Molecular subtyping is essential to infer tumor aggressiveness and predict prognosis. In practice, tumor profiling requires in-depth knowledge of bioinformatics tools involved in the processing and analysis of the generated data. Additionally, data incompatibility (e.g., microarray versus RNA sequencing data) and technical and uncharacterized biological variance between training and test data can pose challenges in classifying individual samples. In this article, we provide a roadmap for implementing bioinformatics frameworks for molecular profiling of human cancers in a clinical diagnostic setting. We describe a framework for integrating several methods for quality control, normalization, batch correction, classification and reporting, and develop a use case of the framework in breast cancer.

Supplementary Data from Differentiation Therapy Exerts Antitumor Effects on Stem-like Glioma Cells

PDF file - 1.1MB, Supplementary Table 1: list of 290 TCGA samples used in the study. Supplementary Table 2: list of miR-mRNA edges identified by CLR. Supplementary Table 3: list of putative direct miR-mRNA edges in the network. Supplementary Table 4: miR-mRNA edges identified as unique to each of the 4 molecular subtypes. (A) Classical (B) Neural (C) Proneural (D) Mesenchymal. Supplementary Table 5: miR and mRNA nodes in regions of copy number aberration in the sub-network corresponding to the 8 discriminatory miRs. Supplementary Table 6: survival analysis results for the 8 discriminatory miRs. Supplementary Table 7: multivariate Cox regression analysis in the TCGA dataset and in the 4 molecular subclasses. Supplementary Table 8: patients' characteristics (validation cohort, n=220). Supplementary Table 9: AQUA values (raw data) for miR-34a expression in Glioblastoma TMAs (validation cohort). Supplementary Table 10: list of transcription factors enriched in the PN subtype

PDF file - 854K, Supplementary Figure 1: miR-mRNA relationship (miR-34a; PDGFRA) detected by CLR in all GBM samples. Supplementary Figure 2: loss of p53 and Pten in murine neural stem cells and progenitor cells promotes the formation of proneural malignant gliomas. Supplementary Figure 3: expression levels of the subtype-specific miRNAs in p53/Pten-/- E13 embryonic neural stem cells infected with lentiviral miRNA precursors. Supplementary Figure 4: miR-34a expression in an independent cohort of GBM by in situ hybridization. Supplementary Figure 5: miR-34a levels in human and mouse proneural GBM. Supplementary Figure 6: miR-34a impairs tumor formation in orthotopic transplants. Supplementary Figure 7: miR-34a impairs the self-renewal of human proneural malignant spheroids. Supplementary Figure 8: miR-34a loss induces the expression of its oncogenic targets. Supplementary Figure 9: validation of PDGFRA as a direct target of miR-34a. Supplementary Figure 10: PDGFRA rescue on the self-renewal phenotype caused by miR-34a over-expression. Supplementary Figure 11: inverse correlation between the expression levels of miR-34a, PDGFRA and Smad4 in the GBM TCGA dataset. Supplementary Figure 12: validation of Smad4 as a direct target of miR-34a. Supplementary Figure 13: Smad4 rescue on the self-renewal phenotype caused by miR-34a over-expression

Background Spinal cord stimulation (SCS) is an effective method to treat neuropathic pain; however, it is challenging to compare different stimulation modalities in an individual patient, and thus, it is largely unknown which of the many available SCS modalities is most effective. Specifically, electrodes leading out through the skin would have to be consecutively connected to different, incompatible SCS devices and be tested over a time period of several weeks or even months. The risk of wound infections for such a study would be unacceptably high and blinding of the trial difficult. The PARS-trial seizes the capacity of a new type of wireless SCS device, which enables a blinded and systematic intra-patient comparison of different SCS modalities over extended time periods and without increasing wound infection rates. Methods The PARS-trial is designed as a double-blinded, randomized, and placebo-controlled multi-center crossover study. It will compare the clinical effectiveness of the three most relevant SCS paradigms in individual patients. The trial will recruit 60 patients suffering from intractable neuropathic pain of the lower extremities, who have been considered for SCS therapy and were already implanted with a wireless SCS device prior to study participation. Over a time period of 35 days, patients will be treated consecutively with three different SCS paradigms (“burst,” “1 kHz,” and “1.499 kHz”) and placebo stimulation. Each SCS paradigm will be applied for 5 days with a washout period of 70 h between stimulation cycles. The primary endpoint of the study is the level of pain self-assessment on the visual analogue scale after 5 days of SCS. Secondary, exploratory endpoints include self-assessment of pain quality (as determined by painDETECT questionnaire), quality of life (as determined by Quality of Life EQ-5D-5L questionnaire), anxiety perception (as determined by the Hospital Anxiety and Depression Scale), and physical restriction (as determined by the Oswestry Disability Index). Discussion Combining paresthesia-free SCS modalities with wireless SCS offers a unique opportunity for a blinded and systematic comparison of different SCS modalities in individual patients. This trial will advance our understanding of the clinical effectiveness of the most relevant SCS paradigms. Trial registration German Clinical Trials Register, DRKS00018929 . Registered on 14 January 2020.

MicroRNAs (miRs) are non-coding master regulators of transcriptome that could act as tumor suppressors (TSs) or oncogenes (oncomiRs). We aimed to systematically investigate the relevance of miRs as prognostic biomarkers in primary glioblastoma multiforme (GBM) treated with postoperative radio(chemo)therapy (PORT). For hypothesis generation, tumor miR expression by Agilent 8x15K human microRNA microarrays and survival data from 482 GBM patients of The Cancer Genome Atlas (TCGA cohort) were analyzed using Cox-PH models. Expression of candidate miRs with prognostic relevance (miR-221/222; miR-17-5p, miR-18a, miR-19b) was validated by qRT-PCR using Taqman technology on an independent validation cohort of GBM patients (n = 109) treated at Heidelberg University Hospital (HD cohort). In TCGA, 50 miRs showed significant association with survival. Among the top ranked prognostic miRs were members of the two miR families miR-221/222 and miR-17-92. Loss of miR-221/222 was correlated with improved prognosis in both cohorts (TCGA, HD) and was an independent prognostic marker in a multivariate analysis considering demographic characteristics (age, sex, Karnofsky performance index (KPI)), molecular markers (O-6-methylguanine-DNA methyltransferase (MGMT) methylation, IDH mutation status) and PORT as co-variables. The prognostic value of miR-17-92 family members was ambiguous and in part contradictory by direct comparison of the two cohorts, thus warranting further validation in larger prospective trials.

BACKGROUND:There is no standard approach to differentiate cerebral radiation necrosis from tumor recurrence and no standard treatment pathway for symptomatic lesions. In addition, reports on histology-proven radiation necrosis and the underlying pathophysiology are scarce and highly relevant. METHODS:Our monocentric, retrospective analysis included 21 histology-proven cerebral radiation necroses. Our study focused on 1) potential risk factors for the development of radiation necrosis, 2) radiologic and histopathologic features of individual necroses, and 3) the suitability of previously reported magnetic resonance imaging (MRI)-based methods to identify radiation necroses based on specific structural image features. RESULTS:Average time between radiation treatment and development of necrosis was 4.68 years (95% confidence interval, 0.19-9.55 years). Matching available MRI data sets with those of patients with tumor lesions, we compared specificity and sensitivity of 3 previously reported methods to identify radionecrosis based on imaging criteria. In our hands, none of these methods reached a sensitivity ≥70%. Radionecrosis presented with large edema and showed increased levels of cell proliferation, as inferred by Ki-67 staining. Surgical removal of radiation necrosis proved to be a safe approach with low permanent morbidity (<5%) and no mortality. CONCLUSIONS:Although the overall incidence of cerebral radiation necrosis is low, our data suggest an increasing incidence over the last 2 decades, which is likely associated with the use of stereotactic radiotherapy. There are no imaging standards to identify radiation necrosis on standard MRI with structural sequences. Surgical removal of radiation necrosis is associated with low morbidity and mortality.

Objective Concussions are the most frequent traumatic brain injuries. Yet, the socioeconomic impact of concussions remains unclear. Socioeconomic effects of concussions on working-age adults were studied on a population scale.Design This population-based, event time study uses administrative data as well as hospital and emergency room records for the population of Denmark.Setting We study all Danish patients, aged 20–59 years, who were treated at a public hospital or at an emergency room between 2003 and 2017 after suffering a concussion without other intracranial or extracranial injuries (n=55 424 unique individuals). None of the patients had a prior diagnosis of intracranial or extracranial injuries within the past 10 years leading up to the incident.Primary and secondary outcome measures As primary endpoint, we investigate the mean effect of concussion on annual salaried income within a 5-year period after trauma. In an exploratory analysis, we study whether the potential impact of concussion on annual salaried income is driven by patient age, education or economic cycle.Results Concussion was associated with an average change in annual salary income of −€1223 (95% CI: −€1540 to −905, p<0.001) corresponding to a salary change of −4.2% (95% CI: −5.2% to −3.1 %). People between 30 and 39 years and those without high school degrees suffered the largest salary decreases. Affected individuals leaving the workforce drove the main part of the decrease. Absolute annual effect sizes were countercyclical to the unemployment rate.Conclusions Concussions have a large and long-lasting impact on salary and employment of working-age adults on a nationwide scale.

OBJECTIVES:A new wireless spinal cord stimulation (SCS) technology, which was introduced in recent years, promises minimal invasive SCS as well as additional advantages such as a wide range of stimulation paradigms and 3-T magnetic resonance imaging (MRI) conditionality.MATERIALS AND METHODS:We prospectively evaluated 12 patients suffering from therapy-resistant neuropathic pain, who were implanted with a wireless SCS system from 2017 to 2019. Potential issues pertaining to handling and usability of the SCS device were evaluated from a patients' as well as from a surgeon's perspective.RESULTS:Mean follow-up was 228.0 days (95% CI, 20.0-518.0 days). We did not record any handling issues nor did we record any relevant local discomfort associated with the implanted SCS device. N = 3/12 patients reported discomfort from wearing the SCS antenna and one patient complained about a short battery life of the controller device. There were no reported incidents during 3-T MRI studies. After an average test period of 51.7 days (95% CI, 11.0-104.0 days), N = 9/12 patients (75%) had reached pain relief of 50% or more with an average pain relief (responders and partial responders) of 67.4% (95% CI, 50.0%-85.0%). On average, patients tested 2.2 different stimulation paradigms, with frequencies ranging from 60 Hz to 10 kHz, but there was no preferred stimulation paradigm.CONCLUSIONS:Minimal invasive implantation of wireless SCS systems was feasible and safe. The device offered a broader range of stimulation paradigms compared to conventional SCS devices, an allowed for a prolonged testing phase and continuous adjustment of SCS programs.

Purpose En bloc resection of retroperitoneal peripheral nerve sheath tumors (PNST) is advocated by a variety of surgical disciplines. Yet, microsurgical, nerve-sparing tumor resection might be better suited to improve symptoms and maintain neurological function, especially in cases where patients present with preoperative neurological deficits. However, neurosurgeons, versed in nerve-sparing techniques to remove PNST, are generally unfamiliar with the visceral approaches to retroperitoneal PNST. Methods We retrospectively evaluate a series of 16 patients suffering from retroperitoneal PNST. Patients were treated by a unique interdisciplinary approach, combining the visceral surgeon’s skills to navigate the complex anatomy of the retroperitoneal space and the neurosurgeon’s familiarity with microsurgical, nerve-sparing tumor removal. Specifically, we assess whether our interdisciplinary approach is suited to improve preoperative symptoms and maintain neurological function and study whether oncological outcome, surgical morbidity, and operative times are comparable to those reported for “classical” retroperitoneal PNST resection. In addition, we study two cases of suspected PNST that were diagnosed as malignant peripheral nerve sheath tumors (MPNST) after surgery. Results Total macroscopic tumor resection was achieved in 14/16 PNST patients. Mean intraoperative blood loss was 680.6 ml (95% CI, 194.3–1167.0 ml) and mean operative time was 162.5 min (95% CI, 121.6–203.4 min). We did not record any major postoperative surgical or neurological complications. A total of 8/11 patients with preoperative pain symptoms reported long-lasting improvement of their symptoms. In terms of oncological outcome, all patients that had been subjected to total tumor removal and for whom follow-up was available, were tumor-free after a mean follow-up of 761.9 days (95% CI, 97.6–1426.0 days). One of the two MPNST patients, who presented with tumor progress 15 months after initial surgery, was subjected to radical re-resection. Conclusions Interdisciplinary, nerve-sparing removal of retroperitoneal PNST is well suited to improve preoperative symptoms and maintain neurological function, while achieving an oncological outcome and a surgical morbidity similar to previously reported results for radical retroperitoneal PNST resection. Radical re-resection was feasible in a patient with post hoc MPNST diagnosis.

Background and objective: Stereotactical procedures require exact trajectory planning to avoid blood vessels in the trajectory path. Innovation in imaging and image recognition techniques have facilitated the automatic detection of blood vessels during the planning process and may improve patient safety in the future. To assess the feasibility of a vessel detection and warning system using currently available imaging and vessel segmentation techniques. Methods: Image data were acquired from post-contrast, isovolumetric T1-weighted sequences (T1CE) and time.-of-flight MR angiography at 3T or 7T from a total of nine subjects. Vessel segmentation by a combination of a vessel-enhancement filter with subsequent level-set segmentation was evaluated using three different methods (Vesselness, FastMarching and LevelSet) in 45 stereotactic trajectories. Segmentation results were compared to a gold-standard of manual segmentation performed jointly by two human experts. Results: The LevelSet method performed best with a mean interclass correlation coefficient (ICC) of 0.76 [0.73, 0.81] compared to the FastMarching method with ICC 0.70 [0.67, 0.73] respectively. The Vesselness algorithm achieved clearly inferior overall performance with a mean ICC of 0.56 [0.53, 0.59]. The differences in mean ICC between all segmentation methods were statistically significant (p < 0.001 with post-hoc p < 0.026). The LevelSet method performed likewise good in MPRAGE and 3T-TOF images and excellent in 7T-TOF image data. The negative predictive value (NPV) was very high (>97%) for all methods and modalities. Positive predictive values (PPV) were found in the overall range of 65-90% likewise depending on algorithm and modality. This pattern reflects the disposition of all segmentation methods - in case of misclassification - to produce preferentially false-positive than false-negative results. In a clinical setting, two to three potential collision warnings would be given per trajectory on average with a PPV of around 50%. Conclusions: It is feasible to integrate a clinically meaningful vessel detection and collision warning system into stereotactical planning software. Both, T1CE and MRA sequences are suitable as image data for such an application. (C) 2019 Elsevier B.V. All rights reserved.

Background Frame-based stereotactic biopsy (FBSB) is a minimally-invasive and effective procedure for the diagnosis of brain lesions and will likely gain clinical importance. Since FBSB procedures comprise a variety of imaging and sampling methods, it is necessary to compare the safety and effectiveness of individual techniques. Objective To assess the safety and effectiveness of FBSB using 1.5T iMRI as a one-stop procedure under general anesthesia without intraoperative histological examination. Methods In this single-center, retrospective analysis, 500 consecutive FBSBs using iMRI were compared to a historic control of 100 biopsies with traditional workflows (computed tomography (CT) with MRI image fusion). All procedures were performed under general anesthesia. Data on surgical procedures, pre- and postoperative neurologic patient status, complications and diagnostic yield were extracted from clinical records. Results Complication rates and diagnostic yield showed no significant differences between both groups. Mortality was 0.6%, 95% CI = [0.12%, 1.74%], in the iMRI and 0.0% [0.00%, 3.62%], in the control group with a morbidity of 5.4% [3.6%, 7.8%] and 6.0% [2.2%, 12.6%] and a diagnostic yield of 96.8% [94.9%, 98.2%] and 96.0% [90.1%, 98.9%]. Mean procedure duration was 124 [121, 127] minutes using iMRI and 112 [106, 118] minutes in the control group. Conclusion FBSB using 1.5T iMRI under general anesthesia is a safe and effective procedure and is equivalent to traditional stereotactic workflows with respect to complication rate and diagnostic yield.

OBJECTIVE: Implantation of deep brain stimulation (DBS) electrodes requires stereotactic imaging. Stereotactic magnetic resonance imaging (MRI) for DBS surgery has become more popular and intraoperative MRI scanners have become more available. We report on our cohort of movement disorder patients who underwent intraoperative stereotactic MRI-only DBS electrode implantation. METHODS: A review of our DBS database for eligible patients over a study period of 8 years was performed. Stereotactic accuracy was calculated as a directional error and the Euclidean distance between planned and controlled electrode positions. Number and choice of microelectrodes, procedural times and complications were documented. RESULTS: n = 86 surgeries in n = 81 patients with Parkinson's Disease (PD), essential tremor and dystonia were performed and n = 167 electrodes were implanted. Mean Euclidean distance between planned and controlled target was 2.1mm (+/- 0.6). The directional error showed that electrodes were implanted more medial (0.3mm +/- 0.9), posterior (0.5mm +/- 1.0) and inferior (0.6mm +/- 1.0) compared to plan. There were no significant differences for stereotactic accuracy between targets, hemispheres or order of implantation. No significant correlations between Euclidean distance and number of microelectrode tracts or volume of intracranial air were observed. N = 539 microelectrodes were applied. In 28.7% non-center trajectories were chosen. Length of tremor (-61 minutes) and PD (-121 minutes) surgeries could be reduced significantly over the course of the study period. N = 1 (1.2%) intracranial hemorrhage occurred. N = 1 (0.6%) electrode had to be repositioned for lack of clinical effect. CONCLUSION: Intraoperative stereotactic MRI for DBS surgery is feasible with high stereotactic accuracy and low rates of complication.

Chronic low back pain (CLBP) is challenging to treat. Minimal invasive neurostimulation therapies, such as subcutaneous peripheral nerve field stimulation (SPNS), improve pain relief and quality of life. The goal of the present study was to assess the usefulness, safety, and efficacy of SPNS in patients with CLBP. Twenty-six consecutive patients with CLBP were prospectively included in the study. For trial neurostimulation, two electrodes were implanted vertically at a depth of 1 cm into the subcutaneous tissue, ≤10 cm from the region of maximum pain. Trial neurostimulation was performed in all patients for 14 days. A successful outcome was defined as at least 50% pain relief. To monitor the effects of permanent neurostimulation, the Visual Analog Scale (VAS), the Oswestry Disability Index (ODI), and quality of life (EQ-5D-3L) were scored preoperatively and at 6-month and 24-month follow-ups. Thirteen patients responded to trial stimulation and had a permanent neurostimulator implanted. The use of pain medication, including opioid analgesics, was reduced in 92% of patients after 24 months. VAS, ODI, and EQ-5D-3L scores were significantly improved in these patients at the 24-month follow-up. The complication rate was 23% (3/13 patients). In non-responders, VAS and ODI at 24 months dropped significantly as well but the decrease was less pronounced compared to responders and had not led to a decrease in pain medication. SPNS is a novel, safe, and effective treatment for CLBP and may have advantages over interventional treatments including intrathecal therapy and spinal cord stimulation.

Background: Intraoperative magnetic resonance imaging (io-MRI) improves the extent of glioma resection. Due to the magnetic field, patients have to be covered with sterile drape and are then transferred into an io-MRI chamber, where ferromagnetic anaesthesia monitors and machines must be kept at distance and can only be applied with limitations. Despite the development of specific paramagnetic equipment for io-MRI use, this method is suspected to carry a higher risk for anaesthesiological and surgical complications. Particularly, serial draping and un-draping cycles as well as the extended surgery duration might increase the risk of perioperative infection. Objective: Given the importance of io-MRI for glioma surgery, the question regarding io-MRI safety needs to be answered. Methods: We prospectively evaluate the perioperative anaesthesiological and surgical complications for 516 cases of brain tumour surgery involving io-MRI (MRI cohort). As a control group, we evaluate a cohort of 610 cases of brain tumour surgery, performed without io-MRI (control group). Results: The io-MRI procedure (including draping/undraping, transfer to and from the MRI cabinet and io-MRI scan) significantly extended surgery, defined as skin to skin time, by 57min (SD=16min) (p0.01). Still, we show low and comparable rates of surgical complications in the MRI cohort and the control group. Postoperative haemorrhage (3.7% versus 3.0% in MRI cohort versus control group; p=0.49) and infections (2.2% versus 1.8% in MRI cohort versus control group; p=0.69) were not significantly different between both groups. No anaesthesiological disturbances were reported. Conclusion: Despite prolonged surgery and serial draping and un-draping cycles, io-MRI was not linked to higher rates of infections and postoperative haemorrhage in this study.

In spite of relentless efforts to devise new treatment strategies, primary glioblastomas invariably recur as aggressive, therapy-resistant relapses and patients rapidly succumb to these tumors. Many therapeutic agents are first tested in clinical trials involving recurrent glioblastomas. Remarkably, however, fundamental knowledge on the biology of recurrent glioblastoma is just slowly emerging. Here, we review current knowledge on recurrent glioblastoma and ask whether and how therapies change intra-tumor heterogeneity, molecular traits and growth pattern of glioblastoma, and to which extent this information can be exploited for therapeutic decision-making. We conclude that the ability to characterize and predict therapy-induced changes in recurrent glioblastoma will determine, whether, one day, glioblastoma can be contained in a state of chronic disease.

Extended tumour resection is imperative to improve the outcome of glioma patients but also carries the risk of increasing morbidity and thus, potentially, of decreasing the patient’s quality of life (QOL). In this pilot study, we evaluated how postoperative neurological and neuropsychological alterations impacted on QOL in patients who underwent glioma resection.

BACKGROUND:A 57-year-old man presented with recurrent episodes of dizziness, weakness of legs, and presyncope. The electrocardiogram showed a sinus bradycardia and recurrent sinus pauses.RESULTS:Cardiac evaluation revealed a normal left ventricular ejection fraction without ischemic, structural, or valvular heart disease. Pronounced limb weakness prompted neurological consultation. Cranial magnetic resonance imaging showed a large right-sided intracranial tumor adjacent to the medial sphenoid wing. Surgical removal of the tumor was accomplished successfully after application of a transient cardiac pacemaker, while decision upon permanent pacemaker implantation was postponed. Histopathology provided evidence of a meningothelial meningioma. Postoperative assessment displayed the absence of sinus node dysfunction after tumor removal.CONCLUSION:Careful differential diagnostic assessment of patients with symptomatic bradycardias needs to rule out reversible causes before implantation of permanent devices.

All-trans-retinoic acid (atRA), an autacoid derived from retinol (vitamin A), regulates energy balance and reduces adiposity. We show that energy status regulates atRA biosynthesis at the rate-limiting step, catalyzed by retinol dehydrogenases (RDH). Six h after re-feeding, Rdh1 expression decreased 80-90% in liver and brown adipose tissue and Rdh10 expression was decreased 45-63% in liver, pancreas, and kidney, all relative to mice fasted 16 h. atRA in the liver was decreased 44% 3 h after reduced Rdh expression. Oral gavage with glucose or injection with insulin decreased Rdh1 and Rdh10 mRNA 50% or greater in mouse liver. Removing serum from the medium of the human hepatoma cell line HepG2 increased Rdh10 and Rdh16 (human Rdh1 ortholog) mRNA expression 2-3-fold by 4 h, by increasing transcription and stabilizing mRNA. Insulin decreased Rdh10 and Rdh16 mRNA in HepG2 cells incubated in serum-free medium by inhibiting transcription and destabilizing mRNA. Insulin action required PI3K and Akt, which suppress FoxO1. Serum removal increased atRA biosynthesis 4-fold from retinol in HepG2 cells, whereas dominant-negative FoxO1 prevented the increase. Thus, energy status via insulin and FoxO1 regulate Rdh expression and atRA biosynthesis. These results reveal mechanisms for regulating atRA biosynthesis and the opposing effects of atRA and insulin on gluconeogenesis, and also suggest an interaction between atRA and insulin signaling related diseases, such as type II diabetes and cancer.