[Show abstract][Hide abstract] ABSTRACT: Cerebellar development is regulated by a coordinated spatiotemporal interplay between granule neuron progenitors (GNPs), Purkinje neurons, and glia. Abnormal development can trigger motor deficits, and more recent data indicate important roles in aspects of memory, behavior, and autism spectrum disorders (ASDs). Germline mutation in the NF1 tumor suppressor gene underlies Neurofibromatosis type 1, a complex disease that enhances susceptibility to certain cancers and neurological disorders, including intellectual deficits and ASD. The NF1 gene encodes for neurofibromin, a RAS GTPase-activating protein, and thus negatively regulates the RAS signaling pathway. Here, using mouse models to direct conditional NF1 ablation in either embryonic cerebellar progenitors or neonatal GNPs, we show that neurofibromin is required for appropriate development of cerebellar folia layering and structure. Remarkably, neonatal administration of inhibitors of the ERK pathway reversed the morphological defects. Thus, our finding
[Show abstract][Hide abstract] ABSTRACT: In a recent Cell paper, Kitambi and colleagues identify a small molecule (Vacquinol-1) that has beneficial effects on a glioblastoma multiforme mouse model by oral administration. In glioblastoma cells, Vacquinol-1 targets macropinocytosis, a cellular process that will not lead to cell death in normal cells.
[Show abstract][Hide abstract] ABSTRACT: Neurofibromatosis Type 1 (NF1) is an autosomal disorder that affects neural crest-derived tissues, leading to a wide spectrum of clinical presentations. Patients commonly present with plexiform neurofibromas, benign but debilitating growths that can transform into malignant peripheral nerve sheath tumors (MPNSTs), a main cause of mortality. Currently, surgery is the primary course of treatment for MPNST, but with the limitation that these tumors are highly invasive. Radiation therapy is another treatment option, but is undesirable because it can induce additional mutations. MPNST patients may also receive doxorubicin as therapy, but this DNA-intercalating agent has relatively low tumor specificity and limited efficacy. In this study, we exploited a robust genetically-engineered mouse model of MPNST that recapitulates human NF1 associated MPNST to identify a novel small chemical compound that inhibits tumor cell growth. Compound 21 (Cpd21) inhibits growth of all available in vitro models of MPNST and human MPNST cell lines, while remaining non-toxic to normally-dividing Schwann cells or mouse embryonic fibroblasts. We show that this compound delays the cell cycle and leads to cellular apoptosis. Moreover, Cpd21 can reduce MPNST burden in a mouse allograft model, underscoring the compound's potential as a novel chemotherapeutic agent.
[Show abstract][Hide abstract] ABSTRACT: Schwann cells are highly plastic cells that dedifferentiate to a progenitor-like state following injury. However, deregulation of this plasticity, may be involved in the formation of neurofibromas, mixed-cell tumors of Schwann cell (SC) origin that arise upon loss of NF1. Here, we show that adult myelinating SCs (mSCs) are refractory to Nf1 loss. However, in the context of injury, Nf1-deficient cells display opposing behaviors along the wounded nerve; distal to the injury, Nf1(-/-) mSCs redifferentiate normally, whereas at the wound site Nf1(-/-) mSCs give rise to neurofibromas in both Nf1(+/+) and Nf1(+/-) backgrounds. Tracing experiments showed that distinct cell types within the tumor derive from Nf1-deficient SCs. This model of neurofibroma formation demonstrates that neurofibromas can originate from adult SCs and that the nerve environment can switch from tumor suppressive to tumor promoting at a site of injury. These findings have implications for both the characterization and treatment of neurofibromas.
[Show abstract][Hide abstract] ABSTRACT: Mitosis is controlled by a network of kinases and phosphatases. We screened a library of small interfering RNAs against a genome-wide set of phosphatases to comprehensively evaluate the role of human phosphatases in mitosis. We found four candidate spindle checkpoint phosphatases, including the tumor suppressor CDKN3. We show that CDKN3 is essential for normal mitosis and G1/S transition. We demonstrate that subcellular localization of CDKN3 changes throughout the cell cycle. We show that CDKN3 dephosphorylates threonine-161 of CDC2 during mitotic exit and we visualize CDC2(pThr-161) at kinetochores and centrosomes in early mitosis. We performed a phosphokinome-wide mass spectrometry screen to find effectors of the CDKN3-CDC2 signaling axis. We found that one of the identified downstream phosphotargets, CKβ phosphorylated at serine 209, localizes to mitotic centrosomes and controls the spindle checkpoint. Finally, we show that CDKN3 protein is down-regulated in brain tumors. Our findings indicate that CDKN3 controls mitosis through the CDC2 signaling axis. These results have implications for targeted anticancer therapeutics.
The Journal of Cell Biology 06/2013; · 10.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: PURPOSE: Glioblastoma multiforme (GBM) is the most lethal form of brain cancer with a median survival of only 12-15 months. Current standard treatment consists of surgery followed by chemoradiation. The poor survival of GBM patients is due to aggressive tumor invasiveness, an inability to remove all tumor tissue, and an innate tumor chemo- and radioresistance. ATM, ataxia telangiectasia (A-T) mutated, is an excellent target for radiosensitizing GBM because of its critical role in regulating the DNA damage response and p53, among other cellular processes. As a first step toward this goal, we recently showed that the novel ATM kinase inhibitor KU-60019 reduced migration, invasion, growth, and potently radiosensitized human glioma cells in vitro. EXPERIMENTAL DESIGN: Using orthotopic xenograft models of GBM, we now show that KU-60019 is also an effective radiosensitizer in vivo. Human glioma cells expressing reporter genes for monitoring tumor growth and dispersal were grown intra-cranially, and KU-60019 was administered intra-tumorally by convection-enhanced delivery or osmotic pump. RESULTS: Our results demonstrate that the combined effect of KU-60019 and radiation significantly increased survival of mice 2-3 fold over controls. Importantly, we show that glioma with mutant p53 is much more sensitive to KU-60019 radiosensitization than genetically matched wild-type glioma. CONCLUSIONS: Taken together, our results suggest that an ATM kinase inhibitor may be an effective radiosensitizer and adjuvant therapy for patients with mutant p53 brain cancers.
Clinical Cancer Research 04/2013; · 7.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVE: Loss of BDNF-TrkB signaling results in obesity in both humans and mice; however, the neural circuit that mediates this effect is unknown. We tested the role of TrkB signaling dopamine-1 receptor expressing neurons in body weight regulation. DESIGN AND METHODS: Mice with a floxed allele of the TrkB gene were paired with mice expressing Cre-recombinase under control of the D1 promoter in order to conditionally knock out expression of TrkB receptors from D1-neurons. RESULTS: Deletion of TrkB receptors from D1 neurons results in obesity in chow fed mice due to increased feed efficiency. In contrast, loss of Trk B signaling in D1 neurons induced hyperphagia and hyperglycemia in mice maintained on high fat diet. CONCLUSIONS: These findings indicate TrkB signaling in D1 neurons regulates body weight by distinct mechanisms for chow and high fat diet and may be important for defending the body against the development of obesity and obesity-related disorders.
[Show abstract][Hide abstract] ABSTRACT: TCR-mediated activation of the Ras signaling pathway is critical for T cell development in the thymus and function in the periphery. However, which members of a family of Ras GTPase-activating proteins (RasGAPs) negatively regulate Ras activation in T cells is unknown. In this study we examined a potential function for the neurofibromin 1 (NF1) RasGAP in the T cell lineage with the use of T cell-specific NF1-deficient mice. Surprisingly, on an MHC class I-restricted TCR transgenic background, NF1 was found to promote thymocyte positive selection. By contrast, NF1 neither promoted nor inhibited the negative selection of thymocytes. In the periphery, NF1 was found to be necessary for the maintenance of normal numbers of naïve CD4(+) and CD8(+) T cells but was dispensable as a regulator of TCR-induced Ras activation, cytokine synthesis, proliferation and differentiation and death. These findings point to a novel unexpected role for NF1 in T cell development as well as a regulator of T cell homeostasis.
[Show abstract][Hide abstract] ABSTRACT: The phosphoinositide signaling system is a crucial regulator of neural development, cell survival, and plasticity. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) negatively regulates phosphatidylinositol 3-kinase signaling and downstream targets. Nse-Cre Pten conditional knockout mice, in which Pten is ablated in granule cells of the dentate gyrus and pyramidal neurons of the hippocampal CA3, but not CA1, recapitulate many of the symptoms of humans with inactivating PTEN mutations, including progressive hypertrophy of the dentate gyrus and deficits in hippocampus-based social and cognitive behaviors. However, the impact of Pten loss on activity-dependent synaptic plasticity in this clinically relevant mouse model of Pten inactivation remains unclear. Here, we show that two phosphatidylinositol 3-kinase- and protein synthesis-dependent forms of synaptic plasticity, theta burst-induced long-term potentiation and metabotropic glutamate receptor (mGluR)-dependent long-term depression, are dysregulated at medial perforant path-to-dentate gyrus synapses of young Nse-Cre Pten conditional knockout mice before the onset of visible morphological abnormalities. In contrast, long-term potentiation and mGluR-dependent long-term depression are normal at CA3-CA1 pyramidal cell synapses at this age. Our results reveal that deletion of Pten in dentate granule cells dysregulates synaptic plasticity, a defect that may underlie abnormal social and cognitive behaviors observed in humans with Pten inactivating mutations and potentially other autism spectrum disorders.
Proceedings of the National Academy of Sciences 03/2013; · 9.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Malignant peripheral nerve sheath tumors (MPNSTs) are soft tissue sarcomas that arise in connective tissue surrounding peripheral nerves. They occur sporadically in a subset of patients with neurofibromatosis type 1 (NF1). MPNSTs are highly aggressive, therapeutically resistant, and typically fatal. Using comparative transcriptome analysis, we identified CXCR4, a G-protein-coupled receptor, as highly expressed in mouse models of NF1-deficient MPNSTs, but not in nontransformed precursor cells. The chemokine receptor CXCR4 and its ligand, CXCL12, promote MPNST growth by stimulating cyclin D1 expression and cell-cycle progression through PI3-kinase (PI3K) and β-catenin signaling. Suppression of CXCR4 activity either by shRNA or pharmacological inhibition decreases MPNST cell growth in culture and inhibits tumorigenesis in allografts and in spontaneous genetic mouse models of MPNST. We further demonstrate conservation of these activated molecular pathways in human MPNSTs. Our findings indicate a role for CXCR4 in NF1-associated MPNST development and identify a therapeutic target.
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: Plexiform neurofibromas are slow-growing chemoradiotherapy-resistant tumours arising in patients with neurofibromatosis type 1 (NF1). Currently, there are no viable therapeutic options for patients with plexiform neurofibromas that cannot be surgically removed because of their proximity to vital body structures. We undertook an open-label phase 2 trial to test whether treatment with imatinib mesylate can decrease the volume burden of clinically significant plexiform neurofibromas in patients with NF1. METHODS: Eligible patients had to be aged 3-65 years, and to have NF1 and a clinically significant plexiform neurofibroma. Patients were treated with daily oral imatinib mesylate at 220 mg/m(2) twice a day for children and 400 mg twice a day for adults for 6 months. The primary endpoint was a 20% or more reduction in plexiform size by sequential volumetric MRI imaging. Clinical data were analysed on an intention-to-treat basis; a secondary analysis was also done for those patients able to take imatinib mesylate for 6 months. This trial is registered with ClinicalTrials.gov, number NCT01673009. FINDINGS: Six of 36 patients (17%, 95% CI 6-33), enrolled on an intention-to-treat basis, had an objective response to imatinib mesylate, with a 20% or more decrease in tumour volume. Of the 23 patients who received imatinib mesylate for at least 6 months, six (26%, 95% CI 10-48) had a 20% or more decrease in volume of one or more plexiform tumours. The most common adverse events were skin rash (five patients) and oedema with weight gain (six). More serious adverse events included reversible grade 3 neutropenia (two), grade 4 hyperglycaemia (one), and grade 4 increases in aminotransferase concentrations (one). INTERPRETATION: Imatinib mesylate could be used to treat plexiform neurofibromas in patients with NF1. A multi-institutional clinical trial is warranted to confirm these results. FUNDING: Novartis Pharmaceuticals, the Indiana University Simon Cancer Centre, and the Indiana University Herman B Wells Center for Pediatric Research.
The Lancet Oncology 10/2012; · 25.12 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Neurofibromatosis type 1 (NF1) is the most common monogenic disorder in which individuals manifest CNS abnormalities. Affected individuals develop glial neoplasms (optic gliomas, malignant astrocytomas) and neuronal dysfunction (learning disabilities, attention deficits). Nf1 genetically engineered mouse models have revealed the molecular and cellular underpinnings of gliomagenesis, attention deficit, and learning problems with relevance to basic neurobiology. Using NF1 as a model system, these studies have revealed critical roles for the NF1 gene in non-neoplastic cells in the tumor microenvironment, the importance of brain region heterogeneity, novel mechanisms of glial growth regulation, the neurochemical bases for attention deficit and learning abnormalities, and new insights into neural stem cell function. Here we review recent studies, presented at a symposium at the 2012 Society for Neuroscience annual meeting, that highlight unexpected cell biology insights into RAS and cAMP pathway effects on neural progenitor signaling, neuronal function, and oligodendrocyte lineage differentiation.
Journal of Neuroscience 10/2012; 32(41):14087-93. · 6.91 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Neurotrophic factors, such as brain-derived neurotrophic factor (BDNF), are associated with the physiology of the striatum and the loss of its normal functioning under pathological conditions. The role of BDNF and its downstream signaling in regulating the development of the striatum has not been fully investigated, however. Here we report that ablation of Bdnf in both the cortex and substantia nigra depletes BDNF in the striatum, and leads to impaired striatal development, severe motor deficits, and postnatal lethality. Furthermore, striatal-specific ablation of TrkB, the gene encoding the high-affinity receptor for BDNF, is sufficient to elicit an array of striatal developmental abnormalities, including de-creased anatomical volume, smaller neuronal nucleus size, loss of dendritic spines, reduced enkephalin expression, diminished nigral dopaminergic projections, and severe deficits in striatal dopamine signaling through DARPP32. In addition, TrkB ablation in striatal neurons elicits a non–cell-autonomous reduction of tyrosine hy-droxylase protein level in the axonal projections of substantia nig-ral dopaminergic neurons. Thus, our results establish an essential function for TrkB in regulating the development of striatal neurons. Huntington disease | neurotrophin T he striatum is a subcortical part of the telencephalon and a major input component of the basal ganglia system. The striatum plays a crucial role in regulating voluntary movements, as well as reward-associated learning and memory. It can be separated into dorsal and ventral parts based on known ana-tomical and functional distinctions, with the dorsal portion as-sociated with movement regulation and the ventral striatum involved predominantly in mediating neurological functions pertaining to motivation, reward, and emotion. Progressive de-generation of the striatal medium spiny neurons (MSNs) and their main dopaminergic inputs from the substantia nigra are the cardinal symptoms of the neurological disorders Huntington disease (HD) and Parkinson disease, respectively. Like other telencephalic regions, the striatum contains in-hibitory neurons generated from the medial ganglionic eminence. The majority of inhibitory GABAergic neurons are MSNs that project outside of the striatum to neighboring structures, such as the globus pallidus and the substantia nigra. The mechanism reg-ulating development of the striatum involves combinatorial codes of transcription factors and parallels the development of other neural structures; however, the molecular mechanisms underlying the maturation and maintenance of the striatum remain largely unknown. Recent studies have shown that brain-derived neuro-trophic factor (BDNF) exerts prosurvival effects on striatal neu-rons in vitro (1, 2) and delays the onset of degeneration in a transgenic mouse model of HD (3). However, it has been shown that during development and in adulthood, Bdnf mRNA is largely absent from striatal neurons and glia. Instead, BDNF is produced in the cortex and substantia nigra and is transported anterogradely to striatal neurons (4). Indeed, cortical ablation of Bdnf leads to cellular deficits in the striatum (5) and global gene expression impairment (6). These findings suggest an important role for BDNF in the striatum; however, given BDNF's complex down-stream signaling capacities, the nature of its intracellular signaling cascades pertaining to development, maturation, and function of the striatum remains to be delineated. In this study, we conditionally ablated BDNF or its receptor TrkB in corticostriatal and nigrostriatal neuronal circuits. We found that Bdnf deletion in both cortex and substantia nigra led to complete depletion of BDNF protein in the striatum. Mutant mice displayed dramatic developmental abnormalities and neurological impairments. Furthermore, specific deletion of TrkB from striatal neurons was sufficient to produce this wide range of developmental deficits. Thus, our results demonstrate that BDNF and TrkB play critical paracrine and cell-autonomous roles, respectively, in the development and maintenance of striatal neurons.
Proceedings of the National Academy of Sciences 09/2012; 109(38):15491-15496. · 9.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Neurotrophic factors, such as brain-derived neurotrophic factor (BDNF), are associated with the physiology of the striatum and the loss of its normal functioning under pathological conditions. The role of BDNF and its downstream signaling in regulating the development of the striatum has not been fully investigated, however. Here we report that ablation of Bdnf in both the cortex and substantia nigra depletes BDNF in the striatum, and leads to impaired striatal development, severe motor deficits, and postnatal lethality. Furthermore, striatal-specific ablation of TrkB, the gene encoding the high-affinity receptor for BDNF, is sufficient to elicit an array of striatal developmental abnormalities, including decreased anatomical volume, smaller neuronal nucleus size, loss of dendritic spines, reduced enkephalin expression, diminished nigral dopaminergic projections, and severe deficits in striatal dopamine signaling through DARPP32. In addition, TrkB ablation in striatal neurons elicits a non-cell-autonomous reduction of tyrosine hydroxylase protein level in the axonal projections of substantia nigral dopaminergic neurons. Thus, our results establish an essential function for TrkB in regulating the development of striatal neurons.
Proceedings of the National Academy of Sciences 09/2012; 109(38):15491-6. · 9.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glioblastoma multiforme is the most common primary malignant brain tumour, with a median survival of about one year. This poor prognosis is due to therapeutic resistance and tumour recurrence after surgical removal. Precisely how recurrence occurs is unknown. Using a genetically engineered mouse model of glioma, here we identify a subset of endogenous tumour cells that are the source of new tumour cells after the drug temozolomide (TMZ) is administered to transiently arrest tumour growth. A nestin-ΔTK-IRES-GFP (Nes-ΔTK-GFP) transgene that labels quiescent subventricular zone adult neural stem cells also labels a subset of endogenous glioma tumour cells. On arrest of tumour cell proliferation with TMZ, pulse-chase experiments demonstrate a tumour re-growth cell hierarchy originating with the Nes-ΔTK-GFP transgene subpopulation. Ablation of the GFP+ cells with chronic ganciclovir administration significantly arrested tumour growth, and combined TMZ and ganciclovir treatment impeded tumour development. Thus, a relatively quiescent subset of endogenous glioma cells, with properties similar to those proposed for cancer stem cells, is responsible for sustaining long-term tumour growth through the production of transient populations of highly proliferative cells.