High resolution mapping of the breast cancer tumor microenvironment using integrated single cell, spatial and in situ analysis of FFPE tissue

Overview

This dataset is associated with the preprint “High resolution mapping of the breast cancer tumor microenvironment using integrated single cell, spatial and in situ analysis of FFPE tissue”, posted to bioRxiv October 7, 2022, in which a single breast cancer FFPE tissue block was assayed with a trio of complementary technologies (Chromium, Visium, Xenium). See the preprint for full details on methods and results. On Dec 6, 2022, the In Situ datasets were reprocessed and updated using the improved xenium-1.0.1 analysis pipeline.

How to view data

Interactively explore data with our Xenium Explorer web demo for Replicate 1, or with Xenium Explorer 1.0, by downloading a Xenium Output Bundle below. Load the Gene Groups CSV to create gene groups based on cell type markers. Follow these instructions to view the registered post-Xenium H&E image for Replicate 1 in Xenium Explorer.

Xenium In Situ

The Xenium In Situ platform uses a microscopy based read-out. A 5 μm FFPE tissue section was sectioned onto a Xenium slide and placed in the Xenium Analyzer instrument for multiple cycles of imaging. Each gene has a unique optical signature, facilitating decoding of the target gene, from which a spatial transcriptomic map was constructed across the entire tissue section. The Xenium data was registered with post-Xenium IF / H&E images (workflow is non-destructive to the tissue) and integrated with Chromium and Visium data.

Channel descriptions for IF staining:

• Channel 1 = 1:200 CD20, Goat anti-Mouse IgG-488 , (Abcam, Cat. No. ab219329, Thermo Fisher Cat. No. A-11029)
• Channel 2 = 1:1500 HER2, Goat anti-Rabbit IgG-594, (Abcam Cat. No. ab134182, Abcam Cat. No. ab150088)
• Channel 3 = DAPI (Thermo Fisher, Cat. No. 62248)

Biomaterial

A single formalin-fixed, paraffin-embedded (FFPE) breast cancer tissue block (T2N1M0, Stage II-B, ER+/HER2+/PR−) was obtained from Discovery Life Sciences. Corresponding dissociated tumor cells, fresh frozen (FF) in liquid nitrogen, were also sampled from the same 2.5 cm biopsy. For the Single Cell Gene Expression Flex (FRP) workflow, two 25 μm curls were pooled as a single replicate. 5 μm sections were taken from the FFPE tissue using a microtome. A 5 μm section was taken for Visium CytAssist and two replicate 5 μm sections were taken for Xenium.

Pathology

The tissue block has two Ductal Carcinoma In Situ (DCIS) regions of interest. DCIS1 had invasive carcinoma lesions scattered throughout the stromal connective tissue, surrounding a large aggregate of highly proliferative ductal carcinoma in situ with two central comedo necrotic formations, while DCIS2 had a smaller and round intermediate nuclear grade ductal hyperplasia of solid type with cells that showed mild to moderate variability in size, shape and placement, variable coarse chromatin, and variably prominent nuclei.