Probes for INS

Choroid plexus (CP) tumors are rare primary brain neoplasms found most commonly in children and are thought to arise from CP epithelial cells. Sox2 is a transcription factor that not only plays a role in development in the ventricular zone, CP, and roof plate, but also contributes to cancer stemness, tumorigenesis, and drug resistance. Gene expression studies demonstrate aberrant Sox2 expression in human CP tumors, suggesting a role in tumor development. A subset of CP tumors exhibit abnormal NOTCH pathway activity. Using animal models, we previously show that sustained NOTCH activity leads to CP tumors. Immunofluorescence, RT-qPCR, and RNA scope assays have revealed increased Sox2 levels in NOTCH-driven CP tumors compared to wild type CP in mice. To investigate the role of Sox2 in CP tumors, we eliminated Sox2 expression in NOTCH-driven CP tumors. Loss of Sox2 almost completely blocked NOTCH-driven CP tumor growth in these mice, supporting a role for Sox2 in these tumors. Ciliation regulation is one proposed functional pathway for tumorigenesis in CP tumors. Using immunofluorescence assays for cilia (ARL13b) and aquaporin transport protein 1 (AQP1) in combination with super resolution microscopy, we observe a stark contrast between wild type CP epithelial cells which are multiciliated and homogeneously express AQP1, indicative of normal epithelial differentiation, compared to NOTCH-driven CP tumors consisting of mono-ciliated cells with loss of AQP1 expression. In Sox2-deficient NOTCH-driven CP tumors, we observe tumor cells remain mono-ciliated and AQP1-negative, indicating that Sox2 loss does not affect the ciliation machinery. Together this warrants further study into the mechanisms of Sox2 functions in CP tumors. By unraveling the role of Sox2 in CP tumors, we may better understand their origin and biology to ultimately design improved treatment options.

Glioblastoma is believed to originate from nervous system cells; however, a putative origin from vessel-associated progenitor cells has not been considered. We deeply single-cell RNA-sequenced glioblastoma progenitor cells of 18 patients and integrated 710 bulk tumors and 73,495 glioma single cells of 100 patients to determine the relation of glioblastoma cells to normal brain cell types. A novel neural network-based projection of the developmental trajectory of normal brain cells uncovered two principal cell-lineage features of glioblastoma, neural crest perivascular and radial glia, carrying defining methylation patterns and survival differences. Consistently, introducing tumorigenic alterations in naïve human brain perivascular cells resulted in brain tumors. Thus, our results suggest that glioblastoma can arise from the brains' vasculature, and patients with such glioblastoma have a significantly poorer outcome.

Glioblastomas are aggressive brain tumors that are largely immunotherapy resistant. This is associated with immunosuppression and a dysfunctional tumor vasculature, which hinder T cell infiltration. LIGHT/TNFSF14 can induce high endothelial venules (HEVs) and tertiary lymphoid structures (TLS), suggesting that its therapeutic expression could promote T cell recruitment. Here, we use a brain endothelial cell-targeted adeno-associated viral (AAV) vector to express LIGHT in the glioma vasculature (AAV-LIGHT). We found that systemic AAV-LIGHT treatment induces tumor-associated HEVs and T cell-rich TLS, prolonging survival in αPD-1-resistant murine glioma. AAV-LIGHT treatment reduces T cell exhaustion and promotes TCF1+CD8+ stem-like T cells, which reside in TLS and intratumoral antigen-presenting niches. Tumor regression upon AAV-LIGHT therapy correlates with tumor-specific cytotoxic/memory T cell responses. Our work reveals that altering vascular phenotype through vessel-targeted expression of LIGHT promotes efficient anti-tumor T cell responses and prolongs survival in glioma. These findings have broader implications for treatment of other immunotherapy-resistant cancers.

Zika virus (ZIKV) can infect human developing brain (HDB) progenitors resulting in epidemic microcephaly, whereas analogous cellular tropism offers treatment potential for the adult brain cancer, glioblastoma (GBM). We compared productive ZIKV infection in HDB and GBM primary tissue explants that both contain SOX2+ neural progenitors. Strikingly, although the HDB proved uniformly vulnerable to ZIKV infection, GBM was more refractory, and this correlated with an innate immune expression signature. Indeed, GBM-derived CD11b+ microglia/macrophages were necessary and sufficient to protect progenitors against ZIKV infection in a non-cell autonomous manner. Using SOX2+ GBM cell lines, we found that CD11b+-conditioned medium containing type 1 interferon beta (IFNβ) promoted progenitor resistance to ZIKV, whereas inhibition of JAK1/2 signaling restored productive infection. Additionally, CD11b+ conditioned medium, and IFNβ treatment rendered HDB progenitor lines and explants refractory to ZIKV. These findings provide insight into neuroprotection for HDB progenitors as well as enhanced GBM oncolytic therapies.

The stemness-associated markers OCT4, NANOG, SOX2, KLF4 and c-MYC are expressed in numerous cancer types suggesting the presence of cancer stem cells (CSCs). Immunohistochemical (IHC) staining performed on 12 lung adenocarcinoma (LA) tissue samples showed protein expression of OCT4, NANOG, SOX2, KLF4 and c-MYC, and the CSC marker CD44. In situ hybridization (ISH) performed on six of the LA tissue samples showed mRNA expression of OCT4, NANOG, SOX2, KLF4 and c-MYC. Immunofluorescence staining performed on three of the tissue samples showed co-expression of OCT4 and c-MYC with NANOG, SOX2 and KLF4 by tumor gland cells, and expression of OCT4 and c-MYC exclusively by cells within the stroma. RT-qPCR performed on five LA-derived primary cell lines showed mRNA expression of all the markers except SOX2. Western blotting performed on four LA-derived primary cell lines demonstrated protein expression of all the markers except SOX2 and NANOG. Initial tumorsphere assays performed on four LA-derived primary cell lines demonstrated 0-80% of tumorspheres surpassing the 50 µm threshold. The expression of the stemness-associated markers OCT4, SOX2, NANOG, KFL4 and c-MYC by LA at the mRNA and protein level, and the unique expression patterns suggest a putative presence of CSC subpopulations within LA, which may be a novel therapeutic target for this cancer. Further functional studies are required to investigate the possession of stemness traits.