Multiscale quantification of morphological heterogeneity with creation of a predictor of longer survival in glioblastoma

Intratumor heterogeneity is a main cause of the dismal prognosis of glioblastoma (GBM). Yet, there remains a lack of a uniform assessment of the degree of heterogeneity. With a multiscale approach, we addressed the hypothesis that intratumor heterogeneity exists on different levels comprising traditional regional analyses, but also innovative methods including computer-assisted analysis of tumor morphology combined with epigenomic data. With this aim, 157 biopsies of 37 patients with therapy-naive IDH-wildtype GBM were analyzed regarding the intratumor variance of protein expression of glial marker GFAP, microglia marker Iba1 and proliferation marker Mib1. Hematoxylin and eosin stained slides were evaluated for tumor vascularization. For the estimation of pixel intensity and nuclear profiling, automated analysis was used. Additionally, DNA methylation profiling was conducted separately for the single biopsies. Scoring systems were established to integrate several parameters into one score for the four examined modalities of heterogeneity (regional, cellular, pixel-level and epigenomic). As a result, we could show that heterogeneity was detected in all four modalities. Furthermore, for the regional, cellular and epigenomic level, we confirmed the results of earlier studies stating that a higher degree of heterogeneity is associated with poorer overall survival. To integrate all modalities into one score, we designed a predictor of longer survival, which showed a highly significant separation regarding the OS. In conclusion, multiscale intratumor heterogeneity exists in glioblastoma and its degree has an impact on overall survival. In future studies, the implementation of a broadly feasible heterogeneity index should be considered.     

Expression of the c-erbB-2-encoded oncoprotein and progesterone receptor in human meningiomas

The present study investigated the expression of c-erbB-2 in 59 meningiomas, including different histological subtypes and anaplastic variants, by immunocytochemistry and molecular biological techniques. Immunohistochemistry using the monoclonal antibody FWP-51 directed against c-erbB-2-encoded oncoprotein gp185 demonstrated variable degrees of immunoreactivity in all meningiomas. The intensity of immunostaining correlated with the degree of expression as assessed by Western analysis in 28 meningiomas using polyclonal antiserum 21N. There was no correlation between the degree of expression and histological variants. Immunoreactivity of all menigiomas was distinctly less intense, however, than that of the human breast cancer cell line SK-BR-3, and slightly lower than that of brain metastases of breast and ovarian carcinomas that served as positive controls for both methods. By Southern analysis all meningiomas showed a single copy of the c-erbB-2 gene. Non-neoplastic arachnoid cap cells also exhibited c-erbB-2 expression and the degree of immunoreactivity was comparable with the majority of meningiomas. These data argue against an overexpression of c-erbB-2 in meningiomas, but rather indicate a cell-type-specific constitutive expression of the c-erbB-2 gene product in meningiomas and their putative progenitor cells. Since a subgroup of meningiomas is known to express progesterone receptors (PR), gp185 immunoreactivity was compared to the hormone receptor status using monoclonal antibody KD68. Fifty-six percent meningiomas showed PR immunoreactivity, but there was no statistically significant correlation with the degree of gp185 expression.This study was supported by a grant of the Tumorzentrum Heidelberg/Mannheim (M.K., No. 10028060)     

HPA-5b (Bra) neonatal alloimmune thrombocytopenia: clinical and immunological analysis of 39 cases

Maternal alloimmunization against fetal platelets can cause fetal and neonatal thrombocytopenia (NAIT). The HPA-1a (PIA1, Zwa) antigen is by far the most common antigen implicated in NAIT. However, today another antigen often linked with that affection is HPA-5b (Br(a)). This is a report of 39 cases of NAIT involving the HPA-5b antigen. Thrombocytopenia may be of grave consequence. Three infants developed intracerebral haemorrhages (ICH). Of these, one died presumably as a consequence of ICH. Central nervous system (CNS) sequelae in the neonatal period was observed in two children. The potential hazards of death or disabling neurologic sequelae following intracerebral haemorrhage call for rapid and reliable diagnosis and effective therapy. Because there is high risk that subsequent pregnancies might be also affected by NAIT, the mothers of a previously affected child should be managed similarly to the HPA-1b mothers (PIA2, Zwb). The antenatal diagnosis of thrombocytopenia should be made and if necessary the in utero therapy instituted.     

Preclinical animal study and clinical trail of modified extraoral craniofacial implants.

We report on our experience using a new endosseous implant designed to provide sufficient retention to various types of facial prostheses. In a preclinical animal experiment implants (N=12, 4 x 3.5 mm) were placed in the frontal calvarial region of nine adult pigs. The animals were sacrificed at 2, 4 and 8 weeks to evaluate the implant incorporation microradiographically. The clinical outcome and patient satisfaction of implant-retained prostheses were evaluated in a group of 10 patients with facial defects by using clinical assessment and standardized questionnaires for patients and relatives. In the prospective clinical study 33 identical modified implants for extraoral anchorage were placed for the fixation of various prostheses in the midfacial (eye, nose) and ear regions in the course of a clinical trial and observed over a follow-up period of 34 months. The bone-implant contact in the animal experiment reached 31% (+/-2) at 2 weeks, 39% (+/-1) after 4 weeks and 51% (+/-5) at 8 weeks. In the clinical trial, no implants were lost and all implants remained osseointegrated as confirmed clinically and radiographically, providing a stable prosthetic restoration. The analysis of the questionnaire indicates an improvement of the quality of life of patients with respect to aesthetic and psychological well-being. The results demonstrate that extraoral implants not only achieve sufficient osseointegration but also show good clinical handling and easy fixation possibilities for prosthetic anchorage.     

Testicular function after OPA/COMP chemotherapy without procarbazine in boys with Hodgkin's disease. Results in 25 patients of the DAL-HD-85 study

Gonadal function was evaluated in 25 boys treated for Hodgkin's disease according to the DAL-HD-85 protocol with OPA- or OPA/COMP-chemotherapy (vincristine-prednisone-adriamycine/cyclophosphamide-vincristine-m ethotrexate- prednisone). All boys were in first continuous complete remission for 6 to 45 months at chronological ages varying from 14.0 to 18.9 years. Testosterone, basal and GnRH-stimulated LH- and FSH-levels were measured. Gonadal function was normal in 16 patients treated with 2 cycles of OPA-chemotherapy in Hodgkin stages I-IIA. 9 patients were treated with 2 OPA- and 2 or 4 COMP-cycles of chemotherapy and had received mean cyclophosphamide doses ranging from 2004 to 3722 mg/m2. Again, no major testicular damage was noted, though some patients had increased stimulated LH-levels possibly indicating compensated Leydig cell-insufficiency. Our results demonstrate, that testicular function is not severely affected when patients are treated for Hodgkin's disease without procarbazine even if cyclophosphamide is given in cumulative doses below 3800 mg/m2. The previously documented severe testicular damage in boys treated according to the DAL-studies HD-78 and HD-82 is thus a result of the gonadotoxic action of procarbazine.     

Computed tomography in depression: association between ventricular size and psychopathology

The relationship between psychopathology and brain alterations, measured by computed tomography (CT), was investigated in 44 depressed patients. Comparisons of ventricle-brain ratio (VBR) between "endogenous" vs. "nonendogenous" subgroups, classified by six distinct diagnostic systems, revealed no significant differences. The VBR and the width of the third ventricle correlated significantly with scores on the Brief Psychiatric Rating Scale, the Global Assessment Scale, the Bech-Rafaelsen Melancholia Scale, the Rating for Emotional Blunting, and the Scale for the Assessment of Negative Symptoms, but not with scores on the Hamilton Rating Scale for Depression and the Hamilton Rating Scale for Anxiety. Item analyses of the Bech-Rafaelsen Melancholia Scale revealed that retardation-related items were most significantly correlated with ventricular size. The wider diameter of the third ventricle in psychotic patients was associated with higher scores on retardation in the psychotic subgroup, whereas the greater distances of both Sylvian fissures showed no relationship to psychomotor retardation. No significant correlations were found between CT values and anxiety, suicidal impulses, somatic complaints, and sleep disturbances.     

Are transforming properties of the bovine papillomavirus E5 protein shared by E5 from high-risk human papillomavirus type 16?

The E5 proteins of bovine papillomavirus type 1 (BPV-1) and human papillomavirus type 16 (HPV-16) are small (44-83 amino acids), hydrophobic polypeptides that localize to membranes of the Golgi apparatus and endoplasmic reticulum, respectively. While the oncogenic properties of BPV-1 E5 have been characterized in detail, less is known about HPV-16 E5 due to its low expression in mammalian cells. Using codon-optimized HPV-16 E5 DNA, we have generated stable fibroblast cell lines that express equivalent levels of epitope-tagged BPV-1 and HPV-16 E5 proteins. In contrast to BPV-1 E5, HPV-16 E5 does not activate growth factor receptors, phosphoinositide 3-kinase or c-Src, and fails to induce focus formation, although it does promote anchorage-independent growth in soft agar. These variant activities are apparently unrelated to differences in intracellular localization of the E5 proteins since retargeting HPV-16 E5 to the Golgi apparatus does not induce focus formation.