ORIGINAL PAPERS Synergy of Nf2 and p53 mutations in development of malignant tumours of neural crest origin Els Robanus-Maandag 1,5,6 , Marco Giovannini 2,6 , Martin van der Valk 3 , Michiko Niwa-Kawakita 2 , Vincent Abramowski 2 , Cristina Antonescu 4 , Gilles Thomas 2 and Anton Berns* ,1 1 Division of Molecular Genetics, Centre for Biomedical Genetics, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands; 2 INSERM U434, Fondation Jean Dausset-CEPH, 27 rue Juliette Dodu, 75010 Paris, France; 3 Department of Animal Pathology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands; 4 Department of Pathology, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA Previously, we have mimicked human neurofibromatosis type 2 (NF2) in conditional Nf2 mutant (P0Cre;Nf2 flox2/flox2 ) mice. Schwannomas, characteristic for NF2, were found at low frequency in older mice. Here, we report that these mice, upon additional hemizygosity for p53, rapidly develop multiple tumours showing features consistent with malig- nant peripheral nerve sheath tumours. Thus, p53 hemi- zygosity promotes tumorigenesis of mutant Nf2 peripheral nerve cells. In contrast, young P0Cre;Nf2 flox2/ þ ;p53 þ / cis mice mainly succumb to Nf2/p53-related osteogenic tumours. Therefore, Cre-mediated early biallelic loss of Nf2 function in neural crest-derived cells hemizygous for p53 results in resistance to osteogenic tumours and increased susceptibility to peripheral nerve sheath tumours. Oncogene (2004) 23, 6541–6547.doi:10.1038/sj.onc.1207858 Published online 28 June 2004 Keywords: MPNST; neural crest; P0 promoter; condi- tional knockout mice; compensatory signalling Introduction Vestibular schwannomas are benign neural sheath tumours of the vestibular branch of the eighth cranial nerve. They occur either as bilateral tumours, the hallmark of the human hereditary cancer syndrome neurofibromatosis type 2 (NF2), or as sporadic uni- lateraltumours.Lossormutationalinactivationofboth alleles of the NF2 gene in the Schwann cell is the rate- limitingstepinneoplastictransformation.NF2patients carryingonemutant NF2 allelearepredisposednotonly to bilateral vestibular schwannomas but also to schwannomas of other cranial, spinal and cutaneous nerves,tocranialandspinalmeningiomas,aswellasto presenile lens opacities and cerebral calcifications (McKusick, 1994). Loss of NF2 function is also frequently found in mesotheliomas (Bianchi et al., 1995). The product of the NF2 gene, called schwanno- min or merlin (Rouleau et al., 1993; Trofatter et al., 1993), has structural similarity with members of the ezrin-radixin-moesin(ERM)familyofproteinsthatlink the cytoskeleton and the cell membrane (for a review, see, Bretscher et al., 2000). Together with ezrin and moesin, schwannomin and the transmembrane receptor CD44formamolecularswitchthatspecifiescellgrowth arrest or proliferation (Morrison etal., 2001). The role of loss of Nf2 function in development andtumorigenesishasbeenstudiedinvariousknockout mousemodels.Embryoshomozygousfor Nf2 mutations causing either indirect skipping of exon 3 or 2–3 or direct deletion of exon 2 are lethal (McClatchey etal.,1998;Giovannini etal.,2000).Theyfailtoinitiate gastrulation between embryonic days 6.5 and 7.0 (McClatchey et al., 1997). Heterozygous Nf2 mutant micedonotmimichumanNF2butdeveloplaterinlife (10–30 months) mainly osteomas and osteosarcomas showing loss of heterozygosity (LOH) for Nf2 (McClatchey et al., 1998; Giovannini et al., 2000). In addition, indications for a role of Nf2 loss in metastatic potential have been found (McClatchey et al., 1998). Previously, we have mimicked NF2 in conditonal Nf2 mutant mice with P0 promoter-directed Cre recombinase activity in Schwann cells (Giovannini et al., 2000). P0Cre;Nf2 flox2/flox2 mice develop Schwann cell hyperplasia and later in life schwannomas due to Cre-mediated biallelic Nf2 exon 2 deletion, whereas these features are absent in P0Cre;Nf2 flox2/ þ mice. The schwannomas occur incidentally or at a low frequency depending on the P0Cre strain. These data suggest that the loss of the Nf2 tumour suppressor function in man and P0Cre;Nf2 flox2/flox2 mice is not observed in heterozygous Nf2 mutant mice possibly becauseofaninsufficientrateofspontaneouslossofthe Nf2 wild-type allele. P0Cre;Nf2 flox2/flox2 mice also show neurocristopathy due to P0 promoter activity in tissues with neural crest-derived components. These include osseous metaplasia and lens cataract, two non-neoplas- tic features of NF2, and the infrequently found osteogenic tumours. Received18January2004;revised26April2004;accepted26April2004; published online 28 June 2004 *Correspondence: A Berns; E-mail: a.berns@nki.nl 5 Current address: Center for Human and Clinical Genetics, Leiden UniversityMedicalCenter,Wassenaarseweg72,2333ALLeiden,The Netherlands 6 These authors contributed equally to this work Oncogene (2004) 23, 6541–6547 & 2004 Nature Publishing Group All rights reserved 0950-9232/04 $30.00 www.nature.com/onc