The role of angiogenesis in Group 3 medulloblastoma pathogenesis and survival

Abstract

Background: Of the 4 medulloblastoma subgroups, Group 3 is the most aggressive but the importance of angiogenesis is unknown. This study sought to determine the role of angiogenesis and identify clinically relevant biomarkers of tumor vascularity and survival in Group 3 medulloblastoma.

Methods: VEGFA mRNA expression and survival from several patient cohorts were analyzed. Group 3 xenografts were implanted intracranially in nude rats. Dynamic susceptibility weighted (DSC) MRI and susceptibility weighted imaging (SWI) were obtained. DSC MRI was used to calculate relative cerebral blood volume (rCBV) and flow (rCBF). Tumor vessel density and rat vascular endothelial growth factor alpha (VEGFA) expression were determined.

Results: Patient VEGFA mRNA levels were significantly elevated in Group 3 compared with the other subgroups (P < 0.001) and associated with survival. Xenografts D283, D341, and D425 were identified as Group 3 by RNA hierarchical clustering and MYC amplification. The D283 group had the lowest rCBV and rCBF, followed by D341 and D425 (P < 0.05). These values corresponded to histological vessel density (P < 0.05), rat VEGFA expression (P < 0.05), and survival (P = 0.002). Gene set enrichment analysis identified 5 putative genes with expression profiles corresponding with these findings: RNH1, SCG2, VEGFA, AGGF1, and PROK2. SWI identified 3 xenograft-independent categories of intratumoral vascular architecture with distinct survival (P = 0.004): organized, diffuse microvascular, and heterogeneous.

Conclusions: Angiogenesis plays an important role in Group 3 medulloblastoma pathogenesis and survival. DSC MRI and SWI are clinically relevant biomarkers for tumor vascularity and overall survival and can be used to direct the use of antivascular therapies for patients with Group 3 medulloblastoma.

Keywords: Group 3; VEGF; angiogenesis; dynamic susceptibility contrast MRI; medulloblastoma.

Fig. 1

VEGF expression is elevated in patients with Group 3 medulloblastoma and is associated with survival. (A) Box and whisker plots from 3 independent cohorts all demonstrating significantly elevated VEGFA mRNA expression in Group 3 medulloblastoma compared with the other 3 subgroups. Top plot from the Medulloblastoma Advanced Genomics International Consortium (MAGIC), middle plot from St Jude Children’s Research Hospital, and lower plot from multiple US institutions (middle line = median, box = upper and lower quartiles). “Relative fold change” is relative to background mRNA expression. (B) Kaplan–Meier survival curves of patients in 2 independent cohorts comparing all patients with “high” versus “low” VEGFA mRNA expression. Top curve from Heidelberg, bottom 2 curves from the same cohort of multiple US institutions. Upper P-value is derived from log-rank test. Lower “Bonf P” is Bonferroni P-value corrected for multiple comparisons used to determine cut points of “high” versus “low” VEGFA expression.

Fig. 2

Hierarchical clustering (HCL) of gene expression from 500 genes of the 7 patient-derived xenografts analyzed.

Fig. 3

Group 3 medulloblastoma vascularity as determined by DSC MRI corresponds to survival. D425 tumors had significantly elevated vascularity as determined by both (A) rCBF and (B) rCBV compared with D283 tumors. Columns are mean values and error bars are standard deviation. (C) Parametric rCBV maps of representative animals from each xenograft cohort demonstrating tumor size and location (red circle). (D) Kaplan–Meier survival curve comparing the 3 xenograft groups.

Fig. 4

Vessel density and rat VEGFA protein expression correspond with rCBV, rCBF, and survival. (A) D283 tumors had significantly less vessel density compared with both D341 and D425 tumors. Arrows indicate vessels. HPF is high power field. (B) Enzyme-linked immunosorbent assay of rat brain VEGFA expression demonstrates a significant increase in D425 tumors compared with D281 tumors. Columns are mean values and error bars are standard deviation.

Fig. 5

Hierarchical clustering of gene expression from 48 key angiogenesis genes of the 3 patient-derived xenografts. Relative expression of the genes RNH1, SCG2, VEGFA, AGGF1, and PROK2 among the 3 tumor types correlated with DSC MRI, vessel density, rat VEGFA expression, and OS.

Fig. 6

There are 3 categories of MRI vessel patterns in Group 3 medulloblastoma found in both rodent xenograft models and in humans: organized, diffuse microvascular, and heterogeneous. (A) Examples of each category in rodents (top row) as noted on SW MRI using ferumoxytol as an intravascular contrast agent. Examples of each category in patients with Group 3 medulloblastoma (bottom row) as noted on GRE MRI without contrast. White circles highlight tumor. (B) Kaplan–Meier survival curve of intracranial Group 3 xenograft models.