Histology-based expression profiling yields novel prognostic markers in human glioblastoma

Abstract

Although the prognosis for patients with glioblastoma is poor, survival is variable, with some patients surviving longer than others. For this reason, there has been longstanding interest in the identification of prognostic markers for glioblastoma. We hypothesized that specific histologic features known to correlate with malignancy most likely express molecules that are directly related to the aggressive behavior of these tumors. We further hypothesized that such molecules could be used as biomarkers to predict behavior in a manner that might add prognostic power to sole histologic observation of the feature. We reasoned that perinecrotic tumor cell palisading, which denotes the most aggressive forms of malignant gliomas, would be a striking histologic feature on which to test this hypothesis. We therefore used laser capture microdissection and oligonucleotide arrays to detect molecules differentially expressed in perinecrotic palisades. A set of RNAs (including POFUT2, PTDSR, PLOD2, ATF5, and HK2) that were differentially expressed in 3 initially studied, microdissected glioblastomas also provided prognostic information in an independent set of 28 glioblastomas that did not all have perinecrotic palisades. On validation in a second, larger independent series, this approach could be applied to other human glioma types to derive tissue biomarkers that could offer ancillary prognostic and predictive information alongside standard histopathologic examination.

FIGURE 1

Laser-capture microdis-section (LCM) process. (A) Typical features of palisades in glioblasto-mas. (B) Frozen section of glioblas-toma with palisading necrosis before LCM. (C) Same section after capture of palisading cells. (D) Laser-captured palisading cells from the same section on the LCM cap. N, necrosis; P, palisading cells; C, “common” tumor cells (see text).

FIGURE 2

(A) Representative results of semiquantitative polymerase chain reaction. The top panel shows gene expression of POFUT2 and the bottom panel shows the β-actin control; the first 3 lanes are control cells and the remaining 3 lanes are pseudopalisading cells from the same case. Note upregulation in 2 of the 3 samples of palisading cells. (B) Immunohistochemistry for HIG2 expression. Most palisading cells show strong cytoplasmic staining compared with the “common” tumor cells. (C) Immunohistochemistry for OLIG2. Approximately half of the “common” tumor cells show nuclear staining but palisading cells are predominantly negative. N, necrosis; P, palisading cells; C, “common” tumor cells.

FIGURE 3

Glioblastoma patient survival based on prognostic markers. Survival curves are drawn with patient survival dichotomized by median expression values for illustration. Cox regression p values are given. (A)POFUT2 (protein-o-fuco-syltransferase 2; p = 0.0005; multiple testing maxT p value = 0.0597); (B)PTDSR (phosphatidylserine factor; p = 0.0021; multiple testing maxT p value = 0.2003).