Matching Quantitative MRI Parameters with Histological Features of Treatment-Naïve IDH Wild-Type Glioma.

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.

Quantitative T1 mapping indicates tumor infiltration beyond the enhancing part of glioblastomas.

The aim of this study was to evaluate whether maps of quantitative T1 (qT1) differences induced by a gadolinium-based contrast agent (CA) are better suited than conventional T1-weighted (T1w) MR images for detecting infiltration inside and beyond the peritumoral edema of glioblastomas. Conventional T1w images and qT1 maps were obtained before and after gadolinium-based CA administration in 33 patients with glioblastoma before therapy. The following data were calculated: (i) absolute qT1-difference maps (qT1 pre-CA - qT1 post-CA), (ii) relative qT1-difference maps, (iii) absolute and (iv) relative differences of conventional T1w images acquired pre- and post-CA. The values of these four datasets were compared in four different regions: (a) the enhancing tumor, (b) the peritumoral edema, (c) a 5 mm zone around the pathology (defined as the sum of regions a and b), and (d) the contralateral normal appearing brain tissue. Additionally, absolute qT1-difference maps (displayed with linear gray scaling) were visually compared with respective conventional difference images. The enhancing tumor was visible both in the difference of conventional pre- and post-CA T1w images and in the absolute qT1-difference maps, whereas only the latter showed elevated values in the peritumoral edema and in some cases even beyond. Mean absolute qT1-difference values were significantly higher (P < 0.01) in the enhancing tumor (838 ± 210 ms), the peritumoral edema (123 ± 74 ms) and in the 5 mm zone around the pathology (81 ± 31 ms) than in normal appearing tissue (32 ± 35 ms). In summary, absolute qT1-difference maps-in contrast to the difference of T1w images-of untreated glioblastomas appear to be able to visualize CA leakage, and thus might indicate tumor cell infiltration in the edema region and beyond. Therefore, the absolute qT1-difference maps are potentially useful for treatment planning.

Delay in diagnosing patients with right-sided glioblastoma induced by hemispheric-specific clinical presentation.

PURPOSE: Cognitive functions are differentially represented in brain hemispheres. Aphasia is an "easy to recognize" symptom of diseases affecting the left side. In contrast, lesions in the right hemisphere cause subtle neuropsychological deficits such as neglect and anosognosia. We evaluated whether right-sided malignant brain tumors are on average larger at the time of first diagnosis as compared to left-sided tumors, and extrapolated the delay in diagnosing right-sided tumors compared to the left side. METHODS: All first-ever diagnosed glioblastoma (GBM) patients between 2005 and 2012 were identified using our hospital-based prospective research registry. Baseline data, information on initial clinical presentation and imaging findings (including tumor volume) were collected. Extrapolation of time since tumor initiation was based on an established gompertzian growth model. RESULTS: We included 173 patients. Mean age of the study population was 58 ± 13 years. Tumors located in the right hemisphere (n = 96) were larger as compared to tumors located in the left hemisphere (n = 77) (median 36.4 mL [interquartile range 13.0-56.0; minimum 0.2, maximum 140.0] vs. 17.2 mL [7.7-45.1 mL; 0.4, 105.2]; p = 0.011). Right-sided tumors grew longer than left-sided tumors (378 ± 95 days vs. 341 ± 74 days; p = 0.006). Initial neuropsychological symptoms differed depending on the affected hemisphere. CONCLUSION: Right-hemispheric symptoms appear to be less clinically conspicuous resulting in a delayed diagnosis of GBM, which might be improved by raising awareness for the corresponding neuropsychological deficits. Whether our findings have prognostic implications needs to be evaluated in future studies.

Low-Dose CCT to Exclude Contraindications to Lumbar Puncture : Benefits and Limitations.

BACKGROUND: Low-dose cranial computed tomography (LD-CCT) based on iterative reconstruction has been shown to have sufficient image quality to assess cerebrospinal fluid spaces (CSF) and midline structures but not to exclude subtle parenchymal pathologies. Patients without focal neurological deficits often undergo CCT before lumbar puncture (LP) to exclude contraindications to LP including brain herniation or increased CSF pressure. We performed LD-CCT to assess if image quality is appropriate for this indication. METHODS: A total of 58 LD-CCT (220 mA/120 kV) of patients before LP were retrospectively evaluated and compared to 79 normal standard dose cranial computed tomography (SD-CCT) (350 mA/120 kV). Iterative reconstruction used for both dose levels was increased by one factor for LD-CCT. We assessed the signal-to-noise (SNR) and contrast-to-noise ratio (CNR), the dose estimates and scored diagnostic image quality by two raters independently. Significance level was set at p < 0.05. RESULTS: The inner and outer CSF spaces except the sulci were equally well depicted by the LD-CCT and SD-CCT; however, depiction of the subtle density differences of the brain parenchyma and the sulci was significantly worse in the LD-CCT (p < 0.0001). The SNR in the gray matter (9.35 vs. 10.61, p < 0.05) and white matter (7.23 vs. 8.15, p < 0.001) were significantly lower in LD-CCT than in SD-CCT with significantly lower dose estimates (1.04 vs. 1.69 mSv, respectively p < 0.0001). CONCLUSION: The use of LD-CCT with a dose reduction of almost 50% is sufficient to exclude contraindications to LP; however, LD-CCT cannot exclude subtle parenchymal pathologies. Therefore, in patients with suspected parenchymal pathology, SD-CCT is still the method of choice.

Pre- and early postoperative GFAP serum levels in glioma and brain metastases.

SUBJECT: To date there is no established tumor marker for the clinical follow-up of glioblastoma, WHO grade IV, (GBM) which constitutes the most frequent and malignant primary brain tumor. However, since there is promising data that the serum glial fibrillary acidic protein (sGFAP) may serve as a biomarker for glial brain tumors, this prospective study aimed at evaluating the diagnostic relevance of perioperative changes in sGFAP levels for the assessment of residual glial tumor tissue in patients undergoing surgery of intracerebral tumors. METHODS: Serum GFAP was measured using an electrochemiluminometric immunoassay (ElecsysR GFAP prototype test, Roche Diagnostics, Penzberg/Germany) in 32 prospectively recruited patients between September 2009 and August 2010. Twenty-five were diagnosed with glioma and seven with brain metastases (BM). We assessed sGFAP levels prior to and at different time points during the early postoperative phase until patient discharge. RESULTS: There were only significant differences in the pre-operative sGFAP levels of patients with gliomas compared to BM (0.18 vs. 0.08 µg/l; p = 0.0198, Welch's t-Test). Even though there was an increase of sGFAP after surgery, there were no significant differences between glioma and BM patients at any other time point. Peak sGFAP levels where reached on postoperative day 1 followed by a slight decrease, but not reaching pre-operative levels until postop day 7. There was no significant correlation between postoperative glioma tumor volume and sGFAP levels in univariate analyses. CONCLUSION: According to our data sGFAP does not appear to be suitable to detect residual glioma tissue in the acute postoperative phase.

"Two is not enough" - Impact of the number of tissue samples obtained from stereotactic brain biopsies in suspected glioblastoma.

OBJECTIVE: Stereotactic procedures are performed in many neurosurgical departments in order to obtain tumor tissue from brain lesions for histopathological evaluation. Biopsies can be performed frame-guided and frame less. Some departments use a biopsy needle (cylinder probe), others a forceps for repetitive smaller tissue samples. Although the applied techniques are somehow different, it is still unclear how many tissue samples have to be taken to establish reliably a final diagnosis based on histopathological and genetic examinations. Only precise histopathological diagnosis results in adequate therapy. METHODS: We included 43 consecutive patients who underwent stereotactic biopsy of a suspected glioblastoma between 02/2013 and 07/2015. All patients showed contrast enhancing tumors in the MRI. The patients underwent stereotactic biopsy with the Leksell frame attached to their head. All stereotactic procedures were performed in the presence of a neuropathologist. Target and Entry Points were calculated with BrainLab iplan software (BrainLab iplan 1.0, Munich, Germany). First the two samples 5mm before the Target (pre-target) and the "Targetpoint" itself were analyzed (group 1), then a histopathological evaluation of all samples was performed (group 2). RESULTS: Mean number of extracted samples was 14. Using classical hematoxylin-eosin stainings, in group 1 histopathological diagnosis was correct in only 30 cases accounting for 73%. Contrariwise a final diagnosis was made in 100% in group 2. CONCLUSION: If only two tissue samples were evaluated in this group of patients with suspected glioblastoma, a correct diagnosis was possible in only 73% of the cases. We conclude that two samples are not enough to establish a final diagnosis even in a subgroup of suspected glioblastoma.

Positive influence of partial resection on overall survival of patients with overlapping glioblastomas.

OBJECTIVES: Patients with overlapping glioblastomas (former known as gliomatosis cerebri according to the 2007 WHO classification) have a poor prognosis. Most of the patients undergo biopsy to confirm histopathological diagnosis. Treatment comprises chemotherapy, radiation and combination of both. We determined whether resection of the contrast enhancing tumor parts leads to a prolonged survival. PATIENTS AND METHODS: We performed a retrospective analysis and included 31 patients with overlapping glioblastomas (OG) who showed WHO IV in the initial histopathological examination. All patients fulfilled criteria of overlapping glioblastomas in the MRI according to WHO criteria (3 or more lobes were affected). We evaluated Karnofsky performance score (KPS), gender, age, IDH-1_R132H status, MGMT promotor methylation status, proliferation index, postoperative therapy, biopsy vs. partial resection and extent of resection as possible factors affecting overall survival (OAS). A matched pair analysis was performed between the biopsy and resection group on basis of age, KPS and combined radio-chemotherapy. RESULTS: 10 Patients underwent resection of the contrast enhancing tumor parts, 21 patients underwent stereotactic biopsy. All included patients showed contrast enhancing lesions in the MRI. Median age was 61 years in the biopsy-group and 53 in the partial resection (PR) group. We found a significant correlation between OAS and age <50 (p=0.02). Median KPS was 80 in the STX group vs. 100 in the PR group. KPS above 80 was significantly associated with longer OAS (p=0.02). Median survival was 174days in the STX group compared to 446days in the PR group (p=0.05). Also the matched pair analysis showed significant p-values for resection. CONCLUSION: Partial resection might have a positive impact on overall survival of patients with overlapping glioblastomas (former known as gliomatosis cerebri), although the prognosis remains limited.

Serum concentrations of glial fibrillary acidic protein (GFAP) do not indicate tumor recurrence in patients with glioblastoma.

Recent studies identified serum concentrations of the astroglial protein glial fibrillary acidic protein (GFAP) to be indicative of glioblastoma (GBM) in patients with newly diagnosed space occupying cerebral mass lesions. Until now, no data is available whether GFAP serum concentrations decrease after first therapy and whether GFAP may be used as a predictor of survival and an indicator of tumor recurrence. In this prospective study, we included 44 patients with a single space occupying cerebral mass lesion suspicious for GBM. GBM was histopathologically proven in 33 cases. After initial therapy, patients were followed up until tumor recurrence (defined according to the RANO criteria) or death (maximum observation period 78 weeks). Blood was sampled on a regular basis, and GFAP serum levels were determined using an immunofluorescence assay. Prior to any intervention, 14 of the 33 GBM patients had elevated GFAP serum concentrations (median 0.25 µg/L, interquartile range 0.13-0.53), whereas only one out of 11 patients having other tumor entities revealed a slightly increased GFAP serum level (0.06 µg/L). Following surgery (i.e., biopsy, full or partial resection), all initially GFAP positive GBM patients showed decreased serum concentrations. During the follow-up period, we found a minimal GFAP increase in one patient only (0.04 µg/L; week 52), although 23 out of 31 available GBM patients developed tumor progression or died. No difference was found regarding the survival rate and the time to tumor recurrence between initially GFAP positive and GFAP negative GBM patients. In GBM patients, initially elevated GFAP serum concentrations decrease after the first diagnostic or therapeutic intervention. GFAP was not predictive for tumor recurrence.

Benefit and Complications of Frame-Based Stereotactic Biopsy in Old and Very Old Patients.

OBJECTIVE: Stereotactic biopsy is an everyday procedure implemented in numerous neurosurgical departments. The procedure is performed to obtain tumor tissue of unclear diagnosis. Going in hand with low complication rates and high diagnostic yield, stereotactic biopsies can be performed in adults and children likewise for histopathologic evaluation of lesions in eloquent localizations. However, little is known about whether aged patients do benefit from stereotactic biopsy or rather the therapy that is derived from histopathologic results. In this study, we therefore focused on old (80-84 years) and very old patients (85 years and older) to evaluate whether stereotactic biopsy should be performed leading to further therapy. We also assessed the complication rates of the procedure in this aged population. METHODS: We performed a retrospective analysis of our database and included all patients older than 80 years who underwent stereotactic biopsy at our department from October 2005 until May 2016. Forty-seven patients were included in this study. These patients were divided into 2 subgroups: group 1 consisted of patients from 80 to 84 years old and group 2 of patients aged 85 years and older. All patients underwent stereotactic biopsy to establish histopathologic diagnosis. We excluded patients who underwent cyst puncture or puncture of a hemorrhage because the procedure was not performed for diagnostic purposes. We assessed gender, neuroradiologic diagnosis, Karnofsky Performance Score (KPS), number of tissue samples taken, histopathologic diagnosis, localization, postoperative hemorrhage, modality of anesthesia anticoagulation, and further therapy. RESULTS: Group 1 consisted of 34 patients and group 2 of 13 patients. KPS was 80 and 70, respectively. A histopathologic diagnosis was possible in all but 1 patient. In group 1, 61.8% of the patients agreed to further postoperative therapy (radiation, 35.3%; chemotherapy, 11.8%; combined radiochemotherapy, 11.8%; complication that prevented therapy, 2.9%), as did 53.8% of the patients in group 2 (resection, 7.7%; radiation, 15.4%; combined radiochemotherapy, 30.7%). In group 1, 38.2% declined further therapy, as did 64.1% in group 2. CONCLUSIONS: Also in old and very old patients, a final histopathologic diagnosis should be established to provide adequate therapy. Our data show that most of these aged patients want to be treated.

Prognostic factors in recurrent glioblastoma patients treated with bevacizumab.

The value of bevacizumab (BEV) in recurrent glioblastoma is unclear. Imaging parameters and progression-free survival (PFS) are problematic endpoints. Few data exist on clinical factors influencing overall survival (OS) in unselected patients with recurrent glioblastoma exposed to BEV. We retrospectively analyzed 174 patients with recurrent glioblastoma treated with BEV at two German brain tumor centers. We evaluated general patient characteristics, MGMT status, pretreatment, concomitant oncologic treatment and overall survival. Karnofsky performance score, number of prior chemotherapies, number of prior recurrences and combined treatment with irinotecan (IRI) were significantly associated with OS in univariate analysis. We did not find differences in OS related to sex, age, histology, MGMT status, prior surgical treatment or number of prior radiotherapies. Combined treatment with IRI and higher KPS both remained significantly associated with prolonged survival in multivariate analysis, but patients receiving IRI co-treatment had less advanced disease. Grouping into clinically relevant categories revealed an OS of 16.9 months from start of BEV in patients with first recurrence and KPS ≥ 80 % (n = 25). In contrast, in patients with second recurrence and KPS < 80 %, OS was 3.6 months (n = 27). Our observational data support an early use of BEV in patients with good performance status. The benefit of co-treatment with IRI in our cohort seems to be the result of biased patient recruitment.

Prospective evaluation of serum glial fibrillary acidic protein (GFAP) as a diagnostic marker for glioblastoma.

Glioblastoma (GBM) is the most common malignant primary brain tumor. Although clinical presentation and brain imaging might be suggestive, histopathological evaluation by means of a brain biopsy is routinely performed to establish the diagnosis. A serum marker indicative of GBM may simplify the diagnostic work-up of patients suspected to having a brain tumor. We prospectively examined 113 patients with newly diagnosed single supratentorial or infratentorial space-occupying brain lesions. Glial fibrillary acidic protein (GFAP) levels were determined from venous blood samples via a prototype ELISA assay prior to any invasive procedures. Serum levels of GFAP were correlated with histopathological findings and MRI parameters. GFAP values were significantly higher in GBM patients (n = 33) compared to all other tumors (p < 0.001). A GFAP serum concentration of ≥0.01 µg/L revealed a sensitivity of 85 % and a specificity of 70 % for differentiating GBM from other entities. By applying a GFAP cut-off point of 0.20 µg/L, specificity was maximized (99 %), but sensitivity dropped to 27 %. In GBM patients, serum GFAP values were significantly correlated with tumor volume. GBM patients with high GFAP levels showed more in vivo GFAP expression as well as more necrosis and perilesional edema compared to GBM patients having low or non-detectable GFAP levels. GFAP serum concentrations differentiated between patients with GBM and patients with cerebral mass lesions of other entities with a moderate diagnostic accuracy. Serum GFAP levels in GBM patients were positively correlated with tumor volume and histopathological tumor characteristics.

Expression Profile of Sonic Hedgehog Pathway Members in the Developing Human Fetal Brain.

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.

Immunohistochemical Assessment of Phosphorylated mTORC1-Pathway Proteins in Human Brain Tumors.

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.

Improved visibility of brain tumors in synthetic MP-RAGE anatomies with pure T1 weighting.

Conventional MRI for brain tumor diagnosis employs T2 -weighted and contrast-enhanced T1 -weighted sequences. Non-enhanced T1 -weighted images provide improved anatomical details for precise tumor location, but reduced tumor-to-background contrast as elevated T1 and proton density (PD) values in tumor tissue affect the signal inversely. Radiofrequency (RF) coil inhomogeneities may further mask tumor and edema outlines. To overcome this problem, the aims of this work were to employ quantitative MRI techniques to create purely T1 -weighted synthetic anatomies which can be expected to yield improved tissue and tumor-to-background contrasts, to compare the quality of conventional and synthetic anatomies, and to investigate optical contrast and visibility of brain tumors and edema in synthetic anatomies. Conventional magnetization-prepared rapid acquisition of gradient echoes (MP-RAGE) anatomies and maps of T1 , PD and RF coil profiles were acquired in comparable and clinically feasible times. Three synthetic MP-RAGE anatomies (PD T1 weighting both with and without RF bias; pure T1 weighting) were calculated for healthy subjects and 32 patients with brain tumors. In healthy subjects, the PD T1 -weighted synthetic anatomies with RF bias precisely matched the conventional anatomies, yielding high signal-to-noise (SNR) and contrast-to-noise (CNR) ratios. Pure T1 weighting yielded lower SNR, but high CNR, because of increased optical contrasts. In patients with brain tumors, synthetic anatomies with pure T1 weighting yielded significant increases in optical contrast and improved visibility of tumor and edema in comparison with anatomies reflecting conventional T1 contrasts. In summary, the optimized purely T1 -weighted synthetic anatomy with an isotropic resolution of 1 mm, as proposed in this work, considerably enhances optical contrast and visibility of brain tumors and edema.

Netrin-1 expression is an independent prognostic factor for poor patient survival in brain metastases.

The multifunctional molecule netrin-1 is upregulated in various malignancies and has recently been presented as a major general player in tumorigenesis leading to tumor progression and maintenance in various animal models. However, there is still a lack of clinico-epidemiological data related to netrin-1 expression. Therefore, the aim of our study was to elucidate the association of netrin-1 expression and patient survival in brain metastases since those constitute one of the most limiting factors for patient prognosis. We investigated 104 brain metastases cases for netrin-1 expression using in-situ hybridization and immunohistochemistry with regard to clinical parameters such as patient survival and MRI data. Our data show that netrin-1 is strongly upregulated in most cancer subtypes. Univariate analyses revealed netrin-1 expression as a significant factor associated with poor patient survival in the total cohort of brain metastasis patients and in sub-entities such as non-small cell lung carcinomas. Interestingly, many cancer samples showed a strong nuclear netrin-1 signal which was recently linked to a truncated netrin-1 variant that enhances tumor growth. Nuclear netrin-1 expression was associated with poor patient survival in univariate as well as in multivariate analyses. Our data indicate both total and nuclear netrin-1 expression as prognostic factors in brain metastases patients in contrast to other prognostic markers in oncology such as patient age, number of brain metastases or Ki67 proliferation index. Therefore, nuclear netrin-1 expression constitutes one of the first reported molecular biomarkers for patient survival in brain metastases. Furthermore, netrin-1 may constitute a promising target for future anti-cancer treatment approaches in brain metastases.