Spatial Profiling of the Tumor Microenvironment

The tumor microenvironment (TME)—the organized area surrounding a tumor that is composed of cancer cells and varieties of immune and other defensive cells—is an exciting locale, where research meets the leading edge of technology. Spatial profiling is beginning to unlock secrets of the TME, holding the promise of more effective cancer immunotherapies. By examining the TME’s diversity of cell types, and expression of RNA and protein markers, researchers are learning more about how the human body fortifies the immune system against the threat of cancer. Here’s a look at how spatial profiling is helping scientists to learn about and bolster these defenses.

Transcriptomics

Today, researchers can perform spatial profiling by measuring the expression of RNA, proteins, or both using different technologies. Vizgen’s MERSCOPE™ platform uses MERFISH (multiplexed error-robust fluorescence in situ hybridization) technology to quantify the spatial distribution of RNAs in tissue specimens. Using libraries designed to probe for cancer genes and others important in the TME, the technology can pinpoint gene expression within cells. “When I analyze tumor samples with MERFISH I can see, with subcellular resolution, exactly the location of tumor cells, non-malignant epithelial cells, immune cells, stromal cells—you name it,” says Colles Price, Oncology Lead at Vizgen. Collaborators have used Vizgen’s platform to study the effects of drugs or genetic manipulations on the gene expression of cells within tumors and the TME.

Price believes that the most exciting developments in our understanding of the TME have come from advances in machine learning and artificial intelligence, which enhance our ability to draw meaningful biological insights from spatial biology data. “In the past five years, we’ve seen this approach applied in digital pathology,” says Price. “[With only a handful of markers,] the research community has been able to stratify patients’ immune cell tissue staining into survival cohorts, significantly predicting those that would respond better to therapy over others.” He adds that the greater number of markers provided by spatial profiling of the TME is likely to accelerate such advances.

Proteomics

Lunaphore’s COMET™ platform uses spatial proteomics technology, with full automation and an open system that allows researchers to use off-the-shelf, label-free antibodies and reagents of their choice. The microfluidics-based multiplex system offers reproducibility that speeds optimization, treating tissue samples gently while avoiding upstream processing steps such as antibody conjugation that can introduce variability. A recent study characterizing the TME of brain metastasis used Lunaphore’s technology to phenotype immune cells and measure intercellular distances.

Diego Dupouy, Co-founder and CTO at Lunaphore, believes that matching patients to the best treatments, and discovering new cancer-fighting medications, are the most exciting advances in the field today. “In recent years, many studies have demonstrated the significance of understanding the landscape of the host immune infiltrates for the prediction of clinical outcomes,” says Dupouy. “This resulted in a fast-growing need for multiplex research, to ultimately develop diagnostic methods that can simultaneously evaluate the immune-tumor-related interactions and its spatial localization in a single tissue sample to more accurately understand and predict patient response to immunotherapy.”

Canopy Biosciences’ CellScape™ platform, using their ChipCytometry™ technology, also provides spatial proteomics at the subcellular and whole-tissue levels, in a benchtop, high-throughput walk-away protein biomarker detection system. The high multiplexing, spatial profiling capabilities, and use of standard antibodies (without proprietary modifications) make it ideal for studying the tumor microenvironment. “Researchers have used this technology to discover rare cell populations in tissue, or profile the spatial relationship between a broad range of cellular phenotypes in the TME,” says Thomas Campbell, Group Product Manager in Spatial Biology at Canopy Biosciences^® (a Bruker company).

Multi-omics

A research group at the Technical University of Munich used Canopy’s ChipCytometry platform to stain for both protein biomarkers and RNA targets in the same tissue specimen. “This advancement will enable researchers to gain additional insights in the TME by uncovering additional rare cell phenotypes through multi-omic analysis of the exact same cells within the spatial context,” says Campbell. He’s excited to move their technology further into clinical development pipelines to improve diagnostics for better patient outcomes.

10x Genomics’ Visium platform also supports multi-omics spatial profiling. “New technologies like Visium are enabling simultaneous measurement of multiple analytes from the same tissue section,” says Abbey Cutchin, Associate Director of Market Development in Oncology at 10x Genomics. “Conceivably, the whole transcriptome, hundreds of proteins, and T and B cell receptor sequences can be profiled in the same tissue section.” Visium can be used for high-resolution transcriptomic mapping of tissues, either alone or in concert with other techniques (single-cell transcriptomics, or immunofluorescence protein detection) for molecular characterization.

A research group at Stanford combined the spatial transcriptomics of Visium with single-cell transcriptomics to examine the architecture of squamous cell carcinoma. They found patterns within the TME and surrounding tissues that were influenced by specific tumor and immune cell dynamics. In particular, they found that the leading edge of the tumor contained a subpopulation of tumor-specific keratinocytes, which are important in intercellular communication and metastasis. “Developing high-resolution maps of tissue architecture is critical to revealing important cancer biology—such as how tumor cells remodel the microenvironmental niche, escape immune surveillance, or metastasize into surrounding tissue,” says Cutchin.

Another group from Emory University used Visium and single-channel transcriptomics to explore phenotypes of CD8+ T cells in metastatic tumors from human brain tissue. The phenotypes varied from naive to exhausted, but some phenotypes were located in particular regions of the tumor. “By linking T cell receptor clones present in Visium and scRNA-seq data, the team was able to map the location of CD8+ T cells with specific phenotypes in the tumor microenvironment,” says Cutchin. “For example, CD8+ T cell clones with an exhausted phenotype are found in the tumor parenchyma, where the signaling milieu is significantly different from the surrounding stroma.”

High-resolution profiling on a subcellular level can reveal even further information about the inner workings of the TME. “[It] will further refine complex cellular relationships and networks, allowing for a deeper understanding of cell-cell and ligand-receptor interactions driving cancer progression,” Cutchin says.

Seeing clinical results

Dupouy notes that the great potential of spatial profiling requires more advanced analytical tools to harness its power fully—for example, to match patient tumor biopsies with appropriate and effective anticancer therapies. “From early biomarker discovery to late-stage translational research, spatial profiling will move into clinical studies,” he says, which will require additional high-throughput multiplexing of large patient cohorts for deep phenotyping. “Capturing intra- and inter-patient variability will reveal pathologically relevant patterns [which can] be correlated with treatment.”

Going forward, Price believes that using machine learning and artificial intelligence to analyze gene expression in individual cells in the TME will lead to further success. “From there we can build predictive patterns of cellular behavior to hypothesize [where these cells came from, and] what these cells might do in the future,” he says. “The past five years have been extremely exciting for spatial biology, but the next five years will be even more incredible.”