JCV T-antigen interacts with the neurofibromatosis type 2 gene product in a transgenic mouse model of malignant peripheral nerve sheath tumors

The human polyomavirus, JC virus, has recently been associated with several human CNS tumors, including medulloblastomas and a broad range of glial-origin tumors. This ubiquitous virus is the causative agent of the rare demyelinating disease, progressive multifocal leukoencephalopathy in immunocompromised individuals. Expression of the viral protein, T-antigen, which possesses the ability to transform cells of neural origin, has been detected in human CNS tumors. In an effort to further understand the transforming potential of JCV T-antigen, transgenic mice expressing JCV T-antigen under the control of the Mad-4 promoter were generated. As described previously, approximately 50% of the animals developed pituitary tumors by 1 year of age. However, a small subset of the animals developed solid masses arising from the soft tissues surrounding the salivary gland, the sciatic nerve, and along the extremities that histologically resemble malignant peripheral nerve sheath tumors, rare neoplasms that occur in individuals with neurofibromatosis type 1 (NF1). JCV T-antigen was detected in tumor tissue by immunohistochemistry and immunoprecipitation/Western blotting, but not in normal tissues and was colocalized with NF2, the putative tumor suppressor protein associated with neurofibromatosis type 2, in the nucleus of some cells. In addition, T-antigen was co-precipitated with NF2, but not with NF1 protein, although NF1 was detectable in tumor tissue. Furthermore, precipitated immunocomplexes contained T-antigen, NF2, and p53, suggesting that these three proteins may form a ternary complex. The importance of these findings on mechanisms of T-antigen-mediated tumorigenesis and the pathogenesis of neurofibromatosis are discussed.Oncogene (2004) 23, 5459-5467. doi:10.1038/sj.onc.1207728 Published online 10 May 2004     

Current status of sporadic and neurofibromatosis type 1-associated malignant peripheral nerve sheath tumors

Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive soft tissue sarcomas that rarely occur in the general population but have a lifetime incidence of 8% to 13% in those with neurofibromatosis type 1 (NF1). Complete surgical resection is the standard treatment for MPNSTs. Unresectable MPNSTs carry a poor prognosis, and survival appears to be worse in NF1-associated tumors than in sporadic tumors. The response rate of MPNSTs to standard chemotherapeutic agents used to treat pediatric and adult soft tissue sarcomas is unknown and is currently undergoing evaluation in a multi-institutional clinical trial. With an increasing understanding of the molecular pathogenesis of MPNSTs, clinical trials with targeted agents have become available and have established that histology-specific trials in this rare malignancy are feasible. This knowledge, coupled with the availability of preclinical MPNST models, likely will accelerate the development of effective treatments for this malignancy.     

MicroRNA‐10b regulates tumorigenesis in neurofibromatosis type 1

MicroRNAs (miRNAs) are frequently deregulated in human tumors, and play important roles in tumor development and progression. The pathological roles of miRNAs in neurofibromatosis type 1 (NF1) tumorigenesis are largely unknown. We demonstrated that miR‐10b was up‐regulated in primary Schwann cells isolated from NF1 neurofibromas and in cell lines and tumor tissues from malignant peripheral nerve sheath tumors (MPNSTs). Intriguingly, a significantly high level of miR‐10b correlated with low neurofibromin expression was found in a neuroectodermal cell line: Ewing’s sarcoma SK‐ES‐1 cells. Antisense inhibiting miR‐10b in NF1 MPNST cells reduced cell proliferation, migration and invasion. Furthermore, we showed that NF1 mRNA was the target for miR‐10b. Overexpression of miR‐10b in 293T cells suppressed neurofibromin expression and activated RAS signaling. Antisense inhibition of miR‐10b restored neurofibromin expression in SK‐ES‐1 cells, and decreased RAS signaling independent of neurofibromin in NF1 MPNST cells. These results suggest that miR‐10b may play an important role in NF1 tumorigenesis through targeting neurofibromin and RAS signaling. (Cancer Sci 2010)     

Lack ofNF1 expression in a sporadic schwannoma from a patient without neurofibromatosis

Summary The neurofibromatosis type 1 (NF1) gene encodes a tumor suppressor protein, neurofibromin, which is expressed at high levels in Schwann cells and other adult tissues. Loss ofNF1 expression has been reported in Schwann cell tumors (neurofibrosarcomas) from patients with NF1 and its loss is associated with increased proliferation of these cells. In this report, we describe downregulation ofNF1 expression in a single spinal schwannoma from an individual without clinical features of neurofibromatosis type 1 or 2. Barely detectable expression ofNF1 RNA was found in this tumor byin situ hybridization using anNF1-specific riboprobe as well as by Northern blot and reverse-transcribed (RT)-PCR analysis. In Schwann cells cultured from this schwannoma, abundant expression ofNF1 RNA could be detected by Northern blot and RT-PCR analysis. These results suggest that, in some tumors, expression ofNF1 may be downregulated by factors produced within the tumor and may represent a novel mechanism for inactivating these growth suppressing genes and allowing for increased cell proliferation in tumors.     

NF2/Merlin in hereditary neurofibromatosis 2 versus cancer: biologic mechanisms and clinical associations

Inactivating germline mutations in the tumor suppressor gene NF2 cause the hereditary syndrome neurofibromatosis 2, which is characterized by the development of neoplasms of the nervous system, most notably bilateral vestibular schwannoma. Somatic NF2 mutations have also been reported in a variety of cancers, but interestingly these mutations do not cause the same tumors that are common in hereditary neurofibromatosis 2, even though the same gene is involved and there is overlap in the site of mutations. This review highlights cancers in which somatic NF2 mutations have been found, the cell signaling pathways involving NF2/merlin, current clinical trials treating neurofibromatosis 2 patients, and preclinical findings that promise to lead to new targeted therapies for both cancers harboring NF2 mutations and neurofibromatosis 2 patients.     

Optic nerve glioma in mice requires astrocyte Nf1 gene inactivation and Nf1 brain heterozygosity

Whereas biallelic neurofibromatosis 1 (NF1) inactivation is observed in NF1-associated gliomas, astrocyte-restricted Nf1 conditional knockout mice do not develop gliomas. These observations suggest that NF1 glioma formation requires additional cellular or genetic conditions. To determine the effect of an Nf1 heterozygous brain environment on NF1 glioma formation, we generated Nf1+/- mice lacking Nf1 expression in astrocytes. In contrast to astrocyte-restricted Nf1 conditional knockout mice, Nf1+/- mice lacking Nf1 in astrocytes develop optic nerve gliomas. This mouse model demonstrates that Nf1+/- cells contribute to the pathogenesis of gliomas in NF1 and provides a tool for the preclinical evaluation of potential therapeutic interventions for these tumors.     

Comparative pathology of nerve sheath tumors in mouse models and humans

Despite the progress made in our understanding of the biology of neurofibromatosis (NF), the long-term clinical outcome for affected patients has not changed significantly in the past decades, and both NF1 and NF2 are still associated with a significant morbidity and a decreased life span. A number of NF1 and NF2 murine models have been generated to aid in the study of NF tumor biology and in the development of targeted therapies for NF patients. A single, universal pathological classification of the lesions generated in these murine models is essential for the validation of the models, for their analysis and comparison with other models, and for their future effective use in preclinical treatment trials. For the formulation of a pathological classification of these lesions, the WHO classification of human tumors was used as a reference. However, it was not adopted for the classification of the GEM lesions because of some important differences between the human and murine lesions. A novel classification scheme for peripheral nerve sheath tumors in murine models was therefore devised.     

NF2 expression levels of gastrointestinal stromal tumors: a quantitative real-time PCR study

Gastrointestinal stromal tumors are the most common mesenchymal tumors of the gastrointestinal tract. Until today, there have been few markers specific for the tumor. This has complicated the differential diagnosis of the neoplasm from tumors of smooth muscle origin. Recently, the proto-oncogene c-kit has been shown to be a very relevant marker as it almost invariably is expressed in gastrointestinal stromal tumors. Radiation exposure, hormonal and genetic factors, particularly neurofibromatosis 2, have been implicated in their development and growth. GIST initiation, either in NF2-associated or in sporadic cases, is linked to inactivation of members of the proteins 4.1 superfamily. The majority of the mutations identified in the NF2 gene result in a truncated protein and are clinically associated with a severe phenotype. Occasionally, missense mutations associated with a mild phenotype may occur. We compared NF2 gene expression in 5 cases with gastrointestinal stromal tumors by quantitative real-time polymerase chain reaction analysis. NF2 gene mRNA expression was assessed in fresh tissue of stomach from 5 consecutive patients. We detected no alterations in NF2 gene expression in the quantitative analyses of the 5 tumors.     

Cutaneous manifestations in neuro-oncology: clinically relevant tumor and treatment associated dermatologic findings

Skin findings are a rare but important aspect of the evaluation and management of patients with tumors of the nervous system. Skin findings have the highest prevalence in genetic tumor syndromes termed neuro-genodermatoses, which include neurofibromatosis type 1 (NF1), neurofibromatosis type 2 (NF2), and tuberous sclerosis. Skin changes are observed in patients with non-syndromic nervous system malignancy, often as a result of pharmacotherapy. The skin may also manifest findings in paraneoplastic conditions that affect the nervous system, providing an early indication of underlying neoplasm, including dermatomyosistis, neuropathic itch, and brachioradial pruritus. In this article, we review the major cutaneous findings in patients with tumors of the brain, spine, and peripheral nervous system focusing on (1) cutaneous manifestations of genetic and sporadic primary nervous system tumor syndromes, and (2) paraneoplastic neurological syndromes with prominent cutaneous features.     

Improving outcomes for neurofibromatosis 1–associated brain tumors

Children and adults with neurofibromatosis type 1 (NF1) are predisposed to developing CNS tumors, including optic pathway gliomas (OPGs), brainstem gliomas (BSGs) and high-grade gliomas. Although current first-line treatments for low-grade gliomas (OPGs and BSGs) may prevent further tumor growth, they rarely result in restoration of the associated visual or neurological deficits. The availability of accurate small-animal models of NF1-associated brain tumors has established tractable experimental platforms for the discovery and evaluation of promising therapeutic agents. On the basis of these preclinical studies, biologically targeted agents are now being evaluated in children with NF1-associated low-grade brain tumors. Collectively, these models have also begun to reveal potential neuroprotective and risk assessment strategies for this brain tumor-prone population.     

Loss of heterozygosity on chromosome 22 in ovarian carcinoma is distal to and is not accompanied by mutations in NF2 at 22q12.

Frequent loss of heterozygosity (LOH) has been reported on 22q in ovarian carcinoma, implying the presence of a tumour-suppressor gene. The neurofibromatosis type 2 gene (NF2) at 22q12 is a plausible candidate. Analysis of 9 of the 17 exons of NF2 by single-strand conformational polymorphism (SSCP) in 67 ovarian carcinomas did not detect any somatic mutations, suggesting that NF2 is not involved in the pathogenesis of ovarian carcinoma. LOH data support this conclusion and that the putative tumour-suppressor gene lies distal to NF2, beyond D22S283.     

Monosomy 7 Myeloproliferative Disease Associated with Neurofibromatosis Type I: a Case Report

Abstract

A 7-year-old girl with neurofibromatosis type I (NF1) was diagnosed to have monosomy 7 myeloproliferative disease (Mo 7-MPD). Of the benign and malignant tumors that are encountered with increased incidence in NF1, those originating from the neural crest are frequent. However, tumors that do not originate from the neural crest may also be seen and among these, myeloid leukemias are prominent. Studies on NF1 patients with Mo 7-MPD and juvenile chronic myeloid leukemia (JCML) have suggested the role of the NF1 gene in the leukemogenesis. The relationship between monosomy 7 and hematological malignancies is already known. These findings are in agreement with the multistep development theory of cancer. In addition, our case is one of the very rare NF1 cases having father to daughter inheritance with a myeloid malignancy. We believe that cytogenetic and molecular genetic studies will contribute to further understanding of leukemogenesis.     

The angiogenic factor midkine is aberrantly expressed in NF1-deficient Schwann cells and is a mitogen for neurofibroma-derived cells

Loss of the tumor suppressor gene NF1 in neurofibromatosis type 1 (NF1) contributes to the development of a variety of tumors, including malignant peripheral nerve sheath tumors (MPNST) and benign neurofibromas. Of the different cell types found in neurofibromas, Schwann cells usually provide between 40 and 80%, and are thought to be critical for tumor growth. Here we describe the identification of growth factors that are upregulated in NF1-/- mouse Schwann cells and are potential regulators of angiogenesis and cell growth. Basic fibroblast growth factor (FGF-2), platelet-derived growth factor (PDGF) and midkine (MK) were found to be induced by loss of neurofibromin and MK was further characterized. MK was induced in human neurofibromas, schwannomas, and various nervous system tumors associated with NF1 or NF2; midkine showed an expression pattern overlapping but distinct from its homolog pleiotrophin (PTN). Immunohistochemistry revealed expression of MK in S-100 positive Schwann cells of dermal and plexiform neurofibromas, and in endothelial cells of tumor blood vessels, but not in normal blood vessels. Furthermore, MK demonstrated potent mitogenic activity for human systemic and brain endothelial cells in vitro and stimulated proliferation and soft agar colony formation of human MPNST derived S100 positive cells and fibroblastoid cells derived from an NF1 neurofibroma. The data support a possible central role for MK as a mediator of angiogenesis and neurofibroma growth in NF1. Oncogene (2001) 20, 97 - 105.     

Analysis of cytogenetic aberrations in sporadic vestibular schwannoma by comparative genomic hybridization

Vestibular schwannomas (VS) are benign tumors of the nervous system that are usually sporadic but also occur in the inherited disorder neurofibromatosis type 2 (NF2). The NF2 gene is a tumor suppressor gene located on chromosome 22. Loss of the NF2 protein product, Merlin, is universal in both sporadic and NF2-related schwannomas and the loss or mutation of the gene is the only established causative event underlying schwannoma formation. Comparative genomic hybridization (CGH) was used to screen 20 sporadic VS to identify additional chromosomal regions that may harbor genes involved in VS-tumorigenesis. The most common change were losses on chromosome 22q. Additionally, losses were observed on chromosome 9p indicating a possible participation of the CDKN2A tumor suppressor gene in the genesis of VS. Gains were observed on 17q, 19p and 19q, which have been reported before in malignant peripheral nerve sheath tumors that are associated with neurofibromatosis type 1. Importantly, high level amplifications have been observed on 16p and 16q as well as on 9q, suggesting the possible involvement of several oncogenes in the tumorigenesis of VS. Our data suggest the involvement of various oncogenes and tumor suppressor genes might play a role in the genesis of the vestibular schwannomas apart from the inactivation of the NF2 gene.     

Susceptible stages in Schwann cells for NF1-associated plexiform neurofibroma development

Stem cells are under strict regulation by both intrinsic factors and the microenvironment. There is increasing evidence that many cancers initiate through acquisition of genetic mutations (loss of intrinsic control) in stem cells or their progenitors, followed by alterations of the surrounding microenvironment (loss of extrinsic control). In neurofibromatosis type 1 (NF1), deregulation of Ras signaling results in development of multiple neurofibromas, complex tumors of the peripheral nerves. Neurofibromas arise from the Schwann cell lineage following loss of function at the NF1 locus, which initiates a cascade of interactions with other cell types in the microenvironment and additional cell autonomous modifications. In this study, we sought to identify whether a temporal "window of opportunity" exists during which cells of the Schwann cell lineage can give rise to neurofibromas following loss of NF1. We showed that acute loss of NF1 in both embryonic and adult Schwann cells can lead to neurofibroma formation. However, the embryonic period when Schwann cell precursors and immature Schwann cells are most abundant coincides with enhanced susceptibility to plexiform neurofibroma tumorigenesis. This model has important implications for understanding early cellular events that dictate neurofibroma development, as well as for the development of novel therapies targeting these tumors.     

Cerebrospinal fluid proteomic analysis reveals dysregulation of methionine aminopeptidase-2 expression in human and mouse neurofibromatosis 1-associated glioma

Individuals affected with the neurofibromatosis 1 (NF1) tumor predisposition syndrome are prone to the development of multiple nervous system tumors, including optic pathway gliomas (OPG). The NF1 tumor suppressor gene product, neurofibromin, functions as a Ras GTPase-activating protein, and has been proposed to regulate cell growth by inhibiting Ras activity. Recent studies from our laboratory have shown that neurofibromin also regulates the mammalian target of rapamycin activity in a Ras-dependent fashion, and that the rapamycin-mediated mammalian target of rapamycin inhibition ameliorates the Nf1-/- astrocyte growth advantage. Moreover, Nf1-deficient astrocytes exhibit increased protein translation. As part of a larger effort to identify protein markers for NF1-associated astrocytomas that could be exploited for therapeutic drug design, we did an objective proteomic analysis of the cerebrospinal fluid from genetically engineered Nf1 mice with optic glioma. One of the proteins found to be increased in the cerebrospinal fluid of OPG-bearing mice was the eukaryotic initiation factor-2alpha binding protein, methionine aminopeptidase 2 (MetAP2). In this study, we show that Nf1 mouse OPGs and NF1-associated human astrocytic tumors, but not sporadic pilocytic or other low-grade astrocytomas, specifically expressed high levels of MetAP2. In addition, we show that Nf1-deficient astrocytes overexpress MetAP2 in vitro and in vivo, and that treatment with the MetAP2 inhibitor fumagillin significantly reduces Nf1-/- astrocyte proliferation in vitro. These observations suggest that MetAP2 is regulated by neurofibromin, and that MetAP2 inhibitors could be potentially employed to treat NF1-associated tumor proliferation.     

24例神经纤维瘤病临床分析

目的:探讨神经纤维瘤病(neurofibromatosis,NF)的临床表现、诊断及治疗方法.方法:收集24例神经纤维瘤病患者的临床资料.回顾性分析该病的临床表现、诊断、治疗方法等.结果:24例患者中23例为Ⅰ型神经纤维瘤病(NF Ⅰ型),均否认家族遗传史.1例为Ⅱ型神经纤维瘤病(NFⅡ型),有家族史.男性14例,女性10例,发病年龄出生至64岁,平均年龄24.7岁.首次就诊20例,复发或再次就诊4例.结论:神经纤维瘤病临床表现多样,对于肿瘤在体表的患者,易于诊断;而肿瘤位于内脏的患者,如何做出准确的诊断,是一个值得探讨的问题.在治疗上,手术达不到根治的目的,术后易复发,需再次手术切除.     

Proteomic analysis reveals hyperactivation of the mammalian target of rapamycin pathway in neurofibromatosis 1-associated human and mouse brain tumors

Individuals with the tumor predisposition syndrome, neurofibromatosis 1 (NF1), are prone to development of nervous system tumors, including neurofibromas and pilocytic astrocytomas. Based on the ability of the NF1 gene product (neurofibromin) to function as a GTPase activating protein for RAS, initial biologically based therapies for NF1-associated tumors focused on the use of RAS inhibitors, but with limited clinical success. In an effort to identify additional targets for therapeutic drug design in NF1, we used an unbiased proteomic approach to uncover unanticipated intracellular signaling pathways dysregulated in Nf1-deficient astrocytes. We found that the expression of proteins involved in promoting ribosome biogenesis was increased in the absence of neurofibromin. In addition, Nf1-deficient astrocytes exhibit high levels of mammalian target of rapamycin (mTOR) pathway activation, which was inhibited by blocking K-RAS or phosphatidylinositol 3-kinase activation. This mTOR pathway hyperactivation was reflected by high levels of ribosomal S6 activation in both Nf1 mutant mouse optic nerve gliomas and in human NF1-associated pilocytic astrocytoma tumors. Moreover, inhibition of mTOR signaling in Nf1-/- astrocytes abrogated their growth advantage in culture, restoring normal proliferative rates. These results suggest that mTOR pathway inhibition may represent a logical and tractable biologically based therapy for brain tumors in NF1.     

Tumor load rather than contrast enhancement is associated with the visual function of children and adolescents with optic pathway glioma – a retrospective Magnetic Resonance Imaging study

Journal of Neuro-Oncology - Optic pathway gliomas are often asymptomatic tumors occurring in children with neurofibromatosis type 1 (NF1?+?OPG) or sporadically (spOPG). Treatment is...     

Conditional N-rasG12V expression promotes manifestations of neurofibromatosis in a mouse model

Human clinical neurofibromatosis type 1 (NF1) and type 2 (NF2) result from mutations and inactivation of neurofibromin and merlin genes, respectively, which negatively regulate Ras pathways. To evaluate the contribution of N-Ras activity to the development of NF, we generated a novel transgenic mouse expressing oncogenic N-ras specifically in central nerve cells, neural crest-derived cells and lens epithelial cells. Soon after birth, the mouse skin showed hyperpigmentation of the epidermis and melanin-laden macrophages in the dermis, as observed in the café-au-lait spots of human cases. At 3 months of age, all the mice had neurofibromas in the skin and neurofibroma-like tumors with structure similar to Wagner-Meissner bodies in the adrenal medulla. At 4 months of age, all the mice developed subcapsular cataract. In the 5th month, some developed protruding dermal neurofibromas involving subcutaneous fat. However, plexiform neurofibroma, schwannoma, astrocytoma and pheochromocytoma were not observed in the mice, suggesting a requirement for signal(s) other than the activated N-Ras pathway to induce these tumors. Thus, the activated N-Ras signal may be a main pathway for the development of the disease phenotypes characteristic of NF.