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Simple Summary: Hyper-angiogenesis is a characteristic of glioblastoma (GBM), and anti-angio-genesis is a crucial strategy to interfere with tumor progression in GBM therapy. We previously found a high expression of ALDH1A3 in proliferating vasculature in GBM patients, which was associated with poor prognosis. The present study generated two ALDH1A3-overexpressing GBM cells (oxGBMs) and demonstrated a potent pro-angiogenesis function of ALDH1A3 under the different conditions of co-culturing oxGBMs with endothelial cells in vitro and in an angiogenesis model in vivo. Moreover, we identified a mechanism underlying the oxALDH1A3-mediated pro-angiogenic effect involving the paracrine PAI-1 and IL-8 derived from oxGBMs. Blockage of PAI-1 or IL-8 hindered the hyper-angiogenesis phenotype resulting from oxALDH1A3. These findings defined a novel function of ALDH1A3 as an angiogenesis promoter in GBM, beyond its role as a well-known cancer stem cell marker, and highlighted ALDH1A3-PAI-1/IL-8 as a novel targeting signaling for future anti-angiogenesis therapy in GBM. Hyper-angiogenesis is a typical feature of glioblastoma (GBM), the most aggressive brain tumor. We have reported the expression of aldehyde dehydrogenase 1A3 (ALDH1A3) in proliferating vasculature in GBM patients. We hypothesized that ALDH1A3 may act as an angiogenesis promoter in GBM. Two GBM cell lines were lentivirally transduced with either ALDH1A3 (ox) or an empty vector (ev). The angiogenesis phenotype was studied in indirect and direct co-culture of endothelial cells (ECs) with oxGBM cells (oxGBMs) and in an angiogenesis model in vivo. Angiogenesis array was performed in oxGBMs. RT2-PCR, Western blot, and double-immunofluorescence staining were performed to confirm the expression of targets identified from the array. A significantly activated angiogenesis phenotype was observed in ECs indirectly and directly co-cultured with oxGBMs and in vivo. Overexpression of ALDH1A3 (oxALDH1A3) led to a marked upregulation of PAI-1 and IL-8 mRNA and protein and a consequential increased release of both proteins. Moreover, oxALDH1A3-induced angiogenesis was abolished by the treatment of the specific inhibitors, respectively, of PAI-1 and IL-8 receptors, CXCR1/2. This study defined ALDH1A3 as a novel angiogenesis promoter. oxALDH1A3 in GBM cells stimulated EC angiogenesis via paracrine upregulation of PAI-1 and IL-8, suggesting ALDH1A3-PAI-1/IL-8 as a novel signaling for future anti-angiogenesis therapy in GBM. [ABSTRACT FROM AUTHOR]

Simple Summary: Spread of prostate cancer to other parts of the body is responsible for the majority of deaths. Tumour cell epithelial mesenchymal plasticity (EMP) increases their metastatic potential and facilitates their survival in the blood as circulating tumour cells (CTCs). The aim of this study was to molecularly characterise CTCs in a panel of prostate cancer patient-derived xenografts using genes associated with epithelial and mesenchymal phenotypes, and to compare the EMP status of CTCs with their matched primary tumours. The study highlights high heterogeneity in CTC enumeration and EMP gene expression between tumour-bearing mice and within individual blood samples, and therefore caution should be taken when interpreting pooled CTC analyses. Critically, tumour cells were present in the epithelial-mesenchymal hybrid state in the circulation. The study also demonstrates that there is high variation in CTC size, which would introduce sample bias to size-based CTC isolation techniques. Metastasis is the leading cause of cancer-related deaths worldwide. The epithelial-mesenchymal plasticity (EMP) status of primary tumours has relevance to metastatic potential and therapy resistance. Circulating tumour cells (CTCs) provide a window into the metastatic process, and molecular characterisation of CTCs in comparison to their primary tumours could lead to a better understanding of the mechanisms involved in the metastatic cascade. In this study, paired blood and tumour samples were collected from four prostate cancer patient-derived xenograft (PDX) models (BM18, LuCaP70, LuCaP96, LuCaP105) and assessed using an EMP-focused, 42 gene human-specific, nested quantitative RT-PCR assay. CTC burden varied amongst the various xenograft models with LuCaP96 having the highest number of CTCs per mouse (mean: 704; median: 31) followed by BM18 (mean: 101; median: 21), LuCaP70 (mean: 73; median: 16) and LuCaP105 (mean: 57; median: 6). A significant relationship was observed between tumour size and CTC number (p = 0.0058). Decreased levels of kallikrein-related peptidase 3 (KLK3) mRNA (which encodes prostate-specific antigen; PSA) were observed in CTC samples from all four models compared to their primary tumours. Both epithelial- and mesenchymal-associated genes were commonly expressed at higher levels in CTCs compared to the bulk primary tumour, although some common EMT-associated genes (CDH1, VIM, EGFR, EPCAM) remained unchanged. Immunofluorescence co-staining for pan-cytokeratin (KRT) and vimentin (VIM) indicated variable proportions of CTCs across the full EMP axis, even in the same model. EMP hybrids predominated in the BM18 and LuCaP96 models, but were not detected in the LuCaP105 model, and variable numbers of KRT+ and human VIM+ cells were observed in each model. SERPINE1, which encodes plasminogen activator inhibitor-1 (PAI-1), was enriched at the RNA level in CTCs compared to primary tumours and was the most commonly expressed mesenchymal gene in the CTCs. Co-staining for SERPINE1 and KRT revealed SERPINE1+ cells in 7/11 samples, six of which had SERPINE+KRT+ CTCs. Cell size variation was observed in CTCs. The majority of samples (8/11) contained larger CTCs ranging from 15.3 to 37.8 µm, whilst smaller cells (10.7 ± 4.1 µm, similar in size to peripheral blood mononuclear cells (PBMCs)) were identified in 6 of 11 samples. CTC clusters were also identified in 9/11 samples, containing 2–100 CTCs per cluster. Where CTC heterogeneity was observed in the clusters, epithelial-like cells (KRT+VIM−) were located on the periphery of the cluster, forming a layer around hybrid (KRT+VIM+) or mesenchymal-like (KRT−VIM+) cells. The CTC heterogeneity observed in these models emphasises the complexity in CTC isolation and classification and supports the increasingly recognised importance of the epithelial-mesenchymal hybrid state in cancer progression and metastasis. [ABSTRACT FROM AUTHOR]

Simple Summary: Hepatoblastoma (HB), the most common form of childhood liver cancer, is associated with dual mutation and/or dysregulation of the Wnt/β-catenin and Hippo pathways in ~50% of cases. However, this mutational simplicity cannot explain HB's biological and histologic diversity. This discussion focuses upon recent work showing that specific β-catenin mutants are key determinants of this HB variability as well as their metabolic and transcriptional signatures. Dysregulation of the anti-oxidant NFE2L2 pathway also contributes to tumorigenesis by being directly transforming in association with either of the other two factors. The transcriptional overlap of tumors generated by pairs of factors identifies crucial targets that likely mediate HB tumorigenesis, behavior and appearance. Hepatoblastoma (HB), the most common childhood liver cancer, is associated with seven distinct histologic subtypes and variable degrees of clinical aggressiveness and presentation. Yet it is among the least genomically altered tumors known, with about half of HBs showing mutation and/or dysregulation of the Wnt/β-catenin and Hippo pathways. This raises the question of how this mutational simplicity can generate such biological and histologic complexity. Recent work shows that the identity of the underlying β-catenin mutation is a major contributor. Mutation or over-expression of the NFE2L2/NRF2 transcription factor, previously thought only to promote anti-oxidant responses, has also recently been shown to accelerate the growth of HBs generated by mutations in the Wnt/β-catenin and Hippo pathways while imparting novel features such as the tumor-associated cysts and necrosis. Moreover, patient-associated NFE2L2 mutations are overtly transforming when co-expressed with either mutant β-catenin or a Hippo pathway effector. The finding that tumorigenesis can be driven by any two arms of the β-catenin/Hippo/NFE2L2 axis has permitted the identification of a small subset of coordinately regulated tumor-specific transcripts, some of whose levels correlate with inferior long-term outcomes in HB and other cancers. Collectively, these findings begin to provide for more refined and molecularly based classification, survival algorithms and design of chemotherapeutic regimens. [ABSTRACT FROM AUTHOR]