PI3 kinase mutations and mutational load as poor prognostic markers in diffuse glioma patients

Abstract

Introduction: Recent advances in molecular diagnostics allow diffuse gliomas to be classified based on their genetic changes into distinct prognostic subtypes. However, a systematic analysis of all molecular markers has thus far not been performed; most classification schemes use a predefined and select set of genes/molecular markers. Here, we have analysed the TCGA dataset (combined glioblastoma (GBM) and lower grade glioma (LGG) datasets) to identify all prognostic genetic markers in diffuse gliomas in order to generate a comprehensive classification scheme.

Results: Of the molecular markers investigated (all genes mutated at a population frequency >1.7 % and frequent chromosomal imbalances) in the entire glioma dataset, 57 were significantly associated with overall survival. Of these, IDH1 or IDH2 mutations are associated with lowest hazard ratio, which confirms IDH as the most important prognostic marker in diffuse gliomas. Subsequent subgroup analysis largely confirms many of the currently used molecular classification schemes for diffuse gliomas (ATRX or TP53 mutations, 1p19q codeletion). Our analysis also identified PI3-kinase mutations as markers of poor prognosis in IDH-mutated + ATRX/TP53 mutated diffuse gliomas, median survival 3.7 v. 6.3 years (P = 0.02, Hazard rate (HR) 2.93, 95 % confidence interval (CI) 1.16 - 7.38). PI3-kinase mutations were also prognostic in two independent datasets. In our analysis, no additional molecular markers were identified that further refine the molecular classification of diffuse gliomas. Interestingly, these molecular classifiers do not fully explain the variability in survival observed for diffuse glioma patients. We demonstrate that tumor grade remains an important prognostic factor for overall survival in diffuse gliomas, even within molecular glioma subtypes. Tumor grade was correlated with the mutational load (the number of non-silent mutations) of the tumor: grade II diffuse gliomas harbour fewer genetic changes than grade III or IV, even within defined molecular subtypes (e.g. ATRX mutated diffuse gliomas).

Conclusion: We have identified PI3K mutations as novel prognostic markers in gliomas. We also demonstrate that the mutational load is associated with tumor grade. The increase in mutational load may partially explain the increased aggressiveness of higher grade diffuse gliomas when a subset of the affected genes actively contributes to gliomagenesis and/or progression.

Fig. 1

PI3-kinase mutations are prognostic in molecular astrocytomas (diffuse gliomas with ATRX and/or TP53 mutations). a Data from TCGA samples (test cohort). Histology and grade of samples presented are listed in Additional file 1: Table S11; b Data from two validation cohorts (combined) from astrocytomas [17] and glioblastomas [6]. In both figures, only samples with an IDH mutation and TP53 or ATRX were selected. In these molecular astrocytomas, PI3 kinase mutations are prognostic for overall survival. P values indicated are calculated using the Log-rank test. Number of samples analysed are N = 144 (PIK3 wt) and N = 13 (PIK3 mutant) for the TCGA cohort and N = 22 (PIK3 wt) and N = 12 (PIK3 mutant) for the validation cohort

Fig. 2

Survival in prognostic molecular subtypes of diffuse glioma stratified by tumor grade. Different subtypes are indicated above each graph. As can be seen, within defined molecular subtypes, tumor grade remains a prognostic factor. Number of samples (grade II, III and IV) for each graph: 10, 42 and 248 (IDH-wt); 73, 65 and 13 (IDH-mut, ATRX/TP53 mut); 42, 32 and 0 (IDH-mut, 1p19q codeleted). 17 samples were IDH-mut but had neither ATRX/TP53 mutations nor 1p19q codeletion. P values indicated are calculated using the Log-rank test; other test for significance show for IDH-wt tumors a P = 0.0092 and for IDH-mut, ATRX/TP53 mut tumors a P = 0.0034 (both logrank test for trend); for IDH-mut, 1p19q codeleted) a P = 0.28 (Gehan Breslow Wilcoxon test). Log rank P values for grade II v. III are 0.43 and 0.063 for IDH-wt and IDH-mut, ATRX/TP53 mut tumors respectively

Fig. 3

Correlation between patient age and mutational load in diffuse gliomas. The number of non-silent genetic changes increases with patient age. This increase is irrespective of histological subtype (not shown) or tumor grade. Two samples (out of the 542 analyzed) with a high mutational load fall outside the y-axis limit in this figure: a grade III astrocytoma, age 47, mutational load 408 and a GBM, age 45, mutational load 181

Fig. 4

Proposed scheme for the prognostic classification of diffuse gliomas. Diffuse gliomas are first stratified based on their IDH-mutation status. Further classification is based on the ATRX and/or TP53 mutation status or determining 1p19q codeletion (these changes are mutually exclusive). Within the ATRX and/or TP53 mutated samples, mutations in PI3 kinase genes PIK3CA and PIK3R1 are associated with poor prognosis. It should be noted that there are genetic changes that associate with each molecular subtype (like EGFR amplification with IDH-wt tumors). They are however, not important for prognostic classification and may occur in several molecular subtypes. For example, PI3K mutations occur in all molecular subtypes but are only significantly prognostic in IDH-mutated, TP53/ATRX mutated diffuse gliomas). Additional prognostic factors include tumor grade and patient age, both of which are correlated to the mutational load of the tumor and are listed below the classification scheme. These additional markers are often correlated to the mutational profile of the tumors: Patients with IDH-wt tumors are often older and most are diagnosed as grade IV. ATRX/TP53 indicates mutation of either/both genes; 1p19q indicates codeletion of these chromosomal arms