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Quantitative MRI allows to probe tissue properties by measuring relaxation times and may thus detect subtle changes in tissue composition. In this work we analyzed different relaxation times (T1, T2, T2* and T2′) and histological features in 321 samples that were acquired from 25 patients with newly diagnosed IDH wild-type glioma. Quantitative relaxation times before intravenous application of gadolinium-based contrast agent (GBCA), T1 relaxation time after GBCA as well as the relative difference between T1 relaxation times pre-to-post GBCA (T1rel) were compared with histopathologic features such as the presence of tumor cells, cell and vessel density, endogenous markers for hypoxia and cell proliferation. Image-guided stereotactic biopsy allowed for the attribution of each tissue specimen to its corresponding position in the respective relaxation time map. Compared to normal tissue, T1 and T2 relaxation times and T1rel were prolonged in samples containing tumor cells. The presence of vascular proliferates was associated with higher T1rel values. Immunopositivity for lactate dehydrogenase A (LDHA) involved slightly longer T1 relaxation times. However, low T2′ values, suggesting high amounts of deoxyhemoglobin, were found in samples with elevated vessel densities, but not in samples with increased immunopositivity for LDHA. Taken together, some of our observations were consistent with previous findings but the correlation of quantitative MRI and histologic parameters did not confirm all our pathophysiology-based assumptions.

Despite multidisciplinary local and systemic therapeutic approaches, the prognosis for most patients with brain metastases is still dismal. The role of adaptive and innate anti-tumor response including the Human Leukocyte Antigen (HLA) machinery of antigen presentation is still unclear. We present data on the HLA class II-chaperone molecule CD74 in brain metastases and its impact on the HLA peptidome complexity. We analyzed CD74 and HLA class II expression on tumor cells in a subset of 236 human brain metastases, primary tumors and peripheral metastases of different entities in association with clinical data including overall survival. Additionally, we assessed whole DNA methylome profiles including CD74 promoter methylation and differential methylation in 21 brain metastases. We analyzed the effects of a siRNA mediated CD74 knockdown on HLA-expression and HLA peptidome composition in a brain metastatic melanoma cell line. We observed that CD74 expression on tumor cells is a strong positive prognostic marker in brain metastasis patients and positively associated with tumor-infiltrating T-lymphocytes (TILs). Whole DNA methylome analysis suggested that CD74 tumor cell expression might be regulated epigenetically via CD74 promoter methylation. CD74high and TILhigh tumors displayed a differential DNA methylation pattern with highest enrichment scores for antigen processing and presentation. Furthermore, CD74 knockdown in vitro lead to a reduction of HLA class II peptidome complexity, while HLA class I peptidome remained unaffected. In summary, our results demonstrate that a functional HLA class II processing machinery in brain metastatic tumor cells, reflected by a high expression of CD74 and a complex tumor cell HLA peptidome, seems to be crucial for better patient prognosis.

Objective: We present the follow-up of a patient with a malignant undifferenciated neuroepithelial tumor of the medulla oblongata who regained a high quality of life after serious deterioration and a difficult decision for treatment Method: This 51-year-old woman complained of nausea without any [for full text, please go to the a.m. URL], 66. Jahrestagung der Deutschen Gesellschaft für Neurochirurgie (DGNC)

Background: Current pathological diagnostics include the analysis of (epi-)genetic alterations as well as oncogenic pathways. Deregulated mammalian target of rapamycin complex 1 (mTORC1) signaling has been implicated in a variety of cancers including malignant gliomas and is considered a promising target in cancer treatment. Monitoring of mTORC1 activity before and during inhibitor therapy is essential. The aim of our study is to provide a recommendation and report on pitfalls in the use of phospho-specific antibodies against mTORC1-targets phospho-RPS6 (Ser235/236; Ser240/244) and phospho-4EBP1 (Thr37/46) in formalin fixed, paraffin embedded material. Methods and findings: Primary, established cell lines and brain tumor tissue from routine diagnostics were assessed by immunocyto-, immunohistochemistry, immunofluorescent stainings and immunoblotting. For validation of results, immunoblotting experiments were performed. mTORC-pathway activation was pharmacologically inhibited by torin2 and rapamycin. Torin2 treatment led to a strong reduction of signal intensity and frequency of all tested antibodies. In contrast phospho-4EBP1 did not show considerable reduction in staining intensity after rapamycin treatment, while immunocytochemistry with both phospho-RPS6-specific antibodies showed a reduced signal compared to controls. Staining intensity of both phospho-RPS6-specific antibodies did not show considerable decrease in stability in a timeline from 0–230 minutes without tissue fixation, however we observed a strong decrease of staining intensity in phospho-4EBP1 after 30 minutes. Detection of phospho-signals was strongly dependent on tissue size and fixation gradient. mTORC1-signaling was significantly induced in glioblastomas although not restricted to cancer cells but also detectable in non-neoplastic cells. Conclusion: Here we provide a recommendation for phospho-specific immunohistochemistry for patient-orientated therapy decisions and monitoring treatment response.

The Sonic Hedgehog (SHH) pathway plays a central role in the developing mammalian CNS. In our study, we aimed to investigate the spatiotemporal SHH pathway expression pattern in human fetal brains. We analyzed 22 normal fetal brains for Shh, Patched, Smoothened, and Gli1-3 expression by immunohistochemistry. In the telencephalon, strongest expression of Shh, Smoothened, and Gli2 was found in the cortical plate (CP) and ventricular zone. Patched was strongly upregulated in the ventricular zone and Gli1 in the CP. In the cerebellum, SHH pathway members were strongly expressed in the external granular layer (EGL). SHH pathway members significantly decreased over time in the ventricular and subventricular zone and in the cerebellar EGL, while increasing levels were found in more superficial telencephalic layers. Our findings show that SHH pathway members are strongly expressed in areas important for proliferation and differentiation and indicate a temporal expression gradient in telencephalic and cerebellar layers probably due to decreased proliferation of progenitor cells and increased differentiation. Our data about the spatiotemporal expression of SHH pathway members in the developing human brain serves as a base for the understanding of both normal and pathological CNS development.

Der Sonic Hedgehog-Signalweg spielt eine zentrale Rolle in der embryonalen Entwicklung des zentralen Nervensystems. Beispielsweise reguliert Shh die Zellproliferation, die Migration sowie die Steuerung der Apoptose. Bislang existieren jedoch kaum Daten über die Verteilung von Sonic Hedgehog (Shh) und seinen Rezeptor- und Signalmolekülen im sich entwickelnden normalen humanen ZNS. Hieraus resultiert das Ziel der vorliegenden Arbeit, Shh selbst, sowie Patched, Smoothened, Gli1, Gli2 und Gli3 in ihrer räumlich-zeitlichen Verteilung im humanen ZNS darzustellen. Hierzu wurden 22 fetale Groß- und Kleinhirne zwischen der 12. und 28. Schwangerschaftswoche mittels Immunhistochemie untersucht. Alle sechs untersuchten Moleküle lassen sich in allen cerebellären, als auch neokortikalen Rindenschichten des sich entwickelnden fetalen Gehirns darstellen. In den fetalen Rindenschichten des Großhirnes lässt sich eine starke Immunreaktivitätsrate für alle untersuchten Signalmoleküle in den ventrikelnahen, proliferationsreichen Rindenschichten zeigen. Im zeitlichen Verlauf konnte ein Abfall (für Shh signifikant) der Immunreaktivitätsintensität des Signalweges in den proliferationsreichen Zonen und zugleich ein Anstieg in den ventrikelfernen Schichten beobachtet werden. Bei der Betrachtung der humanen Kleinhirnentwicklung zeichnet sich die Äußere Körnerzellschicht, als Proliferationszone, durch eine starke Immunreaktivitätsrate aller untersuchten Signalmoleküle aus. Im zeitlichen Verlauf zeigen sich im Kleinhirn ebenfalls eine Abnahme der Frequenz in der proliferationsreichen Rindenschicht und eine ansteigende Immunreaktivitätsintensität in der Inneren Körnerzellschicht. In der hier vorliegenden Arbeit kann sowohl in der Großhirnrinde, als auch in der Kleinhirn-rinde, ein Shh-Gradient dargestellt werden. Dies könnte somit Hinweis auf ein analoges, Gradienten gesteuertes Migrationsverhalten geben. Die Zunahme der Immunreaktivitäts-rate in der Äußeren Körnerzellschicht könnte als mitogene Wirkung von Shh auf die Körnerzellvorläufer gedeutet werden und würde somit bei unzureichender Inaktivierung, eine Grundlage für die Entstehung des Shh-Aktivierungsweg assoziierten Medulloblastoms bieten. Hieraus ergibt sich die enorme Wichtigkeit der genauen Kenntnis des Shh-Aktivierungsweges für das Verständnis der physiologischen Hirnentwicklung sowie pathologischer Veränderungen. The Sonic Hedgehog pathway plays an important role in cell proliferation, cell migration and apoptosis of the embryonic mammalian CNS. Until now, almost no data on the key players of the Sonic Hedgehog (Shh) pathway exists in the context of spatio-temporal patterns of the normal human fetal brain. Against this background we investigated 22 human fetal brain specimens (between the 12th and 28th week of gestation) for Shh, Patched, Smoothened, Gli1, Gli2 and Gli3 expression by immunohistochemistry. All six investigated proteins were expressed throughout the developing human fetal telencephalic and cerebellar cortical layers. In the telencephalon we were able to show a high expression in layers which have a high proliferation rate. During maturation we found a decrease of the signal in the ventricular- and subventricular zone, but also an increase of the immunoreactivity in the developing cortical plate. In the cerebellum the strongest signal was detected on cerebellar external granule layer cells. As brain development continued, there was a drop-off of the immunoreactivity at the external granule layer, but an ascent of the staining intensity in the inner granule layer. We were able to show that there is a Shh-gradient in the telencephalon, as well as in the cerebellum. This leads to the hypothesis, that there could be a gradient dependent migration behavior. The increased levels of immunoreactivity at the external granule layer could also be explained by the mitogenic influence of Shh on the granule precursor cells. The insufficient inhibition of the Shh pathway is likely to be one step in the development of the Shh-associated Medulloblastoma. The knowledge of these factors in the physiologically developing brain is therefore essential for both, the understanding of CNS pathology and the normal ontogenesis.