WT1 Clone 6F-H2 Cytoplasmic Expression Differentiates Astrocytic Tumors from Astrogliosis and Associates with Tumor Grade, Histopathology, IDH1 Status, Apoptotic and Proliferative Indices: A Tissue Microarray Study

Abstract

Objectives: This tissue microarray (TMA) immunohistochemical (IHC) study elucidates the role of Wilms' tumor 1 protein (WT1) in diagnosis and prognostication of astrocytic tumors.

Methods: IHC was applied to 75 astrocytic lesions (18 astrogliosis and 57 astrocytic tumors) using antibodies directed against WT1 clone 6F-H2, isocitrate dehydrogenase 1(IDH1), Bcl2 and Ki67. WT1 IHC staining was evaluated and scored blindly by 2 pathologists. Bcl2 and Ki67 scores and labelling indices were assessed and IDH1 status determined. Statistical analysis was performed using the appropriate methodology.

Results: WT1 cytoplasmic expression was detected in 89.5% of astrocytic tumors but not in astrogliosis. Positive WT1 differentiated astrocytic tumors (92.6% accuracy) and grade II diffuse astrocytomas (93.5% accuracy) from astrogliosis with high sensitivity, specificity and positive predictive values (p<0.001). Increased WT1 score significantly associated higher Bcl2 and Ki67 labelling indices, increasing WHO tumor grade and tumor's histopathologic type (p<0.05) showing staining pattern variability by tumor entity and cell type. Glioblastomas, gliosarcomas and subependymal giant cell astrocytomas were the most frequently WT1 expressing tumors with frequent +3 WT1 score. About 21.4% of pilocytic astrocytomas had +3WT1 score in association with increased Bcl2 and Ki67 indices. Low WT1 scores in grade II and III diffuse astrocytomas were linked to the high frequency of IDH1 positivity, and were associated with low Bcl2 and Ki67 labelling indices. In glioblastomas, WT1 significantly associated poor prognostic variables: older age, negative-IDH1 status, high Bcl2 and Ki67 labelling indices (p=0.04, <0.001, =0.001 and.

Keywords: Astrocytic tumors; Astrogliosis; Bcl2/Ki67; IDH1; WT1.

Figure 1

WT1 Expression in Different Astrocytic Lesions. Astrogliosis Showing Negatively-Stained Reactive Astrocytes Compared to Positive Endothelial Internal Control (a, 100x); score +3 subependymal giant cell astrocytoma (b, 200x); pilocytic astrocytomas with score +2 loose plexiform (c, x100) and score +3 dense plexiform patterns (d, 100x); diffuse astrocytoma (e, 100x) and anaplastic astrocytoma showing +1 focal staining pattern (f; 100x); score +3 glioblastoma (g, 40x) showing palisaded positive cells delineating a focus of necrosis (h, 100x); score +2 glioblastoma (i, 100x); and +3 gliosarcoma (j, 100x). WT1 positive luminal endothelial layer but negative proliferating tumor vessels are noted (d, g, i; arrows)

Figure 2

WT1 Score Association with the Prognostic Markers in High- and Low- Astrocytoma Grades. Grade IV, WT1 score+3 glioblastoma with IDH1 positive status, high Bcl2 and high Ki67 indices (a-e, 100x); and a grade II, WT1 score+2 diffuse astrocytoma with gemistocytic differentiation (inset) showing IDH1 positive status, intermediate Bcl2 index (f-i, 100x) and low Ki67 index (j, 200x)

Figure 3

Statistical Analysis of WT1 Expression in Astrocytomas Distinguished by WHO Grade. Bars represent percentages of tumors within each WT1 score (0, +1, +2, +3) displayed in relation to age, IDH1 status, Bcl2 and Ki67 labelling indices respectively. Corresponding p-values are shown, * p-value is significant if <0.05