Spatially Isolated Macrophage Populations Predict Different Outcomes in Colon Cancer

Introduction

Macrophages are sentinel immune cells in tissues, performing a variety of functions including pathogen defense, antigen presentation, phagocytosis of dead cells, and the secretion of signals that drive tissue repair. In tumors, macrophage infiltration often indicates poor patient prognosis. Consequently, tumor-associated macrophages (TAMs) have attracted widespread attention as potential targets for cancer therapy. However, single-target drugs against TAMs (e.g., CSF1 pathway inhibitors) have shown minimal effectiveness against solid tumors, possibly due to neglecting the heterogeneity of macrophages. Clearly, a better understanding of the molecular and functional diversity of macrophages is needed to rationally target them in cancer treatment and predict clinical outcomes.

Paper Information

This paper was authored by Magdalena Matusiak et al. from Stanford University’s Department of Pathology, Center for Biomedical Informatics Research, Department of Biomedical Data Science, Department of Bioengineering, Department of Microbiology and Immunology, Department of Developmental Biology, Institute for Stem Cell Biology and Regenerative Medicine, and the Stanford Cancer Institute. The paper is published in the 2024 issue of the journal Cancer Discovery.

Research Methods

The study applied multiplex protein analysis techniques to depict the spatial distribution of five human macrophage types (LYVE1+, FOLR2+, IL4I1+, NLRP3+, and SPP1+) in normal breast and colon tissues, as well as in malignant tissues such as breast cancer and colorectal cancer. The study was characterized by simultaneously staining other cell types within the tissues, providing a comprehensive understanding of the multicellular environment surrounding these macrophage populations.

Experimental Steps

1. Data Collection and Processing

   • Data on colorectal cancer and breast cancer were collected and integrated from four public single-cell RNA sequencing datasets.
   • Single-cell transcriptome data were used to define markers for macrophage subpopulations and their distribution in healthy and cancerous tissues.

2. Multiplex Protein Analysis

   • The CODEX multiplex tissue imaging technology, including 36 antibodies, was used to simultaneously visualize multiple protein markers in breast and colon tissue microarrays.
   • The spatial distribution of the five macrophage subpopulations was thoroughly depicted through immunofluorescence and IHC staining techniques.

3. Data Analysis

   • Based on multiplex imaging data, cell proximity analysis was conducted to identify the rich neighboring environments of macrophages and their respective cellular compositions.
   • Spatial transcriptomics data were combined to reveal the specific microenvironments and functional nodes where macrophages are distributed in healthy and diseased tissues.

Research Results

1. Spatial Segregation of Macrophage Subpopulations

   • The distribution of the five macrophages (LYVE1+, FOLR2+, IL4I1+, NLRP3+, and SPP1+) in tissues is non-random, occupying distinct spatial environments.
   • LYVE1+ and FOLR2+ macrophages showed high conservativeness in both tumor types, with the FOLR2+ subpopulation being highly enriched in plasma cell microenvironments.
   • IL4I1+ macrophages exhibited high phagocytic activity in regions with frequent epithelial cell death and predicted good outcomes in colorectal cancer patients. SPP1+ macrophages were enriched in hypoxic and necrotic tumor areas, correlating with poor prognosis in colorectal cancer patients.

2. Cross-Tissue Conservation

   • The study found that macrophage subpopulations and their microenvironments are highly conserved across multiple tissues and clinical states, showing characteristics of tissue “foundational building blocks”.

3. Macrophages and Prognosis

   • In colorectal cancer, a high infiltration of IL4I1+ macrophages correlates with good patient prognosis, while SPP1+ macrophages are often associated with poorer prognosis.

4. Role of NLRP3 Inflammasomes

   • The study indicates that NLRP3+ macrophages are associated with regions of aggressive inflammatory responses, such as neutrophil accumulation in breast cancer, colorectal cancer, and Crohn’s disease.

Conclusion and Significance

This paper showcases the diversity and spatial organization of human macrophages in healthy and cancerous tissues, revealing their specific roles in different microenvironments. These findings propose new strategies for using macrophages as targets for tumor therapy, suggesting that targeting specific macrophage subpopulations may be more therapeutically effective than broadly suppressing all macrophages. Furthermore, it highlights the potential role of IL4I1+ macrophages in anti-CD47 and PD-L1 immunotherapy and provides the first histologic evidence of inflammasome formation in human tumors. The research results not only offer a framework for the study of macrophage function in both healthy and diseased states but also make the images obtained through this research publicly accessible via an online resource (https://magdalenamat.github.io/spatial_mac_map/), providing a lasting resource for related fields of study.