Identification of Therapeutic Potential of Hypoxic Macrophages in Glioblastoma

Academic Background

Glioblastoma (GBM) and isocitrate dehydrogenase (IDH) mutant gliomas are the most common malignant brain tumors in adults. Tumor-associated macrophages (TAMs) are the major immune cell infiltrates in these tumors. These cells interact directly with malignant cells, promote tumor progression, and construct an immunosuppressive microenvironment. Therefore, TAMs have become an attractive therapeutic target and are widely used in various therapeutic strategies to inhibit TAM recruitment or survival, restore TAM phagocytosis, and reprogram TAM phenotypes. However, the effectiveness of large-scale clinical trials is currently limited, mainly because of the cellular diversity and plasticity of TAMs.

Source Information

The study, conducted by Wenying Wang, Tianran Li, Yue Cheng, and others, was published in the journal “Cancer Cell.” The research was supported by multiple scientific institutions, including the Third Military Medical University of China, Southwest Hospital, and Huazhong University of Science and Technology, and was published on May 13, 2024. Corresponding authors are Yi-Fang Ping (pingyifang@126.com), Xiu-Wu Bian (bianxiuwu@263.net), and Yu Shi (shiyu@tmmu.edu.cn).

Research Process

Experimental Design and Methods

The goal of the study was to identify and analyze hypoxic macrophages (Hypoxia-TAMs) in IDH wild-type GBM and IDH mutant gliomas and explore their potential role in tumor vascular normalization and anti-tumor efficacy. The study combined single-cell RNA sequencing (scRNA-seq) with spatial transcriptomics to map the molecular and functional heterogeneity landscape from tumor samples of 51 patients.

Sample Collection and Sequencing

Samples were collected from 51 glioma patients (including 40 GBM-IDHwt, 8 IDH mutant astrocytomas, and 3 IDH mutant oligodendrogliomas) for single-cell RNA sequencing, and a cell lineage annotation algorithm was used for cell lineage annotation.

Preliminary Data Analysis

Unbiased clustering and cell lineage annotation identified 360,214 single cells classified into 12 major lineages, with tumor cells accounting for 46% and myeloid cells for 30%. Further analysis clustered myeloid cells into monocytes, TAMs, microglial TAMs (MG-TAMs), etc., and identified five MG-TAM subtypes and four dendritic cell subtypes.

Identification and Functional Analysis of Hypoxia-TAMs

Using gene module enrichment analysis (GSVA) and unbiased clustering, a Hypoxia-TAM cluster was identified, exhibiting significant hypoxia response characteristics. Non-negative least squares regression (NNLS) analysis confirmed the presence and changes of these clusters in different types of gliomas, with a higher frequency in GBM-IDHwt. Hypoxia-TAMs in hypoxic regions exhibited high levels of glioblastoma (HGBM) cell interaction and spatial association with blood vessels, promoting high vascular permeability by secreting adrenomedullin (ADM), resulting in endothelial cell adhesion junction disruption.

Experimental Results and Analysis

Transcriptional Characteristics and Spatial Distribution of Hypoxia-TAMs

Spatial transcriptomic analysis of HGBM samples indicated that Hypoxia-TAMs primarily localize around necrotic regions and stimulate the disruption of endothelial cell adhesion junctions through ADM secretion, leading to significant tumor vascular leakage. Targeted ADM therapy (including gene knockout and pharmacological blockade) restored vascular integrity, increased intratumoral drug concentration, and enhanced the efficacy of the anti-tumor drug dabrafenib.

Bioinformatics and Functional Experiments

Further bioinformatics and functional experiments revealed the polarization mechanism of Hypoxia-TAMs, identifying SPARC produced by tumor cells and hypoxia-induced lactate as key microenvironmental factors. Activation of the p50 transcription factor in the NF-κB signaling pathway was crucial for the polarization of Hypoxia-TAMs.

Summary of Major Findings

1. Identification of Hypoxia-TAMs: Hypoxia-TAMs exhibited significant hypoxia response characteristics, concentrated around necrotic areas of the tumor.
2. Role of ADM in Vascular Abnormalities: Hypoxia-TAMs secreted ADM, disrupting endothelial cell adhesion junctions and leading to vascular abnormalities.
3. Therapeutic Potential: Targeting ADM restored vascular integrity, significantly improving drug delivery and anti-tumor efficacy.

Conclusions and Value

This study systematically revealed the molecular and functional heterogeneity of Hypoxia-TAMs in gliomas and their central role in vascular abnormalities through high-precision single-cell transcriptomics and spatial transcriptomics. The research suggests that targeted interventions against ADM or its receptor CRLR can restore vascular structural integrity, enhance drug delivery efficiency, and provide new strategies for tumor therapy.

Scientific Significance and Application Prospects

This study not only reveals the value of Hypoxia-TAMs as a new therapeutic target but also provides new insights into the dynamics of immune cells in the tumor microenvironment and their relationship with tumor development. These findings are expected to have far-reaching impacts on therapeutic strategies for gliomas and other tumors, particularly in enhancing drug permeability and immunotherapy efficacy.