Most gastrointestinal stromal tumors (GISTs) carry activating mutations of the KIT gene encoding the receptor tyrosine kinase KIT. In a previous study we were able to show an association between the lack of KIT mutations (wild-type GISTs) and the presence of a significant epithelioid tumor component. A very recent study described the occurrence of PDGFRalpha mutations in KIT wt GISTS. Therefore, we studied a panel of 87 GISTs for mutations in the hot spot regions of the PDGFRalpha gene with single strand conformation polymorphism analysis and sequencing and correlated the PDGFRalpha status with pathomorphological data. We detected 20 cases with exon 18 mutations but none with exon 12 mutations. The mutations were located in the second kinase domain of PDGFRalpha with 16 point mutations, and four larger deletions of 9 to 12 bp. All cases with mutations in the PDGFRalpha gene revealed wild-type KIT in common regions of mutation, ie, exons 9 and 11. Most interestingly, the occurrence of PDGFRalpha mutations was significantly associated with a higher frequency of epithelioid or mixed morphology (18 of 20 cases, P < 0.0001) and gastric location (all cases, P = 0.0008). Our data indicate that GISTs represent distinctive entities, differing in genetic, biological, and morphological features.

Hepatoblastomas are the most frequent malignant liver tumors of childhood. A high frequency of activating b-catenin mutations in hepatoblastomas indicates that the Wnt signaling pathway plays an important role in the development of this embryonic neoplasm. Stabilization of Bcatenin leads to an increased formation of nuclear B-catenin-T-cell factor complexes and altered expression of Wnt-inducible target genes. In this study, we analyzed the mRNA expression levels of nine Wnt genes, including c-JUN, c-MYC, CYCLIN D1, FRA-1, NKD-1, ITF-2, MMP-7, uPAR, and b-TRCP, by competitive reverse transcription-PCR. We analyzed 23 hepatoblastoma biopsies for which matching liver tissue was available, 5 hepatoblastoma cell lines, and 3 human fetal liver samples. b-TRCP and NKD-1 were highly expressed in all hepatoblastoma samples, independent of the b-catenin mutational status, in comparison with their nontumorous counterparts. b-TRCP mRNA overexpression was associated with accumulation of intracytoplasmic and nuclear B-TrCP protein. In human liver tumor cells without b-catenin mutations, Nkd-1 inhibited the Wnt-3a-activated Tcf-responsive-luciferase reporter activity, whereas Nkd-1 in hepatoblastomas with b-catenin mutations had no antagonistic effect. Our data emphasize the inhibitory effect of B-TrCP and Nkd-1 on the Wnt signaling pathway in a manner analogous to Conductin (AXIN2) and Dkk-1, inhibitors shown previously to be up-regulated in hepatoblastomas. Our findings indicate that overexpression of the Wnt antagonists Nkd-1 and B-TrCP reveals an activation of the Wnt signaling pathway as a common event in hepatoblastomas. We propose that Nkd-1 and B-TrCP may be used as possible diagnostic markers for the activated Wnt signaling pathway in hepatoblastomas.

Medulloblastomas (MBs) represent the most common malignant brain tumors in children. Most MBs develop sporadically in the cerebellum, but their incidence is highly elevated in patients with familial adenomatous polyposis coli. These patients carry germline mutations in the APC tumor suppressor gene. APC is part of a multiprotein complex involved in the Wnt signaling pathway that controls the stability of b-catenin, the central effector in this cascade. Previous genetic studies in MBs have identified mutations in genes coding for b-catenin and its partners, APC and AXIN1, which cause activation of Wnt signaling. The pathway is negatively controlled by the tumor suppressor AXIN2 (Conductin), a scaffold protein of this signaling complex. To investigate whether alterations in AXIN2 may also be involved in the pathogenesis of sporadic MBs, we performed a mutational screening of the AXIN2 gene in 116 MB biopsy samples and 11 MB cell lines using single-strand conformation polymorphism and sequencing analysis. One MB displayed a somatic, tumor-specific 2 bp insertion in exon 5, leading to carboxy-terminal truncation of the AXIN2 protein. This tumor biopsy showed nuclear accumulation of b-catenin protein, indicating an activation of Wnt signaling. In 2 further MB biopsies, mutations were identified in exon 5 (Glu408Lys) and exon 8 (Ser738Phe) of the AXIN2 gene, which are due to predicted germline mutations and rare polymorphisms. mRNA expression analysis in 22 MBs revealed reduced expression of AXIN2 mRNA compared to 8 fetal cerebellar tissues. Promoter hypermethylation could be ruled out as a major cause for transcriptional silencing by bisulfite sequencing. To study the functional role of AXIN2 in MBs, wild-type AXIN2 was overexpressed in MB cell lines in which the Wnt signaling pathway was activated by Wnt-3a. In this assay, AXIN2 inhibited Wnt signaling demonstrated in luciferase reporter assays. In contrast, overexpression of mutated AXIN2 with a deleted C-terminal DIX-domain resulted in an activation of the Wnt signaling pathway. These findings indicate that mutations of AXIN2 can lead to an oncogenic activation of the Wnt pathway in MBs. ' 2007 Wiley-Liss, Inc.Key words: medulloblastoma; Wnt; AXIN2; conductin Medulloblastomas (MBs) represent the most frequent malignant brain tumors of childhood with an incidence of 5 cases/million children.1 MBs are thought to arise from neoplastic transformation of cells in the external granule cell layer of the developing cerebellum.2 Although the majority of MBs occur as sporadic cases, familial tumor syndromes, including nevoid basal cell carcinoma syndrome (Gorlin syndrome) and Turcot syndrome, have been associated with a significantly elevated incidence of MBs.3,4 The wild-type APC allele in MBs has been reported to be lost in patients with Turcot syndrome, suggesting that APC functions as a tumor suppressor gene in the cerebellum. 5 Loss of APC function results in transcriptional activation of the Wnt signaling pathway, which plays major roles in tumorigenesis and in the r...

No abstract