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Biomarker multiplexing pathology and panel development

A major focus of the lab is the development and validation of reliable quantitative prognostic and predictive biomarkers.

The lab uses a unique multiplexing technology called multiplex immunofluorescence, developed by GE Global Research Centre, that applies multiple stain-image-bleach cycles on the same tissue slide, thereby allowing for co-localization of up to 100 markers on the same cell.

https://sunnybrook.ca/uploads/1/_research/labs/ge-cytell-multiplexing-platform.jpgGE Cytell Multiplexing Platform
Workflow of multiplexing immunofluorescence (click image for larger view)

Workflow of multiplexing immunofluorescence

Using quantitative methods measuring biomarker signal we are able to define the extent of cancer heterogeneity by characterizing each clone, and investigate potential functional relationships between different biomarkers. This information can be used to delineate further subtypes of cancers for developing treatments that can target heterogeneous tumours.

We are involved in a number of collaborative projects developing and testing biomarker panels for ductal carcinoma in situ (DCIS), immunotherapies for ovarian cancer, immune checkpoint inhibitors in breast cancer and immunotherapies in hypermutant cancers (brain, lung, others).

In the DCIS project, a collaboration with Dr. Nofech-Mozes, we are developing a method that uses a panel of biomarkers to correlate with the prognosis of DCIS in malignant progression. We are investigating a panel of immune checkpoint markers, including programmed cell death proteins PD1 and PD-L1, which are involved in suppressing immune response toward cancer cells. We are also developing quantitative tools for studying cellular arrangements that will provide important clues about the potential interactions between cells that express immune checkpoint proteins and the tumour microenvironment.

A multiplexed composite image of breast cancer tissue showing co-localization of seven biomarkers. A multiplexed composite image of breast cancer tissue showing co-localization of seven biomarkers.

We have also developed an alternative method for biomarker multiplexing based on simultaneous immunohistochemical staining with different chromogens and imaging with a multispectral deconvolution system that can unmix and quantify wavelengths of the different chromogens applied.

Whole mount multispectral image acquisitionLeft: colour-rendering index camera, Nuance Zeiss AxioZoom V16 microscope, middle: tiled region of interest of an image, right: triple and quadruple stains in breast cancer.