Passage of systemically delivered pharmacological agents into the brain is largely blocked by the blood-brain-barrier (BBB), an organotypic specialization of brain endothelial cells (ECs). Tumor vessels in glioblastoma (GBM), the most common malignant brain tumor in humans, are abnormally permeable, but this phenotype is heterogeneous and may differ between the tumor’s center and invasive front. Here, through single-cell RNA sequencing (scRNA-seq) of freshly isolated ECs from human glioblastoma and paired tumor peripheral tissues, we have constructed a molecular atlas of human brain ECs providing unprecedented molecular insight into the heterogeneity of the human BBB and its molecular alteration in glioblastoma. We identified 5 distinct EC phenotypes representing different states of EC activation and BBB impairment, and associated with different anatomical locations within and around the tumor. This unique data resource provides key information for designing rational therapeutic regimens and optimizing drug delivery.

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Background: Clinical studies have shown that first-line use of anti-angiogenetic therapy can prolong progression-free survival but little progress has been made in extending the overall survival of the patients. We explored the role of ELK3 in glioma angiogenesis to improve and design more efficacious therapies. Methods: A tissue microarray and immunohistochemistry analysis were used to determine the expression of ELK3 protein in 400 glioma patients. Cell proliferation, metastasis, cell cycle, and apoptosis were monitored in U87 and U251 cells using CCK-8, EdU, transwell assays, and flow cytometry. A tube-formation assay, a rat aorta ring sprouting assay, and a matrigel plug assay were performed to examine the antiangiogenic activity of ELK3. An ELISA, Western blot, and correlation analysis of the CGGA dataset were used to detect the association between ELK3 and VEGF-A or ELK3 and HIF-1 $$\alpha$$α . Besides, orthotopic transplantation in nude mice and histopathological and immunological analysis of in vitro tumors were used to explore the effect of ELK3 on tumor progression and median survival. Results: ELK3 was upregulated in glioma tissues and associated with a poor prognosis. In vitro, ELK3 promoted cell proliferation and cell cycle progression, induced metastasis, and suppressed apoptosis. Then, silencing ELK3 inhibited VEGF-A expression and secretion by facilitating HIF-1 $$\alpha$$α degradation via ubiquitination. Finally, knockdown ELK3 inhibited tumor progression and angiogenesis in vitro and in vivo, as well as prolonged nude mice’s median survival. Conclusions: Our findings first evidenced that ELK3 is crucial for glioma because it promotes angiogenesis by activating the HIF-1 $$\alpha$$α /VEGF-A signaling axis. Therefore, we suggest that ELK3 is a prognostic marker with a great potential for glioma angiogenesis and ELK3-targeted therapeutic strategies might hold promise in improving the efficacy of anti-angiogenic therapies.

Cancer immunotherapies have shown sustained clinical success in treating primary non-small-cell lung cancer (NSCLC). However, patients with brain metastasis are excluded from the trials because the brain is viewed traditionally as an immune-privileged organ. The composition and properties of tumor-infiltrating myeloid cells in metastatic brain tumors are mostly unknown. To depict the baseline landscape of the composition, gene signature, and functional states of these immune cells, we performed - single-cell RNA sequencing (scRNAseq) for 12,196cellsafter data preprocessing, including 2,241 immunecells from three surgically removed brain lesions of treatment-naïve NSCLC patients. We found a lack of T lymphocyte infiltration and activation, as well as the vast expansion of tumor-associated macrophage(TAM) in the brain lesions of NSCLC patients. By comparing our scRNAseq dataset with published data from early and late-stage primary NSCLC tumors, we showed that this compromised T cell response is unique to brain lesions. We identified a unique alternative activation (M2) gene expression pattern of the TAM in the brain metastasis and a lack of known T cell co-stimulator expression. Accumulation of M2 polarized TAM may, therefore, cause the comprised anti-tumor T cell response in metastatic brain lesions. These findings can contribute to the design of new immunotherapy strategies for NSCLC patients with brain metastasis.