An investigation of WNT pathway activation and association with survival in central nervous system primitive neuroectodermal tumours (CNS PNET)

Abstract

Central nervous system primitive neuroectodermal tumours (CNS PNET) are high-grade, predominantly paediatric, brain tumours. Previously they have been grouped with medulloblastomas owing to their histological similarities. The WNT/beta-catenin pathway has been implicated in many tumour types, including medulloblastoma. On pathway activation beta-catenin (CTNNB1) translocates to the nucleus, where it induces transcription of target genes. It is commonly upregulated in tumours by mutations in the key pathway components APC and CTNNB1. WNT/beta-catenin pathway status was investigated by immunohistochemical analysis of CTNNB1 and the pathway target cyclin D1 (CCND1) in 49 CNS PNETs and 46 medulloblastomas. The mutational status of APC and CTNNB1 (beta-catenin) was investigated in 33 CNS PNETs and 22 medulloblastomas. CTNNB1 nuclear localisation was seen in 36% of CNS PNETs and 27% of medulloblastomas. A significant correlation was found between CTNNB1 nuclear localisation and CCND1 levels. Mutations in CTNNB1 were identified in 4% of CNS PNETs and 20% of medulloblastomas. No mutations were identified in APC. A potential link between the level of nuclear staining and a better prognosis was identified in the CNS PNETs, suggesting that the extent of pathway activation is linked to outcome. The results suggest that the WNT/beta-catenin pathway plays an important role in the pathogenesis of CNS PNETs. However, activation is not caused by mutations in CTNNB1 or APC in the majority of CNS PNET cases.

Figure 1

Kaplan–Meier curves for analysis of the CNS PNET and medulloblastoma patient cohorts. A significant difference in overall survival was seen between CNS PNET patients under the age of 5 years at diagnosis and those over 5 years (P=0.045) (A). Patients who had a complete rather than partial resection had a better prognosis (overall survival, P=0.01) (B). CNS PNET patients treated with both chemotherapy (CT) and radiotherapy (RT) had a significantly better progression-free survival (P=0.01) (C). Medulloblastoma patients under 5 years at diagnosis also showed a significant association with survival (overall survival, P=0.006) (D). Medulloblastoma patients that had not relapsed (P<0.001) (E) or metastasised (overall survival, P=0.01) (F) had a significantly better prognosis. Medulloblastoma patients treated with either radiotherapy or chemotherapy and radiotherapy also had a better outcome (overall survival, P<0.001) (G). All survival times are in months.

Figure 2

Immunohistochemical analysis of CTNNB1, CCND1 and MKI67 in the CNS PNET and medulloblastoma cohorts. Two patterns of CTNNB1 nuclear staining were seen in both CNS PNETs and medulloblastomas. A low level of nuclear staining (less than 10%) was seen in some CNS PNETs (A) and medulloblastomas (B). In others a high level of nuclear staining (greater than 10%) was seen (C CNS PNET and D medulloblastoma). Over expression of CCND1 was also seen in a subset of tumours (E, CNS PNET). MKI67 levels were measured in both cohorts (F, CNS PNET). Levels did not correlate with CCND1. An additional CNS PNET sample containing a mutation in CTNNB1 exon 3 was analysed in a separate experiment and displayed high CTNNB1 nuclear staining (G).

Figure 3

Schematic representation of mutation locations in exon 3 of CTNNB1. Amino acid substitutions are indicated above the sequence; grey changes represent mutations from medulloblastoma and black from CNS PNET.

Figure 4

Kaplan–Meier curves for analysis of CTNNB1 IHC. Comparison of CNS PNETs displaying nuclear CTNNB1 with non-nuclear staining did not reveal a significant difference in overall survival (P=0.852) (A). Comparison of high CTNNB1 nuclear tumours (high nuclear) with the rest of the cohort (rest; tumours displaying low nuclear, cytoplasmic or negative staining), although not statistically significant (overall survival, P=0.113), suggested a trend towards a better prognosis for the high CTNNB1 nuclear group (B). A significant difference in overall survival was seen between CNS PNETs displaying high and low CTNNB1 nuclear staining (overall survival, P=0.007) (C). In the medulloblastoma cohort comparison of nuclear with non-nuclear CTNNB1 tumours was not significant (overall survival, P=0.590), but suggested a trend towards better survival for the nuclear group (D). Comparison of CTNNB1 high nuclear cases (high nuclear) with all other tumours in the medulloblastoma cohort (rest) also suggested the same trend (overall survival, P=0.310) (E). All survival times are in months.