Análise da Frequência de Polimorfismos nos genes IL28B e IL28R1 em pacientes acometidos por Meningioma

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Papiloma de plexo coroide em um cão

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Características tomográficas de neoplasias encefálicas primárias em cães

Computed tomography (CT) can be considered an important test to identify the presence of primary brain neoplasias in dogs. CT results can help define the type of brain tumor when associated with clinical findings. It allows the identification of lesions and their features, such as size, location, compression and invasion of adjacent tissue. One must analyze the density, mass effect, peritumoral edema, calcification, and image enhancement after intravenous injection of contrast medium. Gliomas, meningiomas and tumors of the choroid plexus and pituitary are the most common primary brain neoplasms diagnosed by CT in dogs. The aim of this paper is to review the literature related to primary brain tumors and report their most important tomographic features, in order to help clinicians achieve a presumptive diagnosis of tumor type.

Cytogenetic findings in pediatric radiation-induced atypical meningioma after treatment of medulloblastoma: case report and review of the literature

Ionizing radiation is the most recognized risk factor for meningioma in pediatric long-term cancer survivors. Information in this rare setting is exceptional. We report the clinical and cytogenetic findings in a radiation-induced atypical meningioma following treatment for desmoplastic medulloblastoma in a child. This is the second study to describe the cytogenetic aspects on radiation-induced meningiomas in children. Chromosome banding analysis revealed a 46, XX, t(1;3)(p22;q12), del(1)(p?)[8]/46, XX[12]. Loss of chromosome 1p as a consequence of irradiation has been proposed to be more important in the development of secondary meningiomas in adults. Deletions in the short arm of chromosome 1 also appear to be a shared feature in both pediatric cases so far analyzed.

Biochemistry in healthy and neoplastic human tissues: metabolic alteration revealed by HR-MAS nuclear magnetic resonance spectroscopy

This thesis is focused on the metabolomic study of human cancer tissues by ex vivo High Resolution-Magic Angle Spinning (HR-MAS) nuclear magnetic resonance (NMR) spectroscopy. This new technique allows for the acquisition of spectra directly on intact tissues (biopsy or surgery), and it has become very important for integrated metabonomics studies. The objective is to identify metabolites that can be used as markers for the discrimination of the different types of cancer, for the grading, and for the assessment of the evolution of the tumour. Furthermore, an attempt to recognize metabolites, that although involved in the metabolism of tumoral tissues in low concentration, can be important modulators of neoplastic proliferation, was performed. In addition, NMR data was integrated with statistical techniques in order to obtain semi-quantitative information about the metabolite markers. In the case of gliomas, the NMR study was correlated with gene expression of neoplastic tissues. Chapter 1 begins with a general description of a new “omics” study, the metabolomics. The study of metabolism can contribute significantly to biomedical research and, ultimately, to clinical medical practice. This rapidly developing discipline involves the study of the metabolome: the total repertoire of small molecules present in cells, tissues, organs, and biological fluids. Metabolomic approaches are becoming increasingly popular in disease diagnosis and will play an important role on improving our understanding of cancer mechanism. Chapter 2 addresses in more detail the basis of NMR Spectroscopy, presenting the new HR-MAS NMR tool, that is gaining importance in the examination of tumour tissues, and in the assessment of tumour grade. Some advanced chemometric methods were used in an attempt to enhance the interpretation and quantitative information of the HR-MAS NMR data are and presented in chapter 3. Chemometric methods seem to have a high potential in the study of human diseases, as it permits the extraction of new and relevant information from spectroscopic data, allowing a better interpretation of the results. Chapter 4 reports results obtained from HR-MAS NMR analyses performed on different brain tumours: medulloblastoma, meningioms and gliomas. The medulloblastoma study is a case report of primitive neuroectodermal tumor (PNET) localised in the cerebellar region by Magnetic Resonance Imaging (MRI) in a 3-year-old child. In vivo single voxel 1H MRS shows high specificity in detecting the main metabolic alterations in the primitive cerebellar lesion; which consist of very high amounts of the choline-containing compounds and of very low levels of creatine derivatives and N-acetylaspartate. Ex vivo HR-MAS NMR, performed at 9.4 Tesla on the neoplastic specimen collected during surgery, allows the unambiguous identification of several metabolites giving a more in-depth evaluation of the metabolic pattern of the lesion. The ex vivo HR-MAS NMR spectra show higher detail than that obtained in vivo. In addition, the spectroscopic data appear to correlate with some morphological features of the medulloblastoma. The present study shows that ex vivo HR-MAS 1H NMR is able to strongly improve the clinical possibility of in vivo MRS and can be used in conjunction with in vivo spectroscopy for clinical purposes. Three histological subtypes of meningiomas (meningothelial, fibrous and oncocytic) were analysed both by in vivo and ex vivo MRS experiments. The ex vivo HR-MAS investigations are very helpful for the assignment of the in vivo resonances of human meningiomas and for the validation of the quantification procedure of in vivo MR spectra. By using one- and two dimensional experiments, several metabolites in different histological subtypes of meningiomas, were identified. The spectroscopic data confirmed the presence of the typical metabolites of these benign neoplasms and, at the same time, that meningomas with different morphological characteristics have different metabolic profiles, particularly regarding macromolecules and lipids. The profile of total choline metabolites (tCho) and the expression of the Kennedy pathway genes in biopsies of human gliomas were also investigated using HR-MAS NMR, and microfluidic genomic cards. 1H HR-MAS spectra, allowed the resolution and relative quantification by LCModel of the resonances from choline (Cho), phosphorylcholine (PC) and glycerolphorylcholine (GPC), the three main components of the combined tCho peak observed in gliomas by in vivo 1H MRS spectroscopy. All glioma biopsies depicted an increase in tCho as calculated from the addition of Cho, PC and GPC HR-MAS resonances. However, the increase was constantly derived from augmented GPC in low grade NMR gliomas or increased PC content in the high grade gliomas, respectively. This circumstance allowed the unambiguous discrimination of high and low grade gliomas by 1H HR-MAS, which could not be achieved by calculating the tCho/Cr ratio commonly used by in vivo 1H MR spectroscopy. The expression of the genes involved in choline metabolism was investigated in the same biopsies. The present findings offer a convenient procedure to classify accurately glioma grade using 1H HR-MAS, providing in addition the genetic background for the alterations of choline metabolism observed in high and low gliomas grade. Chapter 5 reports the study on human gastrointestinal tract (stomach and colon) neoplasms. The human healthy gastric mucosa, and the characteristics of the biochemical profile of human gastric adenocarcinoma in comparison with that of healthy gastric mucosa were analyzed using ex vivo HR-MAS NMR. Healthy human mucosa is mainly characterized by the presence of small metabolites (more than 50 identified) and macromolecules. The adenocarcinoma spectra were dominated by the presence of signals due to triglycerides, that are usually very low in healthy gastric mucosa. The use of spin-echo experiments enable us to detect some metabolites in the unhealthy tissues and to determine their variation with respect to the healthy ones. Then, the ex vivo HR-MAS NMR analysis was applied to human gastric tissue, to obtain information on the molecular steps involved in the gastric carcinogenesis. A microscopic investigation was also carried out in order to identify and locate the lipids in the cellular and extra-cellular environments. Correlation of the morphological changes detected by transmission (TEM) and scanning (SEM) electron microscopy, with the metabolic profile of gastric mucosa in healthy, gastric atrophy autoimmune diseases (AAG), Helicobacter pylori-related gastritis and adenocarcinoma subjects, were obtained. These ultrastructural studies of AAG and gastric adenocarcinoma revealed lipid intra- and extra-cellularly accumulation associated with a severe prenecrotic hypoxia and mitochondrial degeneration. A deep insight into the metabolic profile of human healthy and neoplastic colon tissues was gained using ex vivo HR-MAS NMR spectroscopy in combination with multivariate methods: Principal Component Analysis (PCA) and Partial Least Squares Discriminant Analysis (PLS-DA). The NMR spectra of healthy tissues highlight different metabolic profiles with respect to those of neoplastic and microscopically normal colon specimens (these last obtained at least 15 cm far from the adenocarcinoma). Furthermore, metabolic variations are detected not only for neoplastic tissues with different histological diagnosis, but also for those classified identical by histological analysis. These findings suggest that the same subclass of colon carcinoma is characterized, at a certain degree, by metabolic heterogeneity. The statistical multivariate approach applied to the NMR data is crucial in order to find metabolic markers of the neoplastic state of colon tissues, and to correctly classify the samples. Significant different levels of choline containing compounds, taurine and myoinositol, were observed. Chapter 6 deals with the metabolic profile of normal and tumoral renal human tissues obtained by ex vivo HR-MAS NMR. The spectra of human normal cortex and medulla show the presence of differently distributed osmolytes as markers of physiological renal condition. The marked decrease or disappearance of these metabolites and the high lipid content (triglycerides and cholesteryl esters) is typical of clear cell renal carcinoma (RCC), while papillary RCC is characterized by the absence of lipids and very high amounts of taurine. This research is a contribution to the biochemical classification of renal neoplastic pathologies, especially for RCCs, which can be evaluated by in vivo MRS for clinical purposes. Moreover, these data help to gain a better knowledge of the molecular processes envolved in the onset of renal carcinogenesis.

Intracranial neoplasia in 61 cats: localisation, tumour types and seizure patterns

The purpose of this study was to analyse retrospectively a feline population with intracranial neoplastic diseases, to document seizure patterns in these animals and to determine whether partial seizures were more frequently associated with structural brain lesions then generalised seizures. In addition, a comparison was made within the population with intracranial neoplasia between two groups of cats: one with and one without seizures. Special emphasis was given to the evaluation of tumour type, localisation and size of the lesion and its correlation with seizure prevalence. Sixty-one cats with histopathological diagnosis of intracranial tumour were identified. Fourteen cats (23%; group A) had a history of seizure(s). Forty-seven cats (77%; group B) had no history of seizure(s). Generalised tonic-clonic seizures were seen in eight cats (57%) and were the most common seizure pattern in our cats with intracranial neoplasia. Clusters of seizures were observed in six cats. Status epilepticus was observed in one patient. The mean age of the cats was 7.9 years within group A (median 8.5) and 9.3 years (median 10) within group B. The cats with lymphoma within both groups were significantly younger than cats with meningioma. In both groups meningioma and lymphoma were confirmed to be the most frequent tumour type, followed by glial cell tumours. The prevalence of the seizures in patients with glial cell tumours was 26.7%, 26.3% in patients with lymphomas and 15% in cases with meningiomas. In 33 cases (54.1%) the tumours were localised in the forebrain, 15 tumours (24.6%) were in the brainstem, four (6.6%) in the cerebellum and nine tumours (14.7%) had multifocal localisation. Parietal lobe and basal ganglia mostly affected group A. In group B tumours were most frequently located in the parietal and frontal lobes as well as in the diencephalon. A positive association was documented between the localisation of a tumour in the forebrain and seizure occurrence.

Morbidity in 201 patients with small sized meningioma treated by microsurgery

The management of patients with small, often asymptomatic meningiomas is controversial and includes observation, microsurgery (MS) and stereotactic radiosurgery (SRS). The purpose of this retrospective study was to analyze the morbidity and the extent of removal after MS for small (< or =3 cm) intracranial meningiomas and compare these results to those of SRS reported in the literature.

Neuropeptide Y receptors in primary human brain tumors: overexpression in high-grade tumors

Peptide receptors are often overexpressed in tumors, and they may be targeted in vivo. We evaluated neuropeptide Y (NPY) receptor expression in 131 primary human brain tumors, including gliomas, embryonal tumors, meningiomas, and pituitary adenomas, by in vitro receptor autoradiography using the 125I-labeled NPY receptor ligand peptide YY in competition with NPY receptor subtype-selective analogs. Receptor functionality was investigated in selected cases using [35S]GTPgammaS-binding autoradiography. World Health Organization Grade IV glioblastomas showed a remarkably high expression of the NPY receptor subtype Y2 with respect to both incidence (83%) and density (mean, 4,886 dpm/mg tissue); astrocytomas World Health Organization Grades I to III and oligodendrogliomas also exhibited high Y2 incidences but low Y2 densities. In glioblastomas, Y2 agonists specifically stimulated [35S]GTPgammaS binding, suggesting that tumoral Y2 receptors were functional. Furthermore, nonneoplastic nerve fibers containing NPY peptide were identified in glioblastomas by immunohistochemistry. Medulloblastomas, primitive neuroectodermal tumors of the CNS, and meningiomas expressed Y1 and Y2 receptor subtypes in moderate incidence and density. In conclusion, Y2 receptors in glioblastomas that are activated by NPY originating from intratumoral nerve fibers might mediate functional effects on the tumor cells. Moreover, identification of the high expression of NPY receptors in high-grade gliomas and embryonal brain tumors provides the basis for in vivo targeting.

Extracranial expansion of a feline meningioma

Meningioma is the most frequently observed primary brain tumour in cats. Usually, it is associated with an intracranial expansion with consequent brain compression, oedema and brain herniation. Typical features of feline intracranial meningiomas are hyperostosis of the adjacent bone and intratumoral mineralisation. We describe a 13-year-old male neutered cat with a 1-year history of behavioural change. At clinical and neurological examination the cat showed signs consistent with right-sided forebrain lesion. Magnetic resonance images showed a right-sided extra-axial contrast enhancing mass in the region of the frontotemporal lobe. The overlying bone of the calvarium showed a marked defect with extracranial expansion of the tissue. Surgery was performed and the tumour could be exposed by a right-sided temporal approach. After extension of the bony defect the mass could be removed properly. The cat recovered well from surgery and a 12-month follow-up showed no persistent neurological deficits. Histopathological assessment of the tumour revealed a transitional grade 1 meningioma. Despite osteolysis and extracranial expansion of the tumour differentials should include menigioma in feline intracranial neoplasms.

The ROAM/EORTC-1308 trial: Radiation versus Observation following surgical resection of Atypical Meningioma: study protocol for a randomised controlled trial.

BACKGROUND Atypical meningiomas are an intermediate grade brain tumour with a recurrence rate of 39-58 %. It is not known whether early adjuvant radiotherapy reduces the risk of tumour recurrence and whether the potential side-effects are justified. An alternative management strategy is to perform active monitoring with magnetic resonance imaging (MRI) and to treat at recurrence. There are no randomised controlled trials comparing these two approaches. METHODS/DESIGN A total of 190 patients will be recruited from neurosurgical/neuro-oncology centres across the United Kingdom, Ireland and mainland Europe. Adult patients undergoing gross total resection of intracranial atypical meningioma are eligible. Patients with multiple meningioma, optic nerve sheath meningioma, previous intracranial tumour, previous cranial radiotherapy and neurofibromatosis will be excluded. Informed consent will be obtained from patients. This is a two-stage trial (both stages will run in parallel): Stage 1 (qualitative study) is designed to maximise patient and clinician acceptability, thereby optimising recruitment and retention. Patients wishing to continue will proceed to randomisation. Stage 2 (randomisation) patients will be randomised to receive either early adjuvant radiotherapy for 6 weeks (60 Gy in 30 fractions) or active monitoring. The primary outcome measure is time to MRI evidence of tumour recurrence (progression-free survival (PFS)). Secondary outcome measures include assessing the toxicity of the radiotherapy, the quality of life, neurocognitive function, time to second line treatment, time to death (overall survival (OS)) and incremental cost per quality-adjusted life year (QALY) gained. DISCUSSION ROAM/EORTC-1308 is the first multi-centre randomised controlled trial designed to determine whether early adjuvant radiotherapy reduces the risk of tumour recurrence following complete surgical resection of atypical meningioma. The results of this study will be used to inform current neurosurgery and neuro-oncology practice worldwide. TRIAL REGISTRATION ISRCTN71502099 on 19 May 2014.

The scaffold protein KSR1, a novel therapeutic target for the treatment of Merlin-deficient tumors

Merlin has broad tumor-suppressor functions as its mutations have been identified in multiple benign tumors and malignant cancers. In all schwannomas, the majority of meningiomas and 1/3 of ependymomas Merlin loss is causative. In neurofibromatosis type 2, a dominantly inherited tumor disease because of the loss of Merlin, patients suffer from multiple nervous system tumors and die on average around age 40. Chemotherapy is not effective and tumor localization and multiplicity make surgery and radiosurgery challenging and morbidity is often considerable. Thus, a new therapeutic approach is needed for these tumors. Using a primary human in vitro model for Merlin-deficient tumors, we report that the Ras/Raf/mitogen-activated protein, extracellular signal-regulated kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) scaffold, kinase suppressor of Ras 1 (KSR1), has a vital role in promoting schwannomas development. We show that KSR1 overexpression is involved in many pathological phenotypes caused by Merlin loss, namely multipolar morphology, enhanced cell-matrix adhesion, focal adhesion and, most importantly, increased proliferation and survival. Our data demonstrate that KSR1 has a wider role than MEK1/2 in the development of schwannomas because adhesion is more dependent on KSR1 than MEK1/2. Immunoprecipitation analysis reveals that KSR1 is a novel binding partner of Merlin, which suppresses KSR1's function by inhibiting the binding between KSR1 and c-Raf. Our proteomic analysis also demonstrates that KSR1 interacts with several Merlin downstream effectors, including E3 ubiquitin ligase CRL4DCAF1. Further functional studies suggests that KSR1 and DCAF1 may co-operate to regulate schwannomas formation. Taken together, these findings suggest that KSR1 serves as a potential therapeutic target for Merlin-deficient tumors.

Molecular biology and biochemistry of brain tumours

ßElucidating some molecular mechanisms and biochemistry of brain tumours is an important step towards the development of adjuvant medical therapies. The present study concentrates on cholecystokinin (CCK), a gut-brain peptide that has been described to be able to induce mitosis of rat gliomas as well as hormone secretion by the anterior pituitary, via the CCK-B receptor. The significance of a polymorphism in the growth hormone releasing hormone (GHRH) receptor (GHRH-R) gene was also determined. Finally, defects in the ß-catenin gene, an important component of the developmental pathway, in a sub-set of craniopharyngiomas were investigated. Reverse transcription-polymerase chain reaction (RT-PCR), restriction digestion analysis and direct sequencing demonstrated expression of CCK peptide itself and its A and B receptors by human gliomas, meningiomas and pituitary tumours. CCK peptides stimulated growth of cultured gliomas and meningiomas as well as in vitro hormone secretion [growth hormone (GH), luteinizing hormone (LH) and follicle stimulating hormone (FSH)] by human pituitary tumours. These biological effects were reduced or abolished by CCK antagonists. In addition, an antibody to CCK reduced mitosis by gliomas and meningiomas, and the same antibody inhibited hormone secretion by cultured human pituitary tumours. CCK peptides stimulated phosphatidylinositol (PI) hydrolysis, indicating coupling of the CCK receptors to phopsholipase C. Cyclic AMP was unaffected. In addition, caspase-3 activity was significantly and markedly increased, whilst proteasome activity was decreased. Taken together, these results may indicate an autocrine/paracrine role of CCK in the control of growth and/or functioning of gliomas, meningiomas and pituitary tumours. Primer induced restriction analysis (PIRA) of a rarer and alternative polymorphism in the GHRH-R receptor, in which Thr replaces Ala at codon 57, in human GH-secreting pituitary tumours was investigated. Whilst the rarer form correlated with an increased response of the pituitary cells to GHRH in vitro, allele distribution studies revealed that it is unlikely that the polymorphism contributes to increased risk of developing GH-secreting tumours and therefore acromegaly. Further findings of this study, using PCR and direct sequencing, were the demonstration of an association between b-catenin gene alterations and craniopharyngiomas of the adamantinomatous type. Since this gene product is involved with development, these results suggest that p-catenin mutations may contribute to the initiation and subsequent growth of congenital adamantinomatous craniopharyngiomas.

Proton radiotherapy: a therapeutic opportunity for pediatric brain tumor patients

Proton radiation therapy is a form of external radiation that uses charged particles which have distinct physical advantages to deliver the majority of its dose in the target while minimizing the dose of radiation to normal tissues. In children who are particularly susceptible even at low and medium doses of radiation, the significant reduction of integral dose can potentially mitigate the incidence of side effects and improve quality of life. The aim of the first part of the thesis is to describe the physical and radiobiological properties of protons, the Proton Therapy Center of Trento (TCPT) active for clinical purpose since 2014, which use the most recent technique called active pencil beam scanning. The second part of the thesis describes the preliminary clinical results of 23 pediatric patients with central nervous system tumors as well as of two aggressive pediatric meningiomas treated with pencil beam scanning. All the patients were particularly well-suited candidates for proton therapy (PT) for possible benefits in terms of survival and incidence of acute and late side effects. We reported also a multicentric experience of 27 medulloblastoma patients (median age 6 years, M/F ratio 13/14) treated between 2015 and 2020 at TPTC coming from three Pediatric oncology centers: Bologna, Florence, and Ljubljana, with a focus on clinical results and toxicities related to radiotherapy (RT). Proton therapy was associated with mostly mild acute and late adverse effects and no cases of CNS necrosis or high grade of neuroradiological abnormailities. Comparable rates of survival and local control were obtained to those achievable with conventional RT. Finally, we performed a systematic review to specifically address the safety of PT for pediatric CNS patients, late side effects and clinical effectiveness after PT in this patient group.