Spatial transcriptomics reveals the role of immune cells in breast and ovarian cancer at the single cell to tissue microenvironment level

Join us for a two-part session for our May 16 installment of the Women’s Health Series!

Session I: The spatially resolved single-cell atlas of the tumor immune architecture revealed the central role of IFN-alpha and plasmacytoid dendritic cells in triple-negative breast cancer

Background: Multiple studies have confirmed the central role of preexisting immune response measured by stromal tumor-infiltrating lymphocytes (sTILs) in triple-negative breast cancer (TNBC). Emerging studies showed that not only the number of TILs but also the location of TILs is important. There are 3 distinct immune architectures described based on the amount and locations of TILs, namely immune enriched (IN), immune excluded (IE), and immune desert (ID). Here we evaluated outcomes and characteristics associated with each immune landscape.

Methods: NanoString Digital Spatial Profiling (DSP) and CosMx, a spatial multi-omics single-cell imaging platform, were performed in 75 samples from the Mayo Clinic (MC) TNBC cohort (Leon-Ferre BCRT 2018). NanoString IO360 was performed in 114 samples from the FinXX trial (NCT00114816). Firstly, tumors with sTIL quantified by H&E ≤ 30% were classified as ID. The rest of the tumors with high sTIL > 30% were categorized according to the intratumoral CD8 protein expression by DSP, with IE having intratumoral CD8 in the lower median and IN having intratumoral CD8 in the upper median. Chi-square test, gene set enrichment, Cox regression, and Kaplan-Meier analysis were used. Differential expression listed as log 2-fold change (FC) was estimated from the linear mixed model with significance defined as two-sided p< 0.05.

Session II: Digital spatial transcriptomic reveals different epithelial and immune cells signatures across endometriosis-related diseases: From simple ovarian endometriosis to ovarian cancer

Abstract: Endometriosis represents a pressing medical concern, imposing a relevant burden on patients including potential malignant transformation into endometriosis-associated ovarian cancer (EAOC). Despite the process of cancer development remains unclear, immune cells abnormalities arose as putative biomarkers in the neoplastic progression. One-third of ovarian endometriosis cases exhibit PD-1/PD-L1 expression levels comparable to those observed in cancer. To substantiate the biological basis of these findings, this study aimed to elucidate the spatial transcriptomic profile of epithelial and immune components from different endometriosis-related conditions, along with their genetic interplay. Formalin-fixed paraffine embedded (FFPE) specimens from 4 ovarian endometriosis (OE) cases, two of which displayed elevated levels of PD-1/PD-L1 expression and were referred to as cancer-like endometriosis (CLE), 1 Clear Cell Ovarian Cancer (CCOC) and 1 Endometrioid Ovarian Cancer (EOC) were cut in 5um section for spatial transcriptomic analysis using GeoMX Human Whole Transcriptome Atlas (WTA) probe-set (>18000 genes). For all sections, the regions of interest (ROIs) were identified using a set of morphology markers: anti-human CD3, CD68 and PanCK antibodies. Spatial transcriptomic analysis revealed condition-specific gene enrichment of epithelial and immune cells across the different conditions. In spite of similar H&E findings, PanCK^pos epithelial cells from CLE showed an increased expression of genes involved in stress response and defense regulation pathways (e.g. CXCL6, S100A9, and PECAM-1), when compared to OE, in which PanCK^pos cells were enriched in DUSP1/2 and VIM. Interestingly, in CCOC, location-specific gene-enrichment was found in PanCK^pos cells, with a significant up regulation of tumor suppressor genes (e.g. EGR-1, NR4A1) in normal-looking peri-tumoral epithelium compared to cancerous intra-tumoral PanCK^pos cells. Similarly, in the immune compartment, spatial transcriptomic analysis revealed that CD3^pos cells from CLE displayed an up-regulation of genes involved in IL-17 signaling, response to oxydative stress and complement activation (e.g. CXCL6, HMOX1, S100A9, and PECAM-1), when compared to OE. Moreover, in CCOC, CD68^pos cells of the tumoral mass showed over-expression of tumor growth and progression markers (e.g. SPP1) compared to their counterpart in the outer area of the tumoral mass. In this first comprehensive spatial transcriptomic study of endometriosis-related diseases with GeoMx WTA, our findings uncovered remarkable differences in epithelial and immune cells among different stages and tissue locations. Importantly, CLE emerged as a transitional entity with a unique immune profile. Ongoing analysis of six additional samples will further refine our results.