Complete remission of a diffuse pontine glioma.

A patient is described in whom a large diffuse glioma of the pons extending into the midbrain was diagnosed at the age of 2 years. Biopsy showed a fibrillary astrocytoma. After shunting of a hydrocephalus, the clinical symptoms abated without conventional therapy. Repeated MRI studies showed a continuous decrease of the tumour which was no longer visible when the patient was 6.6 years old. In reviews on spontaneous remissions of oncologic disorders we were unable to find a case of a biologically benign brain stem tumour. There is one isolated report on a similar case, though without histologic documentation.

Analysis of p53 mutation and epidermal growth factor receptor amplification in recurrent gliomas with malignant progression.

Genomic alterations and expression of the p53 tumor suppressor gene and the epidermal factor receptor gene (EGFR) were investigated in 22 patients with primary World Health Organization (WHO) grade II gliomas that on recurrence had progressed to malignant gliomas of WHO grades III or IV. Mutations of the p53 gene (exons 5 to 8) were found in 12 of 22 primary tumors (10 of 13 astrocytomas, 1 of 7 oligodendrogliomas, 1 of 2 oligoastrocytomas). In each of these cases identical p53 mutations were present in the respective malignant recurrences. In all instances in which the p53 mutation was associated with p53 protein accumulation (10 of 12 cases) the percentage of p53 immunopositive tumor cells had increased from the primary to the recurrent tumor. None of the primary low-grade and none of the recurrent high-grade tumors (7 anaplastic astrocytomas, 10 anaplastic oligodendrogliomas, 4 anaplastic oligoastrocytomas, and 5 glioblastomas) showed evidence of EGFR gene amplification. Our results thus demonstrate p53 is mutated in a high fraction of low-grade astrocytomas with progression to anaplastic astrocytomas and glioblastomas and that progression in such cases is frequently associated with an increase in the fraction of p53 immunopositive tumor cells. The general absence of EGFR amplification in our tumor series supports the hypothesis that the significance of p53 mutation and EGFR amplification may be different in glioblastomas that developed by progression from low-grade astrocytomas (secondary glioblastomas) compared to glioblastomas that developed rapidly in a de novo manner without a history of previous low-grade tumor (primary glioblastomas).

Epidermal growth factor receptor expression in oligodendroglial tumors.

A series of 13 oligodendrogliomas (WHO grade II) and 20 anaplastic oligodendrogliomas (WHO grade III) was studied for gene amplification and expression of the epidermal growth factor receptor gene (EGFR). EGFR gene amplification was found in only one case of anaplastic oligodendroglioma, which additionally showed a deletion/rearrangement at the 5' end of the gene. Northern blot analysis, however, revealed increases of EGFR mRNA expression relative to non-neoplastic control brain in 6 of 13 oligodendrogliomas and 10 of 18 anaplastic oligodendrogliomas. All cases with increased mRNA expression showed strong immunoreactivity for EGFR protein. Our findings thus indicate that increased expression of EGFR mRNA and protein is common in low-grade and high-grade oligodendroglial tumors and in the vast majority of cases is not caused by gene amplification.

Primary glioblastoma multiforme of the oculomotor nerve. Case report.

Tumors of the oculomotor nerve are rare and most instances reported have been schwannomas. The authors present clinical, neuroradiological, and neuropathological findings in a 70-year-old woman with a glioblastoma multiforme (GBM) growing primarily in the proximal part of the left oculomotor nerve. The patient presented with a 1-month history of transient diplopia. Neurological examination revealed an incomplete left-sided oculomotor nerve palsy with no further signs of neurological dysfunction. Cranial computerized tomography and magnetic resonance imaging showed a tumor of the left oculomotor nerve without any obvious signs of penetration into the midbrain or upper pons. Following subtotal removal of the tumor, neuropathological examination of the operative specimen revealed a GBM that had grown diffusely within peripheral nerve tissue. Six weeks after surgery, the patient suddenly died of pulmonary thromboembolism. Postmortem examination of the brain confirmed a large leptomeningeal GBM at the left pontomesencephalic junction with complete destruction of the left oculomotor nerve. To the authors' knowledge, this represents the first case of a GBM of the oculomotor nerve, probably originating from glial cells within the most proximal part of the nerve or the adjacent leptomeninges.

Mutation of the Von Hippel-Lindau tumour suppressor gene in capillary haemangioblastomas of the central nervous system.

A series of 20 capillary haemangioblastomas of the central nervous system was screened for mutations of the von Hippel-Lindau (VHL) tumour suppressor gene by single strand conformational polymorphism (SSCP) and heteroduplex analysis. Aberrant polymerase chain reaction (PCR) products were detected in ten tumours. DNA sequencing of these PCR products revealed that seven tumours had frameshift mutations due either to deletions of one or more base pairs (six cases) or to insertion of one base pair (one case). The remaining three tumours had either point mutations of intron splice site sequences (two cases) or a point mutation resulting in an amino acid substitution (one case). Evidence for germline alterations of the VHL gene was found in two patients who showed identical mutations in both tumour and corresponding leukocyte DNA. The results suggest that mutation of the VHL tumour suppressor gene represents a significant event in the development of capillary haemangioblastomas.

Coexpression of transforming growth factor-alpha and épidermal growth factor receptor in capillary hemangioblastomas of the central nervous system.

The expression of epidermal growth factor receptor (EGFR) and the pre-pro form of one of its ligands, transforming growth factor-alpha (TGF-alpha), was studied by Northern blotting in a series of 14 capillary hemangioblastomas of the central nervous system. A constant coexpression of EGFR and pre-pro-TGF-alpha mRNAs was found. Immunocytochemical investigation of an extended series of 51 capillary hemangioblastomas revealed that the stromal cells in these tumors showed immunoreactivity with monoclonal antibodies to EGFR and TGF-alpha. Analysis of gene dosage by Southern blotting in 20 tumors indicated a normal gene copy number of EGFR and TGF alpha in all cases. Our findings suggest that autocrine and/or juxtacrine growth stimulation via the EGFR may contribute to tumor growth in capillary hemangioblastomas.

The epidermal growth factor receptor in glioblastoma: genomic amplification, protein expression, and patient survival data in a therapeutic trial.

Human glioblastomas were evaluated for overexpression of the epidermal growth factor receptor (EGFR) in a therapeutic trial with the anti-EGFR antibody EMD 55900. A total of 55 cases were examined by immunohistochemistry using 4 different monoclonal antibodies on frozen or on paraffin sections: EGFR-1 (Amersham), E 62 (Merck), E 30 (Merck), and EMD 55900 (MAB 425, Merck). Definition for inclusion in clinical trials of EMD 55900 was an immunohistochemical overexpression of grade 4+ or 3+ in a scale of 4 grades of staining quality. The use of the 4 different antibodies gave essentially equal results. In 21 cases, the immunohistochemical results were supplemented by molecular genetic analysis of EGFR amplification on the corresponding locus of chromosome 7, using frozen tissue from the same blocks after screening for vital tumor areas. Since no other material was available, the differential polymerase chain reaction technique was applied. Interferon-gamma (IFN-gamma) served as a reference gene. In a preliminary experimental series with cases of known EGFR amplification, a densitometric EGFR/IFN-gamma ratio higher than 3 was determined as indicator for amplification of the EGFR gene. With this experimental approach we were able to identify an amplified EGFR gene in 13 specimens including 2 from recurrent glioblastomas in the same patients. All of these showed an increased immunoreactivity for EGFR protein. The degree of EGFR amplification (EGFR/IFN-gamma ratio as measured by DNA densitometry) showed a positive correlation with the grade of immunohistochemical protein expression, both in regard to the fraction of positive cells and to the overall staining intensity.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular genetic analysis of oligodendroglial tumors shows preferential allelic deletions on 19q and 1p.

The molecular genetic alterations of oligodendroglial tumors and mixed gliomas of the central nervous system were studied in a series of 37 cases (8 oligodendrogliomas, 13 anaplastic oligodendrogliomas, 8 oligoastrocytomas, and 8 anaplastic oligoastrocytomas). A total of 180 polymorphic loci and 5 nonpolymorphic gene loci, distributed over all chromosomes, were examined by restriction fragment length polymorphism analysis. Loss of heterozygosity was most frequently observed for loci on 19q with a commonly deleted region at 19q13.2-q13.4 distal to the CYP2a gene and proximal to the D19S22 locus. The incidence of allelic loss on 19q was particularly high (81%) in oligodendroglial tumors and equal to 31% in mixed gliomas. More than 75% of the tumors with allelic deletions on 19q also showed loss of heterozygosity for loci on 1p with one tumor showing only loss of alleles distal to the NGFB gene (1p13-pter). Seven (19%) tumors had lost alleles from 17p with the deleted region including the TP53 tumor suppressor gene in all cases. Sequencing of the TP53 transcripts from exons 2 to 10, however, did not reveal mutations of the remaining allele in any of these tumors. Anaplastic oligodendrogliomas and anaplastic oligoastrocytomas demonstrated an increased incidence of additional allelic losses involving most frequently chromosomes 9p and 10. Gene amplification was detected in two anaplastic tumors, affecting the epidermal growth factor receptor gene in both cases, with additional amplification of the renin gene at 1q32 in one of these cases. In total our results indicate both differences and similarities between the molecular genetic alterations in tumors with oligodendroglial and astrocytic differentiation. The loss of genetic information from 19q and 1p as well as the rarity of TP53 mutations in oligodendroglial tumors suggests that the early events in their oncogenesis are distinct from those associated with astrocytic tumors. However, similarities are indicated by the allelic losses on 9p and 10 in the anaplastic tumors, suggesting the utilization of common pathways of progression.

The stromal Schwann cell during maturation of peripheral neuroblastomas. Immunohistochemical observations with antibodies to the neuronal class III beta-tubulin isotype (beta III) and S-100 protein.

This immunohistochemical study compares the localization of the neuronal class III beta-tubulin isotype (beta III; analogous to the beta' 1-/beta 2-tubulin isoform) to the Schwann cell-associated S-100 protein focusing on topographic relationships of Schwann-like cells to differentiating neuronal phenotypes during stromal development in human peripheral neuroblastomas. The earliest appearance of Schwann cells in poorly differentiated (classical) neuroblastomas is heralded by S-100 protein-immunoreactive cells in close association with tumor blood vessels. In subsequent stages of maturation, i.e. maturing neuroblastoma (ganglioneuroblastoma and gangliocytoma), S-100 protein-positive cells are mostly confined to the connective tissue septa dividing tumor into lobules, and are not freely interspersed with beta III-immunoreactive neoplastic neurons. Significant ensheathment of individual axon-like processes by Schwann cells occurs only in mature ganglioneuromas. beta III is localized in a full spectrum of neoplastic neuronal phenotypes, ranging from poorly-differentiated apolar neuroblasts (often signaling ensuing neuritogenesis) to mature ganglion cells, but not in Schwann cells, or other cell types of the stroma. Our observations suggest that Schwann cells in peripheral neuroblastomas are stroma-derived cells and not an expression of divergent neoplastic differentiation.

[Alleles in chromosome 10p21-26 in malignant gliomas]. / Allelieuntersuchungen auf Chromosom 10q21-26 in malignen Gliomen.

Loss of genetic material on chromosome 10 is regarded as a prominent feature in the genesis of glioblastomas. To use chromosome 10 deletions as diagnostic markers for glioblastomas we investigated, if the loss of chromosome 10 material could be restricted on the region 10q21-26. By PCR microsatellite analysis on frozen tissue and paraffin material from the ZULCH brain tumor collection we found (1) loss of heterozygosity in 10q21-26 in 75% of the investigated DNA from frozen tissue and (2) an interstitial loss in the region of the microsatellite marker D10S186. The combined immunohistochemical analysis of overexpression of EGFR, EGF and TGF alpha with LOH on chromosome 10 showed that chromosome 10 deletions are not exclusively bound to EGFR overexpression.

Frequency and topographical distribution of CD68-positive macrophages and HIV-1 core proteins in HIV-associated brain lesions.

We report the neuropathological and immunohistochemical findings in the brains of 14 AIDS patients with HIV-related encephalopathy. Clinically, half of the patients presented with severe AIDS dementia complex including advanced psychomotor retardation and behavioural dysfunction. These features correlated with striking cerebral atrophy and subcortical lesions visible in CT and/or MRI scans. In 7 cases early signs of impaired memory and concentration and/or psychomotor slowing were apparent accompanied by subcortical lesions in MRI scans and normal CCTs. In order to investigate the topographical distribution of HIV-1-associated features, in every case tissue samples from the frontal, temporal, parietal, occipital cortex and subcortical white matter, the hippocampus, basal ganglia, midbrain, pons, medulla oblongata and cerebellum were studied. In all patients histological examination disclosed the typical cellular constituents of HIV encephalitis (n = 12) or leukoencephalopathy (n = 2). Antibodies against lymphocyte subsets, CD68 antigen, myelin basic protein and GFAP were used to characterize the phenotype of cells and to highlight the white matter gliosis. The distribution and degree of pathological features were analysed in a semiquantitative scale, based on the number of CD68-positive cells, and disclosed great interindividual differences concerning the affected brain regions which only in part correlated with the severity of the clinical picture. It is noteworthy, that the deep gray matter, in particular putamen and thalamus, was involved in every case, independent from the stage of the disease. In addition, quantity and topographical distribution of HIV-1 core protein p24 were studied by use of two monoclonal antibodies. It is noteworthy, that the number of immunoreactive multinucleated giant cells and microglial cells decreased gradually from the deep gray matter, especially putamen and thalamus, and deep white matter to corpus callosum, cerebellar white matter and subcortical cerebral white matter. The topographical predilection of the deep gray matter even in cases with early cognitive decline indicates that the basal ganglia are affected early in the course of the disease. This observation closely resembles the results of highly sensitive quantitative neuropsychological tests which disclosed slowing and impaired coordination of rapid extremity movements indicating basal ganglia lesions even in early stages of HIV dementia.

neu mutations and loss of normal allele in schwannomas induced by N-ethyl-N-nitrosourea in rats.

The mutational activation of the neu proto-oncogene was analyzed in schwannomas of the peripheral nervous system transplacentally induced by N-ethyl-N-nitrosourea (ENU) in rats. All nine primary schwannomas investigated contained a T-->A transversion mutation at nucleotide 2012 of the neu gene, leading to the substitution of valine to glutamic acid in the transmembrane domain. Loss of the wild-type allele occurred in five of nine (56%) primary schwannomas. All four ENU-induced schwannomas transplanted in syngenic hosts (2-14 passages) showed the T-->A transversion mutation and loss of the wild-type allele. These data suggest that mutation of one allele together with loss of the wild-type allele constitutes two critical steps in the progression of rat schwannomas. Since c-erbB-2, the human counterpart of the rat neu gene is frequently involved by amplification in the development of human gastric cancer [30], we screened 12 rat gastric carcinomas induced by N-methyl-N-nitrosourea (MNU) for mutations in the neu gene. However, none of these tumors contained mutations.

Neuropathological studies in the brains of AIDS patients with opportunistic diseases.

The brains of 70 fatal cases with AIDS were studied by means of immunohistochemistry and in-situ hybridization in a consecutive autopsy series (1985-July 1992). In addition, the neuropathological changes were correlated with the neurological and neuroimaging findings. Opportunistic infections included toxoplasmosis (15 cases), cytomegalovirus (CMV)-encephalitis (6), progressive multifocal leucoencephalopathy (2) and fungal infections (3). Malignant lymphomas were found in 7 patients; 6 involved primarily the CNS, one was metastatic. In 14 cases the neuropathological changes were consistent with HIV encephalitis and HIV leucoencephalopathy. Non-specific lesions occurred in 31 cases. The clinical diagnosis in patients with opportunistic diseases (n = 27) diverged in 15 cases (55%) from the underlying pathology. Toxoplasma gondii, CMV and JC viruses were identified by immunohistochemistry and in-situ hybridization on serial paraffin sections. In addition, antibodies against lymphocyte subsets, tissue macrophages, the glial fibrillary acid protein (GFAP) and myelin basic protein were used to characterize the phenotype of cells and to highlight the degree of gliosis and demyelination. Our results show that the distribution and degree of morphological changes might be helpful for the differential diagnosis antemortem. Since neurological complications may represent the first or sole manifestation of AIDS and risk factors for AIDS are often not known, it should be taken into account that CNS manifestations of AIDS may contribute to a sudden and unexpected death or accident. Opportunistic diseases should be considered as a possible differential diagnosis in cases mimicking the clinical picture of apoplexia or dementia. Furthermore, CNS lesions may be detected postmortem in patients who were not known to suffer from Neuro-Aids during life, indicating that CNS involvement is more widespread than assumed.

Expression of CD15 in tumours of the nervous system.

The expression of the CD15 epitope was investigated by immunohistochemistry, western blotting and immuno-thin-layer chromatography on a large series of human nervous system tumours and ethylnitrosourea-induced rat gliomas. Our results show that CD15 is expressed as a glycoprotein- or glycolipid-associated epitope in normal human and rat brain. In contrast, immunoreactivity for CD15 was absent in tumour cells of experimental rat gliomas. In human tumours we found a more complex expression pattern. While intra- and perivascular granulocytes as well as macrophages in necrotic areas of anaplastic tumours were always strongly CD15-positive, immunoreactive tumour cells were detectable only in a fraction of low-grade gliomas. Anaplastic gliomas and glioblastomas consistently did not express the epitope on their tumour cells. In addition to individual low-grade gliomas, we found CD15-positive cases among metastatic carcinomas, craniopharyngeomas, meningiomas, germinomas and malignant melanomas. Our results suggest that immunohistochemistry for CD15 is potentially useful in diagnostic neuropathology as a marker for granulocytes in paraffin sections, as a supplementary tool for the histopathological grading of gliomas, and as an aid for differentiation between anaplastic glioma cells and non-neoplastic glia. Furthermore, it can be speculated that the lack of CD15 expression on anaplastic glioma cells may potentially be responsible for some of their characteristics--such as altered cellular interaction and loss of contact inhibition.

Immunotherapy with stimulated autologous lymphocytes in a case of a juvenile anaplastic glioma.

The effect of immunotherapy with stimulated autologous lymphocytes (SAL) in malignant gliomas is documented and discussed in a bioptical and autoptical case study. A five-year-old child with a recurrently operated and radiated right hemispheric anaplastic astrocytoma died six weeks after immunotherapy with mitogen-activated killer cells and recombinant Interleukin-2. The autopsy revealed a large butterfly glioma with partially necrotic gelatinous tissue at the site of the SAL reservoir. The tumor cell density on the right was less than on the left hemisphere, and T-lymphocyte content was higher on the right hemisphere. These results demonstrate a local effect of SAL therapy in vivo, although the tumor progression as a whole could not be stopped. They also demonstrate the need of a detailed neuropathological examination in all cases of immunotherapy of malignant gliomas.

Occlusive microangiopathy by immunoglobulin (IgM-kappa) precipitation: pathogenetic relevance in paraneoplastic cryoglobulinemic neuropathy.

Histological, immunohistochemical and ultrastructural sural nerve and skin biopsy findings in a case of cryoglobulinemia secondary to an IgM-kappa-producing non-Hodgkin lymphoma are described. The main finding was an occlusive microangiopathy present in both the sural nerve and the skin. Widespread cryoglobulin deposits of the proliferated vasa nervorum were associated with pronounced changes probably evoked by ischemia. Moderate perivascular inflammation, but no florid vasculitis was additionally present. Our observations indicate that occlusive microangiopathy by precipitated cryoglobulins may be a relevant pathogenetic factor in cryoglobulinemic peripheral neuropathy.

Vascular permeability changes in tumours of the peripheral nervous system.

Vascular permeability changes were examined in 34 tumours of the peripheral nervous system by immunohistochemical demonstration of serum proteins as endogenous tracers. The blood-tumour barrier was impaired in the reticular (Antoni type B) portions of neurinomas (Schwannomas) and in cutaneous neurofibromas but was similar to the normal blood-nerve barrier in fibrillary (Antoni type A) neurinomas, in most neurofibromas, in ganglioneuromas and in anaplastic tumours. These differences in permeability are discussed in relation to aspects of pathological tumour vascularization, the histogenesis of microcystic changes, and systemic therapeutic approaches.

Gliofibroma: immunohistochemical analysis.

A case of gliofibroma occurring in an adult patient as a large circumscribed supratentorial tumor is reported. The bimorphic pattern was substantiated and further analyzed by immunohistochemistry. Some evidence in favor of collagen production by mesenchymal and/or inflammatory cells leading to a progressive fibrous replacement of the glial cells in this particular tumor type is presented.

Immunohistochemical demonstration of proliferating cell nuclear antigen in glioblastomas: pronounced heterogeneity and lack of prognostic significance.

In order to evaluate a possible significance of the expression of proliferating cell nuclear antigen (PCNA) as clinically useful prognostic parameter, we retrospectively investigated a series of 40 glioblastomas by means of immunohistochemistry and compared the results to patient survival. All glioblastomas included in the study had been treated by operation, radiotherapy and intraarterial ACNU [3-(4-amino-2-methyl-5-pyrimidinylmethyl)-1-(2-chloroethyl)-1-nitr osourea] chemotherapy. Patient survival ranged from 2 months to 42 months (mean: 14.2 months). PCNA values varied widely, ranging from 0.5% to 75% (mean: 24.9%). Statistical analysis revealed no significant correlation between PCNA index and patient survival. Our study thus indicates that the expression of PCNA appears not to be a useful prognostic parameter for glioblastoma patients.