Rapid, label-free classification of glioblastoma differentiation status combining confocal Raman spectroscopy and machine learning.

Label-free identification of tumor cells using spectroscopic assays has emerged as a technological innovation with a proven ability for rapid implementation in clinical care. Machine learning facilitates the optimization of processing and interpretation of extensive data, such as various spectroscopy data obtained from surgical samples. The here-described preclinical work investigates the potential of machine learning algorithms combining confocal Raman spectroscopy to distinguish non-differentiated glioblastoma cells and their respective isogenic differentiated phenotype by means of confocal ultra-rapid measurements. For this purpose, we measured and correlated modalities of 1146 intracellular single-point measurements and sustainingly clustered cell components to predict tumor stem cell existence. By further narrowing a few selected peaks, we found indicative evidence that using our computational imaging technology is a powerful approach to detect tumor stem cells in vitro with an accuracy of 91.7% in distinct cell compartments, mainly because of greater lipid content and putative different protein structures. We also demonstrate that the presented technology can overcome intra- and intertumoral cellular heterogeneity of our disease models, verifying the elevated physiological relevance of our applied disease modeling technology despite intracellular noise limitations for future translational evaluation.

Concept of a fully-implantable system to monitor tumor recurrence.

Current treatment for glioblastoma includes tumor resection followed by radiation, chemotherapy, and periodic post-operative examinations. Despite combination therapies, patients face a poor prognosis and eventual recurrence, which often occurs at the resection site. With standard MRI imaging surveillance, histologic changes may be overlooked or misinterpreted, leading to erroneous conclusions about the course of adjuvant therapy and subsequent interventions. To address these challenges, we propose an implantable system for accurate continuous recurrence monitoring that employs optical sensing of fluorescently labeled cancer cells and is implanted in the resection cavity during the final stage of tumor resection. We demonstrate the feasibility of the sensing principle using miniaturized system components, optical tissue phantoms, and porcine brain tissue in a series of experimental trials. Subsequently, the system electronics are extended to include circuitry for wireless energy transfer and power management and verified through electromagnetic field, circuit simulations and test of an evaluation board. Finally, a holistic conceptual system design is presented and visualized. This novel approach to monitor glioblastoma patients is intended to early detect recurrent cancerous tissue and enable personalization and optimization of therapy thus potentially improving overall prognosis.

Profile and Prognosis of Spontaneous Lobar Intracerebral Hemorrhage: Comparison of 6-month Survival with STICH II and the MISTIE III Lobar Hemorrhage Subset.

BACKGROUND: Randomized trials on spontaneous lobar intracerebral hemorrhage (ICH) provided no convincing evidence of the superiority of surgical treatment. Since recruitment in the trials was under the premise of equipoise, a selection bias toward patients who did not need surgery or were in hopeless condition must be suspected. The aim of the actual analysis was to compare outcome and patient profile of an unselected hospital series with recent randomized trials and to develop a prognostic model. METHODS: Of 821 patients with spontaneous ICH managed at the neurosurgical department of the University Hospital Düsseldorf between 2013 and 2018, 159 had lobar bleedings. Patient characteristics, hematoma volume, treatment modality, and 6-month survival were compared with STICH II and the subset of lobar hemorrhage in the MISTIE III trial. In addition, a prognostic model for 6-month survival in our patients was developed using a random forest classifier. RESULTS: One hundred and seven patients were managed by surgical evacuation of the hematoma and 52 without surgical evacuation. Median hemorrhage volume in our surgical cohort was 66 and 42 mL in the conservative cohort, compared with 38 and 36 mL in the STICH II trial, and 46 and 47 mL in the surgical and conservative MISTIE III lobar hemorrhage subset. Median initial Glasgow Coma Scale (GCS) score was 12 in our surgical group and 11 in the conservative group, compared with 13 in the STICH II cohorts and 12 in the MISTIE III lobar hemorrhage subset. Median age in our surgical and conservative cohorts was 73 and 74 years, respectively, compared with 65 years in both STICH II cohorts and 68 years in the MISTIE II subsets. Twenty-nine percent of our surgical cohort and 55% of our conservatively managed patients deceased within the first 6 months, compared with 18 and 24%, respectively, in STICH II and 17 and 24% in the MISTIE III subset. Our prognostic model identified large hemorrhage volumes and low admission GCS score as main unfavorable prognostic factors for 6-month survival. The random forest classifier achieved a predictive accuracy of 78% and an area under curve (AUC)- value of 88% regarding survival at 6 months, on a test set independent of the training set. CONCLUSIONS: In comparison with our surgical group, the STICH II and MISTIE III cohorts, recruited under the premise of physician equipoise, underrepresented patients with large ICHs. The cohorts in the randomized trials were therefore biased toward patients with a favorable perspective under conservative management. Initial hematoma volume and admission GCS were the main prognostic factors in our patients.

Testing the Stability of Drug Resistance on Cryopreserved, Gene-Engineered Human Induced Pluripotent Stem Cells.

Human induced pluripotent stem cells (hiPSCs) have emerged as a powerful tool for in vitro modelling of diseases with broad application in drug development or toxicology testing. These assays usually require large quantities of hiPSC, which can entail long-term storage via cryopreservation of the same cell charges. However, it is essential that cryopreservation does not oppose durable changes on the cells. In this project, we characterize one parameter of functionality of one that is well established in the field, in a different research context, an applied hiPSC line (iPS11), namely their resistance to a medium size library of chemo interventions (>160 drugs). We demonstrate that cells, before and after cryopreservation, do not change their relative overall drug response phenotypes, as defined by identification of the top 20 interventions causing dose-dependent reduction of cell growth. Importantly, also frozen cells that are exogenously enforced for stable overexpression of oncogenes myelocytomatosis (cMYC) or tumor protein 53 mutation (TP53R175H), respectively, are not changed in their relative top 20 drugs response compared to their non-frozen counterparts. Taken together, our results support iPSCs as a reliable in vitro platform for in vitro pharmacology, further raising hopes that this technology supports biomarker-associated drug development. Given the general debate on ethical and economic problems associated with the reproducibly crisis in biomedicine, our results may be of interest to a wider audience beyond stem cell research.

Different Calculation Strategies Are Congruent in Determining Chemotherapy Resistance of Brain Tumors In Vitro.

In cancer pharmacology, a drug candidate's therapeutic potential is typically expressed as its ability to suppress cell growth. Different methods in assessing the cell phenotype and calculating the drug effect have been established. However, inconsistencies in drug response outcomes have been reported, and it is still unclear whether and to what extent the choice of data post-processing methods is responsible for that. Studies that systematically examine these questions are rare. Here, we compare three established calculation methods on a collection of nine in vitro models of glioblastoma, exposed to a library of 231 clinical drugs. The therapeutic potential of the drugs is determined on the growth curves, using growth inhibition 50% (GI50) and point-of-departure (PoD) as the criteria. An effect is detected on 36% of the drugs when relying on GI50 and on 27% when using PoD. For the area under the curve (AUC), a threshold of 9.5 or 10 could be set to discriminate between the drugs with and without an effect. GI50, PoD, and AUC are highly correlated. The ranking of substances by different criteria varies somewhat, but the group of the top 20 substances according to one criterion typically includes 17-19 top candidates according to another. In addition to generating preclinical values with high clinical potential, we present off-target appreciation of top substance predictions by interrogating the drug response data of non-cancer cells in our calculation technology.

Association between health insurance status and malignant glioma.

BACKGROUND: Prior studies have suggested an association between patient socioeconomic status and brain tumors. In the present study we attempt to indirectly validate the findings, using health insurance status as a proxy for socioeconomic status. METHODS: There are 2 types of health insurance in Germany: statutory and private. Owing to regulations, low- and middle-income residents are typically statutory insured, whereas high-income residents have the option of choosing a private insurance. We compared the frequencies of privately insured patients suffering from malignant neoplasms of the brain with the corresponding frequencies among other neurosurgical patients at our hospital and among the German population. To correct for age, sex, and distance from the hospital, we included these variables as predictors in logistic and binomial regression. RESULTS: A significant association (odds ratio [OR] = 1.59, CI = 1.45-1.74, P < .001) between health insurance status and brain tumors was found. The association is independent of patients' sex or age. Whereas privately insured patients generally tend to come from farther away, such a relationship was not observed for patients suffering from brain tumors. Comparing the out of house and in-house brain tumor patients showed no selection bias on our side. CONCLUSION: Previous studies have found that people with a higher income, level of education, or socioeconomic status are more likely to suffer from malignant brain tumors. Our findings are in line with these studies. Although the reason behind the association remains unclear, the probability that our results are due to some random effect in the data is extremely low.

Robot technology identifies a Parkinsonian therapeutics repurpose to target stem cells of glioblastoma.

Aim: Glioblastoma is a heterogeneous lethal disease, regulated by a stem-cell hierarchy and the neurotransmitter microenvironment. The identification of chemotherapies targeting individual cancer stem cells is a clinical need. Methodology: A robotic workstation was programmed to perform a drug concentration to cell-growth analysis on an in vitro model of glioblastoma stem cells (GSCs). Mode-of-action analysis of the selected top substance was performed with manual repetition assays and acquisition of further parameters. Results: We identified 22 therapeutic potential substances. Three suggested a repurpose potential of neurotransmitter signal-modulating agents to target GSCs, out of which the Parkinson's therapeutic trihexyphenidyl was most effective. Manual repetition assays and initial mode of action characterization revealed suppression of cell proliferation, cell cycle and survival. Conclusion: Anti-neurotransmitter signaling directed therapy has potential to target GSCs. We established a drug testing facility that is able to define a mid-scale chemo responsome of in vitro cancer models, possibly also suitable for other cell systems.

Quantification of PpIX-fluorescence of cerebral metastases: a pilot study.

5-ALA fluorescence-guided surgery (FGS) is a major advance in neuro-oncological surgery. So far, Protoporphyrin IX (PpIX)-fluorescence has been observed in about half of cerebral metastases resected with routinely equipped microscopes during 5-ALA FGS. The aim of the present pilot study was to quantify PpIX-induced fluorescence of cerebral metastases with a spectrometer. We hypothesize that non-fluorescing metastases under the operating microscope may have spectrometrically measurable levels of fluorescence. A second aim was to analyze correlations between quantified 5-ALA fluorescence and histology or primary tumor type, respectively. Standard FGS was performed in all patients. The fluorescence intensity of the metastasis was semi-quantitatively determined in vivo by a senior surgeon using a special surgical microscope equipped for FGS. A systematic spectrometric ex vivo evaluation of tumor specimens and PpIX-induced fluorescence was performed using a spectrometer connected by optic fibers to a handheld probe. Quantification of 5-ALA-derived fluorescence was measured in a standardized manner with direct contact between mini-spectrometer and metastasis. The difference between the maximum PpIX-fluorescence at 635 nm and the baseline fluorescence was defined as the PpIX fluorescence intensity of the metastasis and given in arbitrary units (AU). Diagnosis of a cerebral metastasis was confirmed by histopathological analysis. A total of 29 patients with cerebral metastases were included. According to neuropathological analysis, 11 patients suffered from non-small cell lung cancer, 10 patients from breast cancer, 6 patients from cancer originating in the gastro-intestinal tract, 1 patient suffered from a malignant melanoma and one patient from renal cancer. The mean age was 63 years (37-81 years). 15 patients were female, 14 patients male. 13 cerebral metastases were considered as ALA-positive by the surgeon. In nine metastases, 5-ALA fluorescence was not visible to the naked eye and could only be detected using the spectrometer. The threshold for an ALA signal rated as "positive" by the surgeon was PpIX fluorescence above 1.1 × 106 AU. The mean PpIX fluorescence of all analyzed cerebral metastases was 1.29 × 106 ± 0.23 × 106 AU. After quantification, we observed a significant difference between the mean 5-ALA-derived fluorescence in NSCLC and breast cancer metastases (Mean Diff: - 1.2 × 106; 95% CI of difference: - 2.2 × 106 to - 0.15 × 106; Sidák-adjusted p = 0.026). In our present pilot series, about half of cerebral metastases showed a 5-ALA fluorescence invisible to the naked eye. Over 50% of these non-fluorescent metastases show a residual 5-ALA fluorescence which can be detected and quantified using a spectrometer. Moreover, the quantified 5-ALA signal significantly differed with respect to the primary tumor of the corresponding cerebral metastasis. Further studies should evaluate the predictive value of the 5-ALA signal and if a quantified 5-ALA signal enables a reliable intraoperative differentiation between residual tumor tissue and edematous brain-in particular in metastases with a residual fluorescence signal invisible to the naked eye.

Predictors for a further local in-brain progression after re-craniotomy of locally recurrent cerebral metastases.

Treatment of recurrent cerebral metastases is an emerging challenge due to the high local failure rate after surgery or radiosurgery and the improved prognosis of patients with malignancies. A total of 36 patients with 37 metastases who underwent surgery for a local in-brain progression of a cerebral metastasis after previous metastasectomy were retrospectively analyzed. Degree of surgical resection on an early postoperative MRI within 72 h after surgery was correlated with the local in-brain progression rate and overall survival. Complete surgical resection of locally recurrent cerebral metastases as confirmed by early postoperative MRI could only be achieved in 37.8%. Detection of residual tumor tissue on an early MRI following recurrent metastasis surgery correlated with further local in-brain progression when defining a significance level of p = 0.05 but not after Sidák or Bonferroni significance level correction for multiple testing: However, definite local tumor control could finally be achieved in 91.9% after adjuvant therapy. Overall survival after recurrent metastasectomy was significantly higher as predicted by diagnosis-specific graded prognostic assessment (12.9 ± 2.3 vs. 8.4 ± 0.7 months; p < 0.0001). However, our series involved a limited number of heterogeneous patients. A larger, prospective, and controlled study is required. Considering the adequate local tumor control achieved in the vast majority of patients, surgery of recurrent metastases may represent one option in a multi-modal treatment approach of patients suffering from locally recurrent cerebral metastases.

Decompressive hemicraniectomy for malignant middle cerebral artery infarction including patients with additional involvement of the anterior and/or posterior cerebral artery territory-outcome analysis and definition of prognostic factors.

BACKGROUND: According to current evidence, adding decompressive craniectomy (DC) to best medical therapy reduces case fatality rate of malignant middle cerebral artery infarction by 50-75%. There is currently little information available regarding the outcome of subgroups, in particular of patients with extensive infarctions exceeding the territory of the middle cerebral artery. METHODS: The records of 101 patients with large hemispheric infarctions undergoing DC were retrospectively reviewed. Twenty-seven patients had additional ACA and/or PCA infarcts. Sequential CTs were used for postoperative follow-up. Intracranial pressure (ICP) was monitored via a ventricular catheter in comatose patients. The main aim of treatment was to keep midline shift below 10 mm and ICP below 20 mmHg. If midline shift increased despite preceding DC, repeat surgery with removal of clearly necrotic tissue was considered. For the current analysis, Glasgow Coma Scale (GCS) at 14 days and modified Rankin Scale (mRS) at 3 months were used as outcome parameters. mRS 2 and 3 were defined as "moderate disability", mRS 4 as "severe disability", and mRS 5 and 6 as "poor outcome". These outcome parameters were correlated to age, gender, side, vascular territory, and time delay after stroke, GCS at the time of decompression, maximum ICP, maximum midline shift, and delay of maximum shift. RESULTS: The median age of the 39 female and 62 male patients was 56 years (range, 5-79 years). Overall, 12 patients died in the acute stage (11.9%). Twenty-three (22.8%) patients recovered to moderate disability at 3 months (mRS ≤ 3), 45 (44.6%) to severe disability and 33 (32.6%) suffered a poor outcome (mRS 5 or 6). Twenty patients (19.8%) required additional necrosectomy due to secondary increasing midline shift and/or intracranial hypertension. Patients recovering to moderate disability at 3 months were in the median 10 years younger than patients with less favorable outcome (P < 0.001) and had a higher GCS prior to surgery (P < 0.001). Eleven of the 27 patients with infarctions exceeding the MCA territory needed secondary surgery, indicating a higher necrosectomy rate as for isolated MCA infarction. At 3 months, the distribution of the outcomes in terms of mRS was comparable between the patients suffering from extended infarctions and patients having isolated MCA stroke. Infarctions exceeding the territory of the middle cerebral artery were seen in 30% of the group recovering to moderate disability and thus as frequent as in the groups suffering a less favorable outcome. CONCLUSIONS: Intensified postoperative management including possible secondary decompression with necrosectomy may further reduce case fatality rate of patients with large hemispheric infarction. Age above 60 years and severely reduced level of consciousness are the most significant factors heralding unfavorable recovery. Patients suffering infarctions exceeding the MCA territory have a comparable chance of favorable recovery as patients with isolated MCA infarction.

Fluorescence Behavior and Dural Infiltration of Meningioma Analyzed by 5-Aminolevulinic Acid-Based Fluorescence: Operating Microscope Versus Mini-Spectrometer.

OBJECTIVE: To compare fluorescence intensity of tumor specimens, as measured by a fluorescence-guided surgery microscope and a spectrometer, to evaluate tumor infiltration of dura mater around meningiomas with help of these 2 different 5-aminolevulinic acid (5-ALA)-based fluorescence tools, and to correlate fluorescence intensity with histopathologic data. MATERIAL AND METHODS: In a clinical series, meningiomas were resected by 5-ALA fluorescence-guided surgery. Fluorescence intensity was semiquantitatively rated by the surgeon at predefined points. Biopsies were harvested and fluorescence intensity measured by a spectrometer and histopathologically analyzed. Sampling was realized at the level of the dura in a centrifugal direction. RESULTS: A total of 104 biopsies (n = 13 tumors) were analyzed. Specificity and sensitivity of the microscope were 0.96 and 0.53 and of the spectrometer 0.95 and 0.93, respectively. Fluorescence intensity as measured by the spectrometer was correlated to histologically confirmed tumor burden. In a centrifugal direction, tumor burden and fluorescence intensity continuously decreased (along the dural tail). Below a threshold value of 639 arbitrary units no tumor was histologically detectable. CONCLUSIONS: At the level of the dura the spectrometer was highly sensitive for detection of meningioma cells. The surgical microscope showed false negative results and missed residual tumor cells in more than one half of the cases. The complementary use of both fluorescence tools may improve resection quality.

Quality of Life After Surgery for Cerebral Cavernoma: Brainstem Versus Nonbrainstem Location.

OBJECTIVE: We sought to analyze long-term outcome and quality of life after surgery of cerebral cavernomas (CCs) with special regard to localization (brainstem vs. nonbrainstem). METHODS: We conducted a retrospective study in a tertiary care center (2000-2010). Clinical charts were analyzed. Health-related quality of life (QoL) was evaluated with the Short Form-36 questionnaire. RESULTS: The study included 60 patients (21 male, 39 female, mean age 39.8 years). The distribution was 67% supratentorial, 7% cerebellar, and 26% brainstem (BS). In the BS group, 87.5% had a preoperative deficit versus 18.2% in the nonbrainstem group (NBS). Operative neurologic morbidity was 31.3% for BS versus 11.4% for NBS. After mean follow-up of 43 months, neurologic status was better or the same as compared with the preoperative status in 75% of BS and all of NBS. SF-36 showed no significant differences between all cavernoma patients compared with a normative healthy population except for a better "pain" score. Subgroup analysis found the same results when comparing NBS with the normative population. Comparison of BS versus norm and NBS, respectively, showed worse scores for BS in physical but not mental health (P ≤ 0.01). CONCLUSION: Clinical outcome was different depending on location: NBS recovered the same neurologic status as preoperatively and showed better QoL in physical health and lower working inability than BS. Surprisingly, there was no difference in mental health. Moreover, QoL of the operated cavernoma population after long-term follow-up did not differ from the norm. We conclude that surgery of cavernomas even in eloquent areas may result in favorable outcome and high patient satisfaction.

5-ALA fluorescence of cerebral metastases and its impact for the local-in-brain progression.

Aim of the present study was to analyze the oncological impact of 5-ALA fluorescence of cerebral metastases. A retrospective analysis was performed for 84 patients who underwent 5-ALA fluorescence-guided surgery of a cerebral metastasis. Dichotomized fluorescence behavior was correlated to the histopathological subtype and primary site of the metastases, the degree of surgical resection on an early postoperative MRI within 72 hours after surgery, the local in-brain-progression rate and the overall survival. 34/84 metastases (40.5%) showed either strong or faint and 50 metastases (59.5%) no 5-ALA derived fluorescence. Neither the primary site of the cerebral metastases nor their subtype correlated with fluorescence behavior. The dichotomized 5-ALA fluorescence (yes vs. no) had no statistical influence on the degree of surgical resection. Local in-brain progression within or at the border of the resection cavity was observed in 26 patients (30.9%). A significant correlation between 5-ALA fluorescence and local in-brain-progression rate was observed and patients with 5-ALA-negative metastases had a significant higher risk of local recurrence compared to patients with 5-ALA positive metastases. After exclusion of the 20 patients without any form of adjuvant radiation therapy, there was a trend towards a relation of the 5-ALA behavior on the local recurrence rate and the time to local recurrence, although results did not reach significance anymore. Absence of 5-ALA-induced fluorescence may be a risk factor for local in-brain-progression but did not influence the mean overall survival. Therefore, the dichotomized 5-ALA fluorescence pattern might be an indicator for a more aggressive tumor.

A new possibility to assess the perioperative walking capacity using a global positioning system in neurosurgical spine patients: a feasibility study.

PURPOSE: It is well established that the perioperative course in terms of patient satisfaction, neurological function and quality of life, is assessed by monitoring the walking capacity. This examination method is affected by several primary and secondary influences. Therefore, we performed a feasibility study to investigate the possibility of assessing the perioperative walking capacity using a global positioning system (GPS) in neurosurgical spine patients. A step was undertaken to generate objective and reliable data for monitoring control with a cost-effective and easy-to-use measurement tool. METHODS: Everyday life activities of four patients were measured by using a GPS-capable mobile phone (a week preoperatively, FU 3 months later). Our custom-made software for Android systems continuously records the position- and movement-data of all subjects during the day at 1 s intervals. The position date were smoothed and checked for plausibility. This semi-automated process was followed by determining the total distance walked (TL), the average distance (AL), the average walking speed (V) and the total walking duration per day (T). Additionally, we are able to explore the measuring inaccuracy. RESULTS: In three patients, nearly all parameters were increased in the follow-up examination (TL: 650.76, 972.63, and 269.07%. AL: 1213.83, 3117.89, and 72.23%. V: 78.62, -15.50, and 8.54%. T: 148.18, 4089.56, and 9.08%). In one patient, we documented a different motion pattern (TL: -54.37%, AL: -31.56%, V: -9.20%, T increased: 507.91%) due to residual limitations after suffering a heart attack. CONCLUSION: In this feasibility study, we demonstrated that this tool is able to measure the perioperative mobility and walking-capability. The certainty of data is dependent on the patients' compliance. The measuring method is used as a low cost, easily accessible, and easy-to-use technique, which seems to be superior to common methods like a treadmill-tests or walking tests. Nevertheless, these results are still to prove in upcoming analysis.

Microsurgical resection of Spetzler-Martin grades 1 and 2 unruptured brain arteriovenous malformations results in lower long-term morbidity and loss of quality-adjusted life-years (QALY) than conservative management--results of a single group series.

BACKGROUND: The therapeutic benefits of microsurgery for unruptured brain AVM remain unclear. METHODS: A series of 97 microsurgically resected unruptured brain AVM was analyzed in terms of postoperative morbidity and lifetime loss of quality-adjusted life-years (QALY). For comparison, the natural risk of becoming disabled was modeled on the basis of published data. RESULTS: Discharge morbidity was recorded in 11 of the 69 of Spetzler-Martin (SM) grade 1 and 2 AVMs (16 %), eight of 22 (36 %) grade 3, and four of six grade 4 (67 %), permanent morbidity >mRS 1 in 3 (4.3 %) grade 1 and 2, four (18 %) grade 3, and three (50 %) grade 4. Treatment inflicted loss of QALY amounted to 0.5 years for SM grade 1-2, 2.5 years grade 3, 7.3 years for grade 4. For the SM grades 1 and 2, the treatment-related loss of 0.5 QALY was met by the natural course after 2.7-4.3 years. For the Spetzler-Martin grades 3 and 4, the treatment-induced loss QALY was not met by the natural risk within a foreseeable time. Permanent morbidity and treatment inflicted loss of QALY of patients younger than 39 years was lower than that of older patients (7 vs. 15 % and 1.0 vs. 2.1 QALY). CONCLUSIONS: Microsurgically managed SM grades 1 and 2 fared better than the modeled natural course but grades 3 and 4 AVM did not benefit from surgery. Younger patients appear to fare more favorably than older patients.