Combining protein proximity detection with chromogenic multiplex: Discovering the PD1/PDL1 interactions

DISCOVERY Proximity detection is a powerful technique for demonstrating the presence of 2 unique targets in direct proximity of each other in tissue sections. It allows visualization of interacting proteins in distance of 40 nm. Recently this technique became available on the Roche DISCOVERY ULTRA staining platform. With the DISCOVERY Proximity detection you can not only detect the presence/absence of a protein, but also to determine its functional states; i.e., whether it carries post-translational modifications or is present in complex with other proteins, in which cells it is expressed, and in which cell compartments the different states of the protein are present. By having the possibility on the DISCOVERY ULTRA system to add chromogenic IHC multiplexing you can create the advantage of visualizing which cells are undergoing protein interaction. In our recent publication we investigated the interaction between PD1/PDL1 where we studied tumour draining lymph nodes of melanomas and the primary tumours with proximity and chromogenic IHC multiplexing. Drugs targeting the PD1/PDL1 pathway have revolutionized the treatment of multiple cancer types including non-small cell lung cancer, renal cancer, and melanoma with a subset of patients experiencing durable responses. PD1/PDL1 blocking antibodies are believed to act primarily in the tumour microenvironment, by reinvigorating exhausted T cells and thereby reviving pre-existing anti-tumour immunity. In this presentation, we will review this technology by setting up PD1/PDL1 Proximity assay in combination with chromogenic IHC multiplexing with CD8, CD68 and CD11c.

Learning Objectives:

1. Obtain knowledge on how to set up the Proximity detection on Roche DISCOVERY ULTRA Platform

2. Obtain knowledge on how to add chromogenic IHC multiplexing to Proximity detection

3. Understand PD-1/PD-L1 interactions in Tumor Draining Lymph Nodes (TDLN) but not tumor correlate with prognosis in melanoma