developed, containing 173Yb-anti-IgG4 [HP6025] to detect bound pembrolizumab and 166Er-anti-PD-1 [EH12.1] to detect available receptors not bound by pembrolizumab. The panel was tested on samples from 10 patients treated with pembrolizumab. Preliminary statistical analysis was conducted to investigate if there was a significant difference in receptor occupancy of responders and non-responders. The assay measured increasing PD-1 receptor occupancy by pembrolizumab in almost all patients. The results from the preliminary statistical analysis did not show any significant difference in the receptor occupancy of responders and non-responders, but more patients are required to assess this hypothesis. Nevertheless, as this assay enables the investigation of multiple cell populations simultaneously, it could still potentially contribute to predict response of pembrolizumab treated patients.