Integrated analysis of mismatch repair system in malignant astrocytomas

Abstract

Malignant astrocytomas are the most aggressive primary brain tumors with a poor prognosis despite optimal treatment. Dysfunction of mismatch repair (MMR) system accelerates the accumulation of mutations throughout the genome causing uncontrolled cell growth. The aim of this study was to characterize the MMR system defects that could be involved in malignant astrocytoma pathogenesis. We analyzed protein expression and promoter methylation of MLH1, MSH2 and MSH6 as well as microsatellite instability (MSI) and MMR gene mutations in a set of 96 low- and high-grade astrocytomas. Forty-one astrocytomas failed to express at least one MMR protein. Loss of MSH2 expression was more frequent in low-grade astrocytomas. Loss of MLH1 expression was associated with MLH1 promoter hypermethylation and MLH1-93G>A promoter polymorphism. However, MSI was not related with MMR protein expression and only 5% of tumors were MSI-High. Furthermore, the incidence of tumors carrying germline mutations in MMR genes was low and only one glioblastoma was associated with Lynch syndrome. Interestingly, survival analysis identified that tumors lacking MSH6 expression presented longer overall survival in high-grade astrocytoma patients treated only with radiotherapy while MSH6 expression did not modify the prognosis of those patients treated with both radiotherapy and chemotherapy. Our findings suggest that MMR system alterations are a frequent event in malignant astrocytomas and might help to define a subgroup of patients with different outcome.

Figure 1. Representative images of hematoxylin and eosin (H&E), MLH1, MSH2 and MSH6 staining on paraffin-embedded sections of samples from representatives low-grade astrocytomas (grade II) (A-D) and glioblastomas (grade IV) (E-H) according to the 2007 WHO classification [19].

Low-grade astrocytomas are well differentiated and slow-growing tumors with absence of necrosis and microvascular proliferation, whereas high-grade astrocytomas are characterized by high cellularity and mitotic activity, necrosis and microvascular proliferation (arrows). MLH1, MSH2 and MSH6 expression was visualized by staining with specific antibodies and their expression was considered positive when nuclear staining was detected in more than 50% of tumor cells (Magnification x400).

Figure 2. Kaplan-Meier estimates of overall survival in high-grade astrocytomas in the entire patient set according to treatment received.

Treatment with both radiotherapy and chemotherapy confers a significant increase in overall survival time (A). Survival analysis in each treatment group separately showed that loss of MSH6 expression correlated with a better overall survival in patients receiving radiation therapy alone (B), whereas MSH6 expression did not modify prognosis of patients receiving both radiotherapy and chemotherapy (C).