The HOPX and BLBP landscape and gliogenic regions in developing human brain.

Outer radial glial cells (oRGs) give rise to neurons and glial cells and contribute to cell migration and expansion in developing neocortex. HOPX has been described as a marker of oRGs and possible actor in glioblastomas. Recent years' evidence points to spatiotemporal differences in brain development which may have implications for the classification of cell types in the central nervous system and understanding of a range of neurological diseases. Using the Human Embryonic/Fetal Biobank, Institute of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark, HOPX and BLBP immunoexpression was investigated in developing frontal, parietal, temporal and occipital human neocortex, other cortical areas and brain stem regions to interrogate oRG and HOPX regional heterogeneity. Furthermore, usage of high-plex spatial profiling (Nanostring GeoMx® DSP) was tested on the same material. HOPX marked oRGs in several human developing brain regions as well as cells in known gliogenic areas but did not completely overlap with BLBP or GFAP. Interestingly, limbic structures (e.g. olfactory bulb, indusium griseum, entorhinal cortex, fimbria) showed more intense HOPX immunoreactivity than adjacent neocortex and in cerebellum and brain stem, HOPX and BLBP seemed to stain different cell populations in cerebellar cortex and corpus pontobulbare. DSP screening of corresponding regions indicated differences in cell type composition, vessel density and presence of apolipoproteins within and across regions and thereby confirming the importance of acknowledging time and place in developmental neuroscience.

Plasma IL-8 and ICOSLG as prognostic biomarkers in glioblastoma.

BACKGROUND: CNS immune privilege has been challenged in recent years. Glioblastoma (GBM) immune dysfunction includes complex interactions with the immune system outside the CNS. The aim of this study was to determine diagnostic and prognostic potential of immune-related proteins in plasma in GBM and interrogate biomarker presence in the brain tumor microenvironment (TME). METHODS: One hundred and fifty-eight patients with glioma WHO grade II-IV were included. Plasma collected at surgery was screened for 92 proteins using proximity extension assay technology and related to clinical outcome. Secretion and expression of candidate prognostic biomarkers were subsequently analyzed in 8 GBM cell lines and public RNAseq data. RESULTS: Plasma levels of 20 out of 92 screened proteins were significantly different in patients with GBM compared to patients with astrocytoma WHO grade II-III. High plasma interleukin-8 (IL-8) (hazard ratio [HR] = 1.52; P = .0077) and low CD244 (HR = 0.36; P = .0004) were associated with short progression-free survival and high plasma IL-8 (HR = 1.40; P = .044) and low ICOS ligand (ICOSLG) (HR = 0.17; P = .0003) were associated with short overall survival (OS) in newly diagnosed patients with GBM. A similar trend was found for ICOSLG (HR = 0.34; P = .053) in recurrent GBM. IL-8 was mostly secreted and expressed by mesenchymal GBM cell lines and expressed by vascular cells and immune cells in the TME. This was also the case for ICOSLG, although less consistent, and with additional expression in tumor-associated oligodendrocytes. CONCLUSIONS: High plasma IL-8 and low ICOSLG at surgery are associated with short OS in newly diagnosed GBM. Source of plasma ICOSLG may be found outside the TME.

Perspective: targeting VEGF-A and YKL-40 in glioblastoma - matter matters.

Glioblastomas (GBM) are heterogeneous highly vascular brain tumors exploiting the unique microenvironment in the brain to resist treatment and anti-tumor responses. Anti-angiogenic agents, immunotherapy, and targeted therapy have been studied extensively in GBM patients over a number of decades with minimal success. Despite maximal efforts, prognosis remains dismal with an overall survival of approximately 15 months.Bevacizumab, a humanized anti-vascular endothelial growth factor (VEGF) antibody, underwent accelerated approval by the U.S. Food and Drug Administration in 2009 for the treatment of recurrent GBM based on promising preclinical and early clinical studies. Unfortunately, subsequent clinical trials did not find overall survival benefit. Pursuing pleiotropic targets and leaning toward multitarget strategies may be a key to more effective therapeutic intervention in GBM, but preclinical evaluation requires careful consideration of model choices. In this study, we discuss bevacizumab resistance, dual targeting of pro-angiogenic modulators VEGF and YKL-40 in the context of brain tumor microenvironment, and how model choice impacts study conclusions and its translational significance.

Systemic Immune Modulation in Gliomas: Prognostic Value of Plasma IL-6, YKL-40, and Genetic Variation in YKL-40.

Background: Complex local and systemic immune dysfunction in glioblastoma (GBM) may affect survival. Interleukin (IL)-6 and YKL-40 are pleiotropic biomarkers present in the tumor microenvironment and involved in immune regulation. We therefore analyzed plasma IL-6, YKL-40, and genetic variation in YKL-40 and explored their ability to distinguish between glioma subtypes and predict survival in GBM. Methods: One hundred fifty-eight patients with glioma WHO grade II-IV were included in the study. Plasma collected at surgery was analyzed for IL-6 and YKL-40 (CHI3L1) by ELISA. CHI3L1 rs4950928 genotyping was analyzed on whole-blood DNA. Results: Neither plasma IL-6 nor YKL-40 corrected for age or rs4950928 genotype could differentiate GBM from lower grade gliomas. GC and GG rs4950928 genotype were associated with lower plasma YKL-40 levels (CC vs. GC, p = 0.0019; CC vs. GG, p = 0.01). Only 10 and 14 out of 94 patients with newly diagnosed GBM had elevated IL-6 or YKL-40, respectively. Most patients received corticosteroid treatment at time of blood-sampling. Higher pretreatment plasma IL-6 was associated with short overall survival (OS) [HR = 1.19 (per 2-fold change), p = 0.042] in univariate analysis. The effect disappeared in multivariate analysis. rs4950928 genotype did not associate with OS [HR = 1.30, p = 0.30]. In recurrent GBM, higher YKL-40 [HR = 2.12 (per 2-fold change), p = 0.0005] but not IL-6 [HR = 0.99 (per 2-fold change), p = 0.92] were associated with short OS in univariate analysis. Conclusion: In recurrent GBM high plasma YKL-40 may hold promise as a prognostic marker. In newly diagnosed GBM perioperative plasma IL-6, YKL-40, and genetic variation in YKL-40 did not associate with survival. Corticosteroid use may complicate interpretation of results.

Astrogliogenesis in human fetal brain: complex spatiotemporal immunoreactivity patterns of GFAP, S100, AQP4 and YKL-40.

The astroglial lineage consists of heterogeneous cells instrumental for normal brain development, function and repair. Unfortunately, this heterogeneity complicates research in the field, which suffers from lack of truly specific and sensitive astroglial markers. Nevertheless, single astroglial markers are often used to describe astrocytes in different settings. We therefore investigated and compared spatiotemporal patterns of immunoreactivity in developing human brain from 12 to 21 weeks post conception and publicly available RNA expression data for four established and potential astroglial markers - GFAP, S100, AQP4 and YKL-40. In the hippocampal region, we also screened for C3, a complement component highly expressed in A1-reactive astrocytes. We found diverging partly overlapping patterns of the established astroglial markers GFAP, S100 and AQP4, confirming that none of these markers can fully describe and discriminate different developmental forms and subpopulations of astrocytes in human developing brain, although AQP4 seems to be the most sensitive and specific marker for the astroglial lineage at midgestation. AQP4 characterizes a brain-wide water transport system in cerebral cortex with regional differences in immunoreactivity at midgestation. AQP4 distinguishes a vast proportion of astrocytes and subpopulations of radial glial cells destined for the astroglial lineage, including astrocytes determined for the future glia limitans and apical truncated radial glial cells in ganglionic eminences, devoid of GFAP and S100. YKL-40 and C3d, previously found in reactive astrocytes, stain different subpopulations of astrocytes/astroglial progenitors in developing hippocampus at midgestation and may characterize specific subpopulations of 'developmental astrocytes'. Our results clearly reflect that lack of pan-astrocytic markers necessitates the consideration of time, region, context and aim when choosing appropriate astroglial markers.

Outcome of Bevacizumab Therapy in Patients with Recurrent Glioblastoma Treated with Angiotensin System Inhibitors.

PURPOSE: Antihypertensive therapy may improve bevacizumab efficacy in cancer patients. We examined efficacy and toxicity of angiotensin system inhibitors (ASI) and other antihypertensive drugs in bevacizumab treated recurrent glioblastoma patients. METHODS: We retrospectively combined a national prescription registry with a clinical database with recurrent glioblastoma patients (n = 243). RESULTS: Patients who initiated ASI after bevacizumab (n = 26) showed a tendency towards improved progression-free survival and overall survival (OS) with hazard rate (HR) reductions (HR = 0.70 and HR = 0.79, respectively). Calcium antagonists during bevacizumab therapy significantly improved OS (HR = 0.57). CONCLUSIONS: Overall the study supports a potential beneficial effect of antihypertensive treatment on prognosis of bevacizumab treated glioblastoma patients.