Understanding Glioblastoma, One Cell at a Time
Glioma is the most prevalent malignant tumor affecting the central nervous system, and glioblastoma (GBM) is its most aggressive and fatal form.1–3 Despite recent advances in therapy, GBM remains challenging to treat, with a median survival of only 14-17 months and a five-year survival rate of approximately 5%.1,2 A significant factor contributing to these grim statistics is the heterogeneity of tumor cells and their interactions with the tumor microenvironment (TME).3 The TME plays a key role in determining tumor behavior, progression and metastasis, and can also induce resistance to therapeutic agents like chemotherapy and immunotherapy.2,3 In glioblastoma, the TME is highly immunosuppressive,2,3 which has recently been shown to be generated by a symbiotic interaction between the TME and the glioma cells themselves.3 Therefore, targeting this interaction has become a potential strategy to improve the efficacy of anti-tumor therapies in GBM.3
Among TME components, tumor-associated macrophages (TAMs) are the most common immune cells, constituting up to 50% of the tumor mass in GBM.2,3 TAMs play a critical role in promoting tumor angiogenesis, immune evasion and tumor proliferation,2 but attempts to target them have not been successful in clinical trials, likely due to our limited knowledge and characterization of these cells and their behavior.3,4 Recent advances in single-cell technologies have opened the door to a more in-depth understanding, which could pave the way to new, more effective therapeutic options.1,3,4
Single-Cell Advances
In a recent study, researchers from the Institute of Pathology and Southwest Cancer Center in Chongqing, China, used single-cell and spatial transcriptomics to characterize subsets of TAMs known as monocyte-derived TAMs (Mo-TAMs).4 Samples from 51 patients with glioblastoma or glioma were included into the study.4
First, six discrete clusters of Mo-TAMs were identified by functional and spatial characteristics. They decided to focus on one of these clusters that was strongly associated with hypoxia (referred to as Hypoxia-TAM) due to the role of hypoxia as a hallmark of tumors in processes like neovascularization, inflammation, and treatment resistance. The team explored how hypoxia impacts Hypoxia-TAM and how Hypoxia-TAM interacts with other cells to further tumor progression.4
Uncovering Instability
Next, it was discovered that Hypoxia-TAM was spatially associated with destabilized microvessels in the peri-necrotic regions of GBMs. They showed that Hypoxia-TAM oversecrete adrenomedullin (ADM) which destabilizes endothelial adherens junctions, promoting a hyperpermeable tumor vasculature.4
To explore the impact on tumor development and treatment, the researchers used a knockout mouse model to study how human GBM xenografts were impacted by the absence of ADM. This demonstrated a restoration of the VE-cadherin – a marker for endothelial junction adhesion – and improved vascular integrity with reduced vascular leakage in the xenografts.4 Similarly, the use of an ADM antagonist (AMA) in a xenograft model of GBM led to the restoration of VE-cadherin expression, the preservation of endothelial junctions, and a decrease in tumor vascular permeability.4
The team then investigated whether AMA could be combined with anti-tumor agents to improve their efficacy. Dabrafenib is used to treat brain tumors with specific genetic mutations but is usually only able to achieve low intratumoral concentrations, partly due to the hyperpermeable vasculature of brain tumors. In a xenograft model, the team co-administered dabrafenib and AMA and compared the results with those of the control or agent alone. They found that the combination provided synergistic therapeutic effects, reducing tumor burden and leading to a corresponding improvement in survival.4
Advancing Therapeutic Applications
Writing in Cancer Cell, the team says that their findings could lead to strategies to normalize tumor vasculature, particularly by targeting ADM, which they have identified as an “orchestrator” of destabilized endothelial connections. They suggest that in the future, therapies that block ADM could be used as an alternative to focused ultrasound therapy, which is associated with a risk of serious adverse effects. The blockade of ADM could not only enhance the perfusion of anti-tumor drugs in GBM but could also have a role in improving the efficacy of immunotherapy. What’s more, since destabilized vasculature and tumor perfusion are common across brain tumors, the team states that the results could have implications that extend beyond GBM.4
Adapted from Wang et al, 2024, Figure 3E. H&E staining and immunostaining of ADM, galectin-3, and CD31 in the peri-necrotic region of hGBMs. Dashed lines indicate the distances (600 μm/line) from necrotic cores. An area indicated with a square is enlarged and shown in each color or the combination of different colors at the bottom. Scale bar, 200 μm.
Synergy between Methods
These findings were made possible through the use of the TissueFAXS Spectra microscopy-based scanning system and StrataQuest analysis software. TissueFAXS Spectra provides researchers with a versatile tool for whole-slide multispectral imaging and analysis. It is capable of automated scanning of up to 8 standard-size slides (with possibility to upgrade to 120-slide loader) and can scan up to 8 markers in one run, making it ideal for studying the spatial association of markers, as in this study.
The team combined this with TissueGnostics’ most powerful image-processing software, StrataQuest, to provide automated context-based quantitative analysis and took advantage of its ability to remove nonspecific background and optimize marker identification. StrataQuest now comes equipped with over 50 apps to perform automated analysis, but advanced users can also develop their own solutions, making this a highly adaptable, powerful tool for a diverse range of imaging purposes.
This paper by Wang and colleagues shows a prime example of how spatial transcriptomics and tissue cytometry can be synergistically combined to verify the results or deepen the understanding of TME processes both on mRNA and protein level. By utilizing multispectral imaging and image anaylsis of multiplex immunostaining for ADM, vessel marker CD31, and Mo-TAM marker galectin-3, the researchers could show the influence of ADM on tumor vasculature in glioblastoma.
Not sure if StrataQuest is a suitable image analysis solution for your project? Take a look at a case study showcasing the lastest features of the new StrataQuest 8 version. If you plan to perform your own project using spatial transcriptomics and search for a solution to verify your findings by staining, contact our team to consult what we can offer to extract maximum of information from your samples.
References
- Batchu S, Hanafy KA, Redjal N, et al. Single-cell analysis reveals diversity of tumor-associated macrophages and their interactions with T lymphocytes in glioblastoma. Sci Rep. 2023;13(1):20874. doi: 10.1038/s41598-023-48116-2.
- Tang F, Wang Y, Zeng Y, et al. Tumor-associated macrophage-related strategies for glioma immunotherapy. NPJ Precis Oncol. 2023;7(1):78. doi: 10.1038/s41698-023-00431-7.
- Khan F, Pang L, Dunterman M, L et al. Macrophages and microglia in glioblastoma: heterogeneity, plasticity, and therapy. J Clin Invest. 2023;133(1):e163446. doi: 10.1172/JCI163446.
- Wang W, Li T, Cheng Y, et al. Identification of hypoxic macrophages in glioblastoma with therapeutic potential for vasculature normalization. Cancer Cell. 2024;42(5):815-832.e12.
