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Introduction: This study aimed to test the diagnostic significance of FET-PET imaging combined with machine learning for the differentiation between multiple sclerosis (MS) and glioma II°-IV°., Methods: Our database was screened for patients in whom FET-PET imaging was performed for the diagnostic workup of newly diagnosed lesions evident on MRI and suggestive of glioma. Among those, we identified patients with histologically confirmed glioma II°-IV°, and those who later turned out to have MS. For each group, tumor-to-brain ratio (TBR) derived features of FET were determined. A support vector machine (SVM) based machine learning algorithm was constructed to enhance classification ability, and Receiver Operating Characteristic (ROC) analysis with area under the curve (AUC) metric served to ascertain model performance., Results: A total of 41 patients met selection criteria, including seven patients with MS and 34 patients with glioma. TBR values were significantly higher in the glioma group (TBRmax glioma vs. MS: p = 0.002; TBRmean glioma vs. MS: p = 0.014). In a subgroup analysis, TBR values significantly differentiated between MS and glioblastoma (TBRmax glioblastoma vs. MS: p = 0.0003, TBRmean glioblastoma vs. MS: p = 0.0003) and between MS and oligodendroglioma (ODG) (TBRmax ODG vs. MS: p = 0.003; TBRmean ODG vs. MS: p = 0.01). The ability to differentiate between MS and glioma II°-IV° increased from 0.79 using standard TBR analysis to 0.94 using a SVM based machine learning algorithm., Conclusions: FET-PET imaging may help differentiate MS from glioma II°-IV° and SVM based machine learning approaches can enhance classification performance.

Purpose: The aim of the present study based on the PriCoTTF-phase I/II trial is the quantification of skin-normal tissue complication probabilities of patients with newly diagnosed glioblastoma multiforme treated with Tumor Treating Field (TTField) electrodes, concurrent radiotherapy, and temozolomide. Furthermore, the skin-sparing effect by the clinically applied strategy of repetitive transducer array fixation around their center position shall be examined., Material and Methods: Low-dose cone-beam computed tomography (CBCT) scans of all fractions of the first seven patients of the PriCoTTF-phase I/II trial, used for image guidance, were applied for the dosimetric analysis, for precise TTField transducer array positioning and contour delineation. Within this trial, array positioning was varied from fixation-to-fixation period with a standard deviation of 1.1 cm in the direction of the largest variation of positioning and 0.7 cm in the perpendicular direction. Physical TTField electrode composition was examined and a respective Hounsfield Unit attributed to the TTField electrodes. Dose distributions in the planning CT with TTField electrodes in place, as derived from prefraction CBCTs, were calculated and accumulated with the algorithm Acuros XB. Dose-volume histograms were obtained for the first and second 2 mm scalp layer with and without migrating electrodes and compared with those with fixed electrodes in an average position. Skin toxicity was quantified according to Lyman's model. Minimum doses in hot-spots of 0.05 cm 2 and 25 cm 2 ( Δ D 0.05cm 2 , Δ D 25cm 2 ) size in the superficial skin layers were analyzed., Results: Normal tissue complication probabilities (NTCPs) for skin necrosis ranged from 0.005% to 1.474% (median 0.111%) for the different patients without electrodes. NTCP logarithms were significantly dependent on patient (P < 0.0001) and scenario (P < 0.0001) as classification variables. Fixed positioning of TTField arrays increased skin-NTCP by a factor of 5.50 (95%, CI: 3.66-8.27). The variation of array positioning increased skin-NTCP by a factor of only 3.54 (95%, CI: 2.36-5.32) (P < 0.0001, comparison to irradiation without electrodes; P = 0.036, comparison to irradiation with fixed electrodes). NTCP showed a significant rank correlation with D25cm 2 over all patients and scenarios (r s  = 0.76; P < 0.0001)., Conclusion: Skin-NTCP calculation uncovers significant interpatient heterogeneity and may be used to stratify patients into high- and low-risk groups of skin toxicity. Array position variation may mitigate about one-third of the increase in surface dose and skin-NTCP by the TTField electrodes., (© 2020 The Authors. Journal of Applied Clinical Medical Physics published by Wiley Periodicals, Inc. on behalf of American Association of Physicists in Medicine.)

Background: Symptomatic epilepsy is a common symptom of glioblastoma, which may occur in different stages of disease. There are discrepant reports on association between early seizures and glioblastoma survival, even less is known about the background of these seizures. We aimed at analyzing the risk factors and clinical impact of perioperative seizures in glioblastoma., Methods: All consecutive cases with de-novo glioblastoma treated at our institution between 01/2006 and 12/2018 were eligible for this study. Perioperative seizures were stratified into seizures at onset (SAO) and early postoperative seizures (EPS, ≤21days after surgery). Associations between patients characteristics and overall survival (OS) with SAO and EPS were addressed., Results: In the final cohort ( n = 867), SAO and EPS occurred in 236 (27.2%) and 67 (7.7%) patients, respectively. SAO were independently predicted by younger age ( P = .009), higher KPS score ( P = .002), tumor location (parietal lobe, P = .001), GFAP expression (≥35%, P = .045), and serum chloride at admission (>102 mmol/L, P = .004). In turn, EPS were independently associated with tumor location (frontal or temporal lobe, P = .013) and pathologic laboratory values at admission (hemoglobin < 12 g/dL, [ P = .044], CRP > 1.0 mg/dL [ P = 0.036], and GGT > 55 U/L [ P = 0.025]). Finally, SAO were associated with gross-total resection ( P = .006) and longer OS ( P = .030), whereas EPS were related to incomplete resection ( P = .005) and poorer OS ( P = .009)., Conclusions: In glioblastoma patients, SAO and EPS seem to have quite different triggers and contrary impact on treatment success and OS. The clinical characteristics of SAO and EPS patients might contribute to the observed survival differences., (© The Author(s) 2020. Published by Oxford University Press, the Society for Neuro-Oncology and the European Association of Neuro-Oncology.)

Pancreatic ductal adenocarcinoma (PDAC) is resistant to virtually all chemo- and targeted therapeutic approaches. Epigenetic regulators represent a novel class of drug targets. Among them, BET and HDAC proteins are central regulators of chromatin structure and transcription, and preclinical evidence suggests effectiveness of combined BET and HDAC inhibition in PDAC. Here, we describe that TW9, a newly generated adduct of the BET inhibitor (+)-JQ1 and class I HDAC inhibitor CI994, is a potent dual inhibitor simultaneously targeting BET and HDAC proteins. TW9 has a similar affinity to BRD4 bromodomains as (+)-JQ1 and shares a conserved binding mode, but is significantly more active in inhibiting HDAC1 compared to the parental HDAC inhibitor CI994. TW9 was more potent in inhibiting tumor cell proliferation compared to (+)-JQ1, CI994 alone or combined treatment of both inhibitors. Sequential administration of gemcitabine and TW9 showed additional synergistic antitumor effects. Microarray analysis revealed that dysregulation of a FOSL1-directed transcriptional program contributed to the antitumor effects of TW9. Our results demonstrate the potential of a dual chromatin-targeting strategy in the treatment of PDAC and provide a rationale for further development of multitarget inhibitors., (© 2020 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Background: The protease inhibitor ritonavir (RTV) is a clinical-stage inhibitor of the human immunodeficiency virus. In a drug repositioning approach, we here exhibit the additional potential of RTV to augment current treatment of glioblastoma, the most aggressive primary brain tumour of adulthood., Methods: We explored the antitumour activity of RTV and mechanisms of action in a broad spectrum of short-term expanded clinical cell samples from primary and recurrent glioblastoma and in a cohort of conventional cell lines and non-tumour human neural controls in vitro. To validate RTV efficacy in monotherapeutic and in combinatorial settings, we used patient-derived xenograft models in a series of in vivo studies., Results: RTV monotherapy induced a selective antineoplastic response and demonstrated cytostatic and anti-migratory activity at clinical plasma peak levels. Additional exposure to temozolomide or irradiation further enhanced the effects synergistically, fostered by mechanisms of autophagy and increased endoplasmic reticulum stress. In xenograft models, we consequently observed increasing overall survival under the combinatorial effect of RTV and temozolomide., Conclusions: Our data establish RTV as a valuable repositioning candidate for further exploration as an adjunct therapeutic in the clinical care of glioblastoma., Competing Interests: Conflict of interest statement The authors have declared no conflicts of interest., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Bony defects of the orbit with connection to the paranasal sinuses can lead to orbital emphysema. A case of compression of the optic nerve due to orbital emphysema in a 39-year-old man is reported. Visual acuity was hand movements only and intraocular pressure was increased to 32 mm Hg with an exophthalmus of 6 mm in comparison to the unaffected side. A sufficient pressure relief could be achieved by canthotomy with cantholysis and radial peritomy.

Glioblastoma is a devastating disease, for which biomarkers allowing a prediction of prognosis are urgently needed. microRNAs have been described as potentially valuable biomarkers in cancer. Here, we studied a panel of microRNAs in extracellular vesicles (EVs) from the serum of glioblastoma patients and evaluated their correlation with the prognosis of these patients. The levels of 15 microRNAs in EVs that were separated by size-exclusion chromatography were studied by quantitative real-time PCR, followed by CD44 immunoprecipitation (SEC + CD44), and compared with those from the total serum of glioblastoma patients ( n = 55) and healthy volunteers ( n = 10). Compared to total serum, we found evidence for the enrichment of miR-21-3p and miR-106a-5p and, conversely, lower levels of miR-15b-3p, in SEC + CD44 EVs. miR-15b-3p and miR-21-3p were upregulated in glioblastoma patients compared to healthy subjects. A significant correlation with survival of the patients was found for levels of miR-15b-3p in total serum and miR-15b-3p, miR-21-3p, miR-106a-5p, and miR-328-3p in SEC + CD44 EVs. Combining miR-15b-3p in serum or miR-106a-5p in SEC + CD44 EVs with any one of the other three microRNAs in SEC + CD44 EVs allowed for a prognostic stratification of glioblastoma patients. We have thus identified four microRNAs in glioblastoma patients whose levels, in combination, can predict the prognosis for these patients., Competing Interests: U.H. reports grants and personal fees from Roche; personal fees and non-financial support from Medac and Bristol-Myers Squibb; and personal fees from Novocure, Novartis, Daichii-Sankyo, Riemser, and Noxxon. N.S. reports personal fees and other support from Roche and received honoraria for advisory board from Bayer. M.G. reports grants, personal fees, and other support from Novocure and Medac, and personal fees from Merck. C.S. reports personal fees from Roche. S.W. and C.C. are employed by Miltenyi Biomedicine. All other authors declare no conflicts of interest.

Extracellular vesicles, including exosomes/microvesicles (EMVs), have been described as sensitive biomarkers that represent disease states and response to therapies. In light of recent reports of disease-mirroring EMV molecular signatures, the present study profiled two EMVs from different Parkinson's disease (PD) tissue sources: (a) neural progenitor cells derived from an endogenous adult stem/progenitor cell, called adult human neural progenitor (AHNP) cells, that we found to be pathological when isolated from postmortem PD patients' substantia nigra; and (b) leucine-rich repeat kinase 2 (LRRK2) gene identified patient induced pluripotent stem cells (iPSCs), which were used to isolate EMVs and begin to characterize their cargoes. Initial characterization of EMVs derived from idiopathic patients (AHNPs) and mutant LRRK2 patients showed differences between both phenotypes and when compared with a sibling control in EMV size and release based on Nanosight analysis. Furthermore, molecular profiling disclosed that neurodegenerative-related gene pathways altered in PD can be reversed using gene-editing approaches. In fact, the EMV cargo genes exhibited normal expression patterns after gene editing. This study shows that EMVs have the potential to serve as sensitive biomarkers of disease state in both idiopathic and gene-identified PD patients and that following gene-editing, EMVs reflect a corrected state. This is relevant for both prodromal and symptomatic patient populations where potential responses to therapies can be monitored via non-invasive liquid biopsies and EMV characterizations., (© 2019 Wiley Periodicals, Inc.)

Purpose: In the EF-14 trial for newly diagnosed glioblastoma (ndGBM) patients addition of Tumour Treating Fields (TTFields) to temozolomide treatment resulted in a significantly improved overall survival (OS). In the NOA-09/CeTeG trial, combination of lomustine and temozolomide was superior to temozolomide monotherapy in patients with O6-methylguanine DNA methyltransferase (MGMT) promoter methylated (MGMTm) ndGBM. We evaluated combination of these two treatment modalities in patients with MGMTm ndGBM. There have been so far no data on the combination of these two efficient regimens., Methods: This bicentric retrospective analysis investigated 16 patients. Parameters evaluated included safety outcome as measured by Common Toxicity Criteria for Adverse Events (CTCAE), clinical outcomes, and compliance to treatment., Results: Hematologic adverse events CTCAE ≥ 3 were observed in seven, hepatotoxic adverse events of CTCAE ≥ 3 in four patients. Mild to moderate skin toxicity was detected in six patients. At data cutoff, patients demonstrated a median progression-free survival (PFS) of 20 months. The usage rate of TTFields showed a high median adherence (83%) to the therapy., Conclusions: This analysis provides first indication that the combination of TTFields/lomustine/temozolomide is safe and feasible. The observed survival outcomes might suggest potential beneficial effects.

Objective: To determine the role of olfactory function in patients with glioblastoma multiforme (GBM) as a prognostic clinical measure., Methods: In a prospective case-control study, olfactory testing was performed in 73 patients with primary GBM at baseline during first-line treatment and at later follow-ups. An age-matched control cohort consisted of 49 patients with neurologic diseases, excluding those known to affect olfactory function per se. Depending on the olfactory testing score, patients were allotted to a hyposmia group (HG) or normosmia group (NG). MRI analysis was performed to assess whether tumor location affects olfactory pathways., Results: Patients with GBM had olfactory dysfunction significantly more often compared to the control cohort ( p = 0.003). Tumor location could not explain this finding since no relevant difference in MRI-based olfactory pathway involvement was found between HG and NG ( p = 0.131). Patients with olfactory dysfunction had significantly worse overall survival (OS) and progression-free survival (PFS) compared to those without dysfunction (median OS 20.9 vs 40.6 months, p = 0.035; median PFS, 9 vs 19 months, p = 0.022). Multivariate analysis in patients without MRI-based involvement of olfactory pathways confirmed olfaction is an independent prognostic factor for OS (hazard ratio [HR] 0.43; p = 0.042) and PFS (HR 0.51; p = 0.049)., Conclusion: This pilot study provides the first indication that olfactory dysfunction is frequently observed in GBM and may be associated with worse survival outcome in GBM. However, validation of these results in an independent cohort is needed., (© 2019 American Academy of Neurology.)

Early detection of cancer holds high promise for reducing cancer-related mortality. Detection of circulating tumor-specific nucleic acids holds promise, but sensitivity and specificity issues remain with current technology. We studied cell-free RNA (cfRNA) in patients with non-small cell lung cancer (NSCLC; n = 56 stage IV, n = 39 stages I-III), pancreatic cancer (PDAC, n = 20 stage III), malignant melanoma (MM, n = 12 stage III-IV), urothelial bladder cancer (UBC, n = 22 stage II and IV), and 65 healthy controls by means of next generation sequencing (NGS) and real-time droplet digital PCR (RT-ddPCR). We identified 192 overlapping upregulated transcripts in NSCLC and PDAC by NGS, more than 90% of which were noncoding. Previously reported transcripts (e.g., HOTAIRM1) were identified. Plasma cfRNA transcript levels of POU6F2-AS2 discriminated NSCLC from healthy donors (AUC = 0.82 and 0.76 for stages IV and I-III, respectively) and significantly associated ( p = 0.017) with the established tumor marker Cyfra 21-1. cfRNA yield and POU6F2-AS transcript abundance discriminated PDAC patients from healthy donors (AUC = 1.0). POU6F2-AS2 transcript was significantly higher in MM ( p = 0.044). In summary, our findings support further validation of cfRNA detection by RT-ddPCR as a biomarker for early detection of solid cancers., Competing Interests: The authors declare no conflict of interest.

Background: Leptomeningeal metastasis (LM) is a predominantly late stage, devastating complication of a variety of malignant solid tumors. Diagnosis relies predominantly on neurological, radiographic, and cerebrospinal fluid (CSF) assessments. Recently, liquid biopsy tests derived from CSF has shown to be a feasible, noninvasive promising approach to tumor molecular profiling for proper brain cancer diagnostic treatment, thereby providing an opportunity for CSF-based personalized medicine. However, LM is typically misleadingly assumed to originate from only one primary tumor type., Case Presentation: In this case report, we provide first evidence of the co-occurrence of LM originating from more than one primary tumor types., Discussion and Conclusions: Based on this patient case profile, the co-occurrence of LM from two or more primary tumor types should be accounted for when deriving diagnostic conclusions from liquid biopsy tests.

Dynamic alterations in the unique brain extracellular matrix (ECM) are involved in malignant brain tumors. Yet studies of brain ECM roles in tumor cell behavior have been difficult due to lack of access to the human brain. We present a tunable 3D bioengineered brain tissue platform by integrating microenvironmental cues of native brain-derived ECMs and live imaging to systematically evaluate patient-derived brain tumor responses. Using pediatric ependymoma and adult glioblastoma as examples, the 3D brain ECM-containing microenvironment with a balance of cell-cell and cell-matrix interactions supports distinctive phenotypes associated with tumor type-specific and ECM-dependent patterns in the tumor cells' transcriptomic and release profiles. Label-free metabolic imaging of the composite model structure identifies metabolically distinct sub-populations within a tumor type and captures extracellular lipid-containing droplets with potential implications in drug response. The versatile bioengineered 3D tumor tissue system sets the stage for mechanistic studies deciphering microenvironmental role in brain tumor progression.

Background: A high-quality reference genome is an essential tool for applied and basic research on arthropods. Long-read sequencing technologies may be used to generate more complete and contiguous genome assemblies than alternate technologies; however, long-read methods have historically had greater input DNA requirements and higher costs than next-generation sequencing, which are barriers to their use on many samples. Here, we present a 2.3 Gb de novo genome assembly of a field-collected adult female spotted lanternfly (Lycorma delicatula) using a single Pacific Biosciences SMRT Cell. The spotted lanternfly is an invasive species recently discovered in the northeastern United States that threatens to damage economically important crop plants in the region., Results: The DNA from 1 individual was used to make 1 standard, size-selected library with an average DNA fragment size of ∼20 kb. The library was run on 1 Sequel II SMRT Cell 8M, generating a total of 132 Gb of long-read sequences, of which 82 Gb were from unique library molecules, representing ∼36× coverage of the genome. The assembly had high contiguity (contig N50 length = 1.5 Mb), completeness, and sequence level accuracy as estimated by conserved gene set analysis (96.8% of conserved genes both complete and without frame shift errors). Furthermore, it was possible to segregate more than half of the diploid genome into the 2 separate haplotypes. The assembly also recovered 2 microbial symbiont genomes known to be associated with L. delicatula, each microbial genome being assembled into a single contig., Conclusions: We demonstrate that field-collected arthropods can be used for the rapid generation of high-quality genome assemblies, an attractive approach for projects on emerging invasive species, disease vectors, or conservation efforts of endangered species., (© The Author(s) 2019. Published by Oxford University Press.)

Aims: Amino acid PET and magnetic resonance spectroscopy (MRS) are at the forefront of noninvasive imaging techniques used for detection and subtyping of glioma-suspicious lesions. In this pilot study, we compare L-methyl-C-methionine PET and MRS for their ability to predict glioma subtypes., Methods: Nineteen patients with histologically, confirmed newly diagnosed glioma underwent preoperative L-methyl-C-methionine PET and MRS in 1 diagnostic session. According to the molecular portfolio and histopathologic diagnosis, patients were subdivided in isocitrate dehydrogenase (IDH) wild-type glioblastoma, IDH wild-type grade II/III glioma, IDH-mutant grade II/III glioma without 1p/19q codeletion, and with 1p/19q codeletion subgroups. Maximum tumor-to-brain ratio (TBRmax), creatine, choline, and N-acetyl aspartate peaks were correlated with postoperative histopathologic tumor diagnoses., Results: Maximum tumor-to-brain ratio was highest in glioblastoma patients (4.18) followed by patients with IDH wild-type grade II and III glioma (3.41). The latter TBRmax values were higher compared with those in patients with IDH-mutant grade II/III glioma without 1p/19q codeletion (1.95) and in patients with IDH-mutant 1p/19q codeleted grade II and III glioma (2.79). Magnetic resonance spectroscopy marker distribution showed no clear trend. Receiver operating characteristic analysis revealed TBRmax to be the best performing parameter in identifying IDH status (area under the curve, 0.67) and all spectroscopy markers combined in identifying glioma subgroups (area under the curve, 0.68), respectively., Conclusions: L-Methyl-C-methionine PET and MRS bear limited potential in glioma subgrouping. L-Methyl-C-methionine PET appears to be superior in differentiating IDH status, whereas MRS is more helpful in glioma subgrouping.

Upland cotton ( Gossypium hirsutum L.) produces terpenoid aldehydes (TAs) that protect the plant from microbial and insect infestations. Foliar TAs include plus (+)- and minus (-)-gossypol, hemigossypolone, and heliocides. To examine foliar TAs' response to physical wounding, the four TA derivatives of a fully glanded G. hirsutum variety JACO GL were quantified by ultra-high performance liquid chromatography. The results show that foliar heliocides increased by 1.7-fold in younger leaves after wounding. While the hemigossypolone level was not affected by the physical wounding, the level of heliocides was significantly increased up to 1.8-fold in the younger leaves. Upland cotton accumulates concentrated carbohydrates, amino acids, and fatty acids in foliar extrafloral nectar (EFN) to serve as a nutrient resource, which attracts both beneficial insects and damaging pests. To better understand the nectar physiology, particularly to determine the temporal dynamics of EFN metabolites in response to the wounding, a gas chromatograph-mass spectrometer (GC-MS) was used to perform metabolic profiling analyses of a G. hirsutum variety Deltapine 383 that has fully developed extrafloral nectaries. A total of 301 compounds were monitored, specifically 75 primary metabolites, two secondary metabolites and 224 unidentified compounds. The physical wounding treatment changed the EFN composition and lowered overall production. The accumulation of 30 metabolites was altered in response to the wounding treatment and threonic acid levels increased consistently. GC-MS combined with Kovat's analysis enabled identification of EFN secondary metabolites including furfuryl alcohol and 5-hyrdomethoxyfurfural, which both have antioxidant and antimicrobial properties that may protect the nectar against microbial pathogens. This study provides new insights into the wounding response of cotton plants in terms of cotton metabolites found in leaf glands and extrafloral nectar as well as highlighting some protective functions of secondary metabolites produced in foliar glands and extrafloral nectaries., Competing Interests: The authors declare no conflict of interest.

A collection of cultivated and non-cultivated species of cotton ( Gossypium spp.) has been maintained for the last four decades in Multan, Pakistan. This geographical location has been observed as a hotspot for the evolution of begomoviruses and satellites associated with cotton leaf curl disease (CLCuD). Recent studies showed that begomoviruses responsible for the CLCuD epidemic in the 1990s, and that almost disappeared from the CLCuD complex in 2000s, have been observed again in CLCuD-infected cotton fields. To identify host species that acted as probable reservoirs for these viruses, we characterized begomoviruses and satellites in non-cultivated cotton species G. raimondii , G. thurberi and G. mustelinum and identified several species of CLCuD associated begomoviruses and satellites. Further, phylogenetic analysis indicated that the identified begomoviruses and beta/alphasatellites are closely related to the ones associated with the most recent CLCuD complex. qPCR indicated that the comparative level of virus significantly decreased in the presence of alphasatellites. Our results indicated that non-cultivated cotton species have been continuously challenged by diverse begomoviruses and associated satellites and act as reservoirs for CLCuD associated begomoviruses. These results provide novel insights into understanding the spread of begomoviruses and associated satellites in New World cotton species introduced into the Old World.

Purpose: Antiangiogenic treatment approaches have failed to improve outcome in randomized trials of high-grade astrocytoma. One key mechanism of resistance to antiangiogenic treatment may concern the upregulation of alternative pro-angiogenic pathways. Regorafenib is a potent multikinase inhibitor that may alter some of those pathways. In this retrospective study, we investigated efficacy and radiographic tumor growth patterns of regorafenib in recurrent high-grade astrocytoma., Methods: We screened for patients with high-grade astrocytoma in whom regorafenib was administered for at least 4 weeks. We assessed treatment efficacy in terms of progression-free survival (PFS), overall survival, and adverse events defined by Common Toxicity Criteria (CTC). In addition, radiographic tumor growth patterns were determined at baseline and recurrence., Results: A total of 6 patients met eligibility criteria. The number of recurrences prior to regorafenib varied between 2 and 6. Patients were on regorafenib treatment for at least 4 weeks and maximally 14 weeks. Median PFS was 3.5 months and ranged from 2.0 to 4.0 months. Radiographic response was progressive disease in all patients with an objective response rate of 0%. CTC°3 adverse events were observed in all but one patient. The most common radiographic growth pattern was local with no change in growth pattern at recurrence. An infiltrative tumor growth was not induced in any patient., Conclusions: This retrospective study indicates a very poor performance of regorafenib in recurrent high-grade astrocytoma with a fairly high number of CTC°3 adverse events. In addition, regorafenib does not seem to bear a potential for infiltrative tumor growth promotion.

Background: Molecularly targeted therapies using receptor inhibitors, small molecules or monoclonal antibodies are routinely applied in oncology. Verification of target expression should be mandatory prior to initiation of therapy, yet, determining the expression status is most challenging in recurrent glioblastoma (GBM) where most patients are not eligible for second-line surgery. Because very little is known on the consistency of expression along the clinical course we here explored common drug targets in paired primary vs. recurrent GBM tissue samples., Methods: Paired surgical tissue samples were derived from a homogeneously treated cohort of 34 GBM patients. All patients received radiotherapy and temozolomide chemotherapy. Verification of common drug targets included immunohistological analysis of PDGFR-β, FGFR-2, FGFR-3, and mTOR-pathway component (phospho-mTOR Ser2448 ) as well as molecular, MLPA-based analysis of specific copy number aberrations at the gene loci of ALK, PDGFRA, VEGFR2/KDR, EGFR, MET, and FGFR1., Results: Paired tumor tissue exhibited significant changes of expression in 9 of the 10 investigated druggable targets (90%). Only one target (FGFR1) was found "unchanged", since dissimilar expression was observed in only one of the 34 paired tumor tissue samples. All other targets were variably expressed with an 18-56% discordance rate between primary and recurrent tissue., Conclusions: The high incidence of dissimilar target expression status in clinical samples from primary vs. recurrent GBM suggests clinically relevant heterogeneity along the course of disease. Molecular target expression, as determined at primary diagnosis, may not necessarily present rational treatment clues for the clinical care of recurrent GBM. Further studies need to analyze the therapeutic impact of longitudinal heterogeneity in GBM.

Introduction: Treatment of patients with metastatic melanoma is hampered by drug-resistance and often requires combination with radiotherapy as last-resort option. However, also after radiotherapy, clinical relapses are common., Methods & Results: Our preclinical models indicated a higher rate of tumour relapse when melanoma cells were first treated with BRAF V600E inhibition (BRAFi) followed by radiotherapy as compared to the reverse sequence. Accordingly, retrospective follow-up data from 65 stage-IV melanoma patients with irradiated melanoma brain metastases confirmed a shortened duration of local response of mitogen-activated protein kinase (MAPK)-inhibitor-pretreated compared with MAPK-inhibitor-naïve intracranial metastases. On the molecular level, we identified JARID1B/KDM5B as a cellular marker for cross-resistance between BRAFi and radiotherapy. JARID1B high cells appeared more frequently under upfront BRAFi as compared with upfront radiation. JARID1B favours cell survival by transcriptional regulation of genes controlling cell cycle, DNA repair and cell death., Conclusion: The level of cross-resistance between combined MAPK inhibition and radiotherapy is dependent on the treatment sequence. JARID1B may represent a novel therapy-overarching resistance marker., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Purpose: With the advent of the revised WHO classification from 2016, molecular features, including isocitrate dehydrogenase (IDH) mutation have become important in glioma subtyping. This pilot trial analyzed the potential for C-methionine (MET) PET/MRI in classifying glioma according to the revised WHO classification using a machine learning model., Methods: Patients with newly diagnosed WHO grade II-IV glioma underwent preoperative MET-PET/MRI imaging. Patients were retrospectively divided into four groups: IDH wild-type glioblastoma (GBM), IDH wild-type grade II/III glioma (GII/III-IDHwt), IDH mutant grade II/III glioma with codeletion of 1p19q (GII/III-IDHmut1p19qcod) or without 1p19q-codeletion (GII/III-IDHmut1p19qnc). Within each group, the maximum tumor-to-brain-ratio (TBRmax) of MET-uptake was calculated. To gain generalizable implications from our data, we made use of a machine learning algorithm based on a development and validation subcohort. A support vector machine model was fit to the development subcohort and evaluated on the validation subcohort. Receiver operating characteristic (ROC) analysis served as metric to assess model performance., Results: Of a total of 259 patients, 39 patients met the inclusion criteria. TBRmax was highest in the GBM cohort (TBRmax 3.83 ± 1.30) and significantly higher (P = 0.004) compared to GII/III-IDHmut1p19qnc group, where TBRmax was lowest (TBRmax 2.05 ± 0.94). ROC analysis showed poor AUC for glioma subtyping (AUC 0.62) and high AUC of 0.79 for predicting IDH status. In the GII/III-IDHmut1p19qcod group, TBR values were slightly higher than in the IDHmut1p19qnc group., Conclusions: MET-PET/MRI imaging in pre-operatively classifying glioma entities appears useful for the assessment of IDH status. However, a larger trial is needed prior to translation into the clinical routine.

In a subset of glioblastoma (GBM) patients, the differentiation between tumor progression and tumor pseudoprogression (PsP) is challenging. This case describes a male patient suffering from isocitrate dehydrogenase 1 (IDH1) mutant GBM who demonstrated an increasing contrast-enhancing (CE) lesion on a cranial magnetic resonance imaging (cMRI) scan 8 months after radiochemotherapy. In accordance with the response assessment in neuro-oncology (RANO) criteria, the cMRI lesion was classified as recurrent tumor, although 18 F-fluoroethyl-L-tyrosine positron emission tomography ( 18 F-FET-PET) did not indicate vital tumor tissue. The patient underwent re-surgery but histopathology only revealed reactive and necrotic tissue, consistent with PsP. Nine weeks after complete resection of the CE lesion, a new lesion emerged that later regressed in the follow-up cMRI scans, thereby retrospectively establishing the diagnosis of recurrent PsP., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Main Conclusion: Some naturally occurring cotton accessions contain commercially attractive seed oil fatty acid profiles. The likely causal factor for a high-oleate trait in pima cotton ( Gossypium barbadense ) accession GB-713 is described here. Vegetable oils are broadly used in the manufacture of many human and animal nutritional products, and in various industrial applications. Along with other well-known edible plant oils from soybean, corn, and canola, cottonseed oil is a valuable commodity. Cottonseed oil is a co-product derived from the processing of cottonseed fiber. In the past, it was used extensively in a variety of food applications. However, cottonseed oil has lost market share in recent years due to less than optimal ratios of the constituent fatty acids found in either traditional or partially hydrogenated oil. Increased awareness of the negative health consequences of dietary trans-fats, along with the public wariness associated with genetically modified organisms has created high demand for naturally occurring oil with high monounsaturate/polyunsaturate ratios. Here, we report the discovery of multiple exotic accessions of pima cotton that contain elevated seed oil oleate content. The genome of one such accession was sequenced, and a mutant candidate fatty acid desaturase-2 (FAD2-1D) gene was identified. The mutant protein produced significantly less linoleic acid in infiltrated Arabidopsis leaf assays, compared to a repaired version of the same enzyme. Identification of this gene provides a valuable resource. Development of markers associated with this mutant locus will be very useful in efforts to breed the high-oleate trait into agronomic fiber accessions of upland cotton.

Rationale: Timely detection of pseudoprogression (PSP) is crucial for the management of patients with high-grade glioma (HGG) but remains difficult. Textural features of O-(2-[18F]fluoroethyl)-L-tyrosine positron emission tomography (FET-PET) mirror tumor uptake heterogeneity; some of them may be associated with tumor progression., Methods: Fourteen patients with HGG and suspected of PSP underwent FET-PET imaging. A set of 19 conventional and textural FET-PET features were evaluated and subjected to unsupervised consensus clustering. The final diagnosis of true progression vs. PSP was based on follow-up MRI using RANO criteria., Results: Three robust clusters have been identified based on 10 predominantly textural FET-PET features. None of the patients with PSP fell into cluster 2, which was associated with high values for textural FET-PET markers of uptake heterogeneity. Three out of 4 patients with PSP were assigned to cluster 3 that was largely associated with low values of textural FET-PET features. By comparison, tumor-to-normal brain ratio (TNRmax) at the optimal cutoff 2.1 was less predictive of PSP (negative predictive value 57% for detecting true progression, p=0.07 vs. 75% with cluster 3, p=0.04)., Principal Conclusions: Clustering based on textural O-(2-[18F]fluoroethyl)-L-tyrosine PET features may provide valuable information in assessing the elusive phenomenon of pseudoprogression.

Purpose: Investigation of clonal heterogeneity may be key to understanding mechanisms of therapeutic failure in human cancer. However, little is known on the consequences of therapeutic intervention on the clonal composition of solid tumors., Experimental Design: Here, we used 33 single cell-derived subclones generated from five clinical glioblastoma specimens for exploring intra- and interindividual spectra of drug resistance profiles in vitro In a personalized setting, we explored whether differences in pharmacologic sensitivity among subclones could be employed to predict drug-dependent changes to the clonal composition of tumors., Results: Subclones from individual tumors exhibited a remarkable heterogeneity of drug resistance to a library of potential antiglioblastoma compounds. A more comprehensive intratumoral analysis revealed that stable genetic and phenotypic characteristics of coexisting subclones could be correlated with distinct drug sensitivity profiles. The data obtained from differential drug response analysis could be employed to predict clonal population shifts within the naïve parental tumor in vitro and in orthotopic xenografts. Furthermore, the value of pharmacologic profiles could be shown for establishing rational strategies for individualized secondary lines of treatment., Conclusions: Our data provide a previously unrecognized strategy for revealing functional consequences of intratumor heterogeneity by enabling predictive modeling of treatment-related subclone dynamics in human glioblastoma. Clin Cancer Res; 23(2); 562-74. ©2016 AACR., Competing Interests: The authors declare no conflict of interests, (©2016 American Association for Cancer Research.)

The conserved coat or V2 gene of begomoviruses is responsible for viral movement in the plant cells. RNAi technology was used to silence V2 gene for resistance against these viruses in transgenic plants. The transformation of the RNAi-based gene construct targeting V2 gene of CLCuKoV-Bur, cloned under 35S promoter, was done in two elite cotton varieties MNH-786 and VH-289 using shoot apex cut method of gene transformation. The transformation efficiency was found to be 3.75 and 2.88 % in MNH-786 and VH-289, respectively. Confirmation of successful transformation was done through PCR in T 0 , T 1 , and T 2 generations using gene-specific primers. Transgenic cotton plants were categorized on the basis of the virus disease index in T 1 generation. Copy number and transgene location were observed using FISH and karyotyping in T 2 generation which confirmed random integration of V2 RNAi amplicon at chromosome 6 and 16. Real-time quantitative PCR analyses of promising transgenic lines showed low virus titer compared to wild-type control plants upon challenging them with viruliferous whiteflies in a contained environment. From the results, it was concluded that amplicon V2 RNAi construct was able to limit virus replication and can be used to control CLCuV in the field., Competing Interests: The authors declare no conflict of interests.

Background: Critical illness myopathy (CIM) and polyneuropathy (CIP) are a common complication of critical illness. Both cause intensive-care-unit-acquired (ICU-acquired) muscle weakness (ICUAW) which increases morbidity and delays rehabilitation and recovery of activities of daily living such as walking ability. Focused physical rehabilitation of people with ICUAW is, therefore, of great importance at both an individual and a societal level. A recent systematic Cochrane review found no randomised controlled trials (RCT), and thus no supporting evidence, for physical rehabilitation interventions for people with defined CIP and CIM to improve activities of daily living. Therefore, the aim of our study is to compare the effects of an additional physiotherapy programme with systematically augmented levels of mobilisation with additional in-bed cycling (as the parallel group) on walking and other activities of daily living., Methods/design: We will conduct a prospective, rater-masked RCT of people with ICUAW with a defined diagnosis of CIM and/or CIP in our post-acute hospital. We will randomly assign patients to one of two parallel groups in a 1:1 ratio and will use a concealed allocation. One intervention group will receive, in addition to standard ICU treatment, physiotherapy with systematically augmented levels of mobilisation (five times per week, over 2 weeks; 20 min each session; with a total of 10 additional sessions). The other intervention group will receive, in addition to standard ICU treatment, in-bed cycle sessions (same number, frequency and treatment time as the intervention group). Standard ICU treatment includes sitting balance exercise, stretching, positioning, and sit-to-stand training, and transfer training to get out of bed, strengthening exercise (in and out of bed), and stepping and assistive standing exercises. Primary efficacy endpoints will be walking ability (defined as a Functional Ambulation Category (FAC) level of ≥3) and the sum score of the Functional Status Score for the Intensive Care Unit (FSS-ICU) (range 0-22 points) assessed by a blinded tester immediately after 2 weeks of additional therapy. Secondary outcomes will include assessment of sit-to-stand recovery, overall limb strength (Medical Research Council, MRC) and grip strength, the Physical Function for the Intensive Care Unit Test-Scored (PFIT-S), the EuroQol 5 Dimensions (EQ-5D) questionnaire and the Reintegration to Normal Living Index (RNL-Index) assessed by a blinded tester. We will measure primary and secondary outcomes with blinded assessors at baseline, immediately after 2 weeks of additional therapy, and at 3 weeks and 6 months and 12 months after the end of the additional therapy intervention. Based on our sample size calculation 108 patients will be recruited from our post-acute ICU in the next 3 to 4 years., Discussion: This will be the first RCT comparing the effects of two physical rehabilitation interventions for people with ICUAW due to defined CIP and/or CIM to improve walking and other activities of daily living. The results of this trial will provide robust evidence for physical rehabilitation of people with CIP and/or CIP who often require long-term care., Trial Registration: We registered the study on 6 April 2016 before enrolling the first patient in the trial at the German Clinical Trials Register ( www.germanctr.de ) with the identifier DRKS00010269 . This is the first version of the protocol (FITonICU study protocol).

The advent of the genome sequences of Arachis duranensis and Arachis ipaensis has ushered in a new era for peanut genomics. With the goal of producing a gene atlas for cultivated peanut ( Arachis hypogaea ), 22 different tissue types and ontogenies that represent the full development of peanut were sequenced, including a complete reproductive series from flower to peg elongation and peg tip immersion in the soil to fully mature seed. Using a genome-guided assembly pipeline, a homeolog-specific transcriptome assembly for Arachis hypogaea was assembled and its accuracy was validated. The assembly was used to annotate 21 developmental co-expression networks as tools for gene discovery. Using a set of 8816 putative homeologous gene pairs, homeolog expression bias was documented, and although bias was mostly balanced, there were striking differences in expression bias in a tissue-specific context. Over 9000 alterative splicing events and over 6000 non-coding RNAs were further identified and profiled in a developmental context. Together, this work represents a major new resource for cultivated peanut and will be integrated into peanutbase.org as an available resource for all peanut researchers.

Purpose: Dovitinib (TKI258) is an oral multi-tyrosine kinase inhibitor of FGFR, VEGFR, PDGFR β, and c-Kit. Since dovitinib is able to cross the blood-brain barrier and targets brain tumor-relevant pathways, we conducted a phase I trial to demonstrate its safety in recurrent glioblastoma (GBM)., Patients and Methods: Patients with first or second GBM recurrence started treatment with the maximal tolerated dose (MTD) previously established in systemic cancer patients (500 mg/d, 5 days on/2 days off). A modified 3 + 3 design in three cohorts (500, 400, 300 mg) was used., Results: Twelve patients were enrolled. Seventy-two adverse events (AEs) occurred and 16.7 % of AEs were classified as ≥CTC grade 3 toxicity, mainly including hepatotoxicity and hematotoxicity. Only one out of six patients of the 300-mg cohort showed grade 3 toxicity. The PFS-6 rate was 16.7 %, and it was not associated with detection of the FGFR-TACC gene fusion in the tumor., Conclusion: Dovitinib is safe in patients with recurrent GBM and showed efficacy in only some patients unselected for target expression. The recommended phase II dose of 300 mg would be substantially lower than the recently established MTD in systemic cancer patients. Further personalized trials are recommended.

Catfish represent 12% of teleost or 6.3% of all vertebrate species, and are of enormous economic value. Here we report a high-quality reference genome sequence of channel catfish (Ictalurus punctatus), the major aquaculture species in the US. The reference genome sequence was validated by genetic mapping of 54,000 SNPs, and annotated with 26,661 predicted protein-coding genes. Through comparative analysis of genomes and transcriptomes of scaled and scaleless fish and scale regeneration experiments, we address the genomic basis for the most striking physical characteristic of catfish, the evolutionary loss of scales and provide evidence that lack of secretory calcium-binding phosphoproteins accounts for the evolutionary loss of scales in catfish. The channel catfish reference genome sequence, along with two additional genome sequences and transcriptomes of scaled catfishes, provide crucial resources for evolutionary and biological studies. This work also demonstrates the power of comparative subtraction of candidate genes for traits of structural significance.

Purpose: Pseudoprogression (PsP) is characterized by therapy-associated but not tumor growth-associated increases of contrast-enhancing glioblastoma lesions on MRI. Although typically occurring during the first 3 months after radiochemotherapy, PsP may occur later in the course of the disease and may then be particularly difficult to distinguish from true tumor progression. We explored PET using O-(2-[(18)F]fluoroethyl)-L-tyrosine ((18)F-FET-PET) to approach the diagnostic dilemma., Experimental Design: Twenty-six patients with glioblastoma that presented with increasing contrast-enhancing lesions later than 3 months after completion of radiochemotherapy underwent (18)F-FET-PET. Maximum and mean tumor/brain ratios (TBRmax and TBRmean) of (18)F-FET uptake as well as time-to-peak (TTP) and patterns of the time-activity curves were determined. The final diagnosis of true progression versus late PsP was based on follow-up MRI using RANO criteria., Results: Late PsP occurred in 7 patients with a median time from radiochemotherapy completion of 24 weeks while the remaining patients showed true tumor progression. TBRmax and TBRmean were significantly higher in patients with true progression than in patients with late PsP (TBRmax 2.4 ± 0.1 vs. 1.5 ± 0.2, P = 0.003; TBRmean 2.1 ± 0.1 vs. 1.5 ± 0.2, P = 0.012) whereas TTP was significantly shorter (mean TTP 25 ± 2 vs. 40 ± 2 min, P < 0.001). ROC analysis yielded an optimal cutoff value of 1.9 for TBRmax to differentiate between true progression and late PsP (sensitivity 84%, specificity 86%, accuracy 85%, P = 0.015)., Conclusions: O-(2-[(18)F]fluoroethyl)-L-tyrosine PET provides valuable information in assessing the elusive phenomenon of late PsP. Clin Cancer Res; 22(9); 2190-6. ©2015 AACR., (©2015 American Association for Cancer Research.)

Orthognathic surgery has always been a classical focus of maxillofacial surgery. Since more than 100 years, various surgical techniques for mandibular repositioning have been developed and clinically tested. Since the establishment of plate and screw osteosynthesis, orthognathic surgery became more stable and safe. Nowadays, different surgical methods for mobilising the mandible are existing. This international multicenter analysis (n = 51 hospitals) is providing first evidence based data for the current use of different surgical methods. The dominating techniques were Obwegeser/dal Pont (61%) followed by Hunsuck/Epker (37%) and Perthes/Schlössmann (29%). The main osteosynthesis materials were plates (82%), bicortical screws (23.5%), or a combination of both (5.9%). 47% of all centers reported to use several surgical methods at the same time, depending on the anatomical problem and the surgeon's preference. This shows that different surgical methods seem to work as comparable, safe, and reliable procedures in everydays clinical practise. On this basis, further prospective studies could evaluate possible advantages for our patients., (Copyright © 2016 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.)

Weeds reduce yield in soybeans (Glycine max) through incompletely defined mechanisms. The effects of weeds on the soybean transcriptome were evaluated in field conditions during four separate growing seasons. RNASeq data were collected from six biological samples of soybeans growing with or without weeds. Weed species and the methods to maintain weed-free controls varied between years to mitigate treatment effects, and to allow detection of general soybean weed responses. Soybean plants were not visibly nutrient- or water-stressed. We identified 55 consistently downregulated genes in weedy plots. Many of the downregulated genes were heat shock genes. Fourteen genes were consistently upregulated. Several transcription factors including a PHYTOCHROME INTERACTING FACTOR 3-like gene (PIF3) were included among the upregulated genes. Gene set enrichment analysis indicated roles for increased oxidative stress and jasmonic acid signaling responses during weed stress. The relationship of this weed-induced PIF3 gene to genes involved in shade avoidance responses in Arabidopsis provide evidence that this gene may be important in the response of soybean to weeds. These results suggest that the weed-induced PIF3 gene will be a target for manipulating weed tolerance in soybean., (No claim to original US government works New Phytologist © 2015 New Phytologist Trust.)

The first nearly cryptic species of scorpionfly from the United States, Panorpa cryptica Bicha and Schiff, n. sp., is described from northern Georgia, southwestern North Carolina and northwestern South Carolina. This insect was initially differentiated from the very similar Panorpa nebulosa Westwood by its unique cytochrome oxidase subunit I (COI) mitochondrial DNA. Habitat details, distribution, and biology are described.

Current advances in sequencing technologies and bioinformatics revealed the genomic background of rice, a staple food for the poor people, and provided the basis to develop large genomic variation databases for thousands of cultivars. Proper analysis of this massive resource is expected to give novel insights into the structure, function, and evolution of the rice genome, and to aid the development of rice varieties through marker assisted selection or genomic selection. In this work we present sequencing and bioinformatics analyses of 104 rice varieties belonging to the major subspecies of Oryza sativa. We identified repetitive elements and recurrent copy number variation covering about 200 Mbp of the rice genome. Genotyping of over 18 million polymorphic locations within O. sativa allowed us to reconstruct the individual haplotype patterns shaping the genomic background of elite varieties used by farmers throughout the Americas. Based on a reconstruction of the alleles for the gene GBSSI, we could identify novel genetic markers for selection of varieties with high amylose content. We expect that both the analysis methods and the genomic information described here would be of great use for the rice research community and for other groups carrying on similar sequencing efforts in other crops.

All eukaryotic cells utilize autophagy for protein and organelle turnover, thus assuring subcellular quality control, homeostasis, and survival. In order to address recent advances in identification of human autophagy associated genes, and to describe autophagy on a system-wide level, we established an autophagy-centered gene interaction network by merging various primary data sets and by retrieving respective interaction data. The resulting network ('AXAN') was analyzed with respect to subnetworks, e.g. the prime gene subnetwork (including the core machinery, signaling pathways and autophagy receptors) and the transcription subnetwork. To describe aspects of evolution within this network, we assessed the presence of protein orthologs across 99 eukaryotic model organisms. We visualized evolutionary trends for prime gene categories and evolutionary tracks for selected AXAN genes. This analysis confirms the eukaryotic origin of autophagy core genes while it points to a diverse evolutionary history of autophagy receptors. Next, we used module identification to describe the functional anatomy of the network at the level of pathway modules. In addition to obvious pathways (e.g., lysosomal degradation, insulin signaling) our data unveil the existence of context-related modules such as Rho GTPase signaling. Last, we used a tripartite, image-based RNAi - screen to test candidate genes predicted to play a role in regulation of autophagy. We verified the Rho GTPase, CDC42, as a novel regulator of autophagy-related signaling. This study emphasizes the applicability of system-wide approaches to gain novel insights into a complex biological process and to describe the human autophagy pathway at a hitherto unprecedented level of detail.

Benzo[a]pyrene (BaP) is an established carcinogen and reproductive and developmental toxicant. BaP exposure in humans and animals has been linked to infertility and multigenerational health consequences. DNA methylation is the most studied epigenetic mechanism that regulates gene expression, and mapping of methylation patterns has become an important tool for understanding pathologic gene expression events. The goal of this study was to investigate aberrant changes in promoter DNA methylation in zebrafish embryos and larvae following a parental and continued embryonic waterborne BaP exposure. A total of 21 genes known for their role in human diseases were selected to measure percent methylation by multiplex deep sequencing. At 96hpf (hours post fertilization) compared to 3.3hpf, dazl, nqo1, sox3, cyp1b1, and gstp1 had higher methylation percentages while c-fos and cdkn1a had decreased CG methylation. BaP exposure significantly reduced egg production and offspring survival. Moreover, BaP decreased global methylation and altered CG, CHH, and CHG methylation both at 3.3 and 96hpf. CG methylation changed by 10% or more due to BaP in six genes (c-fos, cdkn1a, dazl, nqo1, nrf2, and sox3) at 3.3hpf and in ten genes (c-fos, cyp1b1, dazl, gstp1, mlh1, nqo1, pten, p53, sox2, and sox3) at 96hpf. BaP also induced gene expression of cyp1b1 and gstp1 at 96hpf which were found to be hypermethylated. Further studies are needed to link aberrant CG, CHH, and CHG methylation to heritable epigenetic consequences associated with disease in later life., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: After the failure of radiotherapy and temozolomide, there is no established standard therapy for patients with recurrent glioblastoma (GBM). Based on the promising data of a previous trial (NOA-01) for primary GBM and some retrospective case series for GBM recurrence, the combination of nimustine and teniposide (VM26) was commonly used in this setting. When nimustine was no longer available in Europe, we switched to intrvaveneous carmustine (BCNU). Data on the toxicity and efficacy of BCNU and VM26 in recurrent GBM are lacking., Methods: In our neurooncological center, all patients with recurrent GBM or with progressed glioma and a typical MRI lesion suggesting GBM treated with BCNU (130-150 mg/m(2), day 1/42) and VM26 (45-60 mg/m(2), days 1-3/42) were analyzed retrospectively for progression-free survival, overall survival and toxicity., Results: Fifteen patients (median age 52 years) were identified. Median progression-free survival was 2 months and median overall survival was 4 months. Two patients (14%) developed grade 3/4 hematotoxicity. Nonhematological toxicity ≥grade 3 was not observed., Conclusion: Our data do not support the application of BCNU/VM26 in patients with late stages of recurrent GBM., (© 2014 S. Karger AG, Basel.)

The extracellular matrix (ECM) protein tenascin-C (TN-C) is upregulated within glioma tissues and cultured glioma cell lines. TN-C possesses a multi-modular structure and a variety of functional properties have been reported for its domains. We describe five novel monoclonal antibodies identifying different domains of TN-C. The epitopes for these antibodies were investigated by using recombinantly expressed fibronectin type III domains of TN-C. The biological effects of TN-C fragments on glioma cell proliferation and adhesion were analyzed. The expression pattern of TN-C in human glioma tissue sections and in glioma cell lines was studied with the novel library of monoclonal antibodies. The immunocytochemical analyses of the established human glioma cell lines U-251-MG, U-373-MG and U-87-MG revealed distinct staining patterns for each antibody. Robust expression of TN-C was found within the tumor mass of surgery specimens from glioblastoma. In many cases, the expression of this ECM molecule was clearly associated with blood vessels, particularly with microvessels. Three of the new antibodies highlighted individual TN-C-expressing single cells in glioma tissues. The effect of TN-C domains on glioma cells was examined by a BrdU-proliferation assay and an adhesion assay. Short fragments of constitutively expressed TN-C-domains did not exert significant effects on the proliferation of glioma cells, whereas the intact molecule increased cell division rates. In contrast, the long fragment TNfnALL containing all of the FNIII domains of TN-C decreased proliferation. Additionally, we found strong differences between the adhesion-influencing properties of the recombinant fragments on glioma cells.

Glioblastoma (GBM) remains the most pervasive and lethal of all brain malignancies. One factor that contributes to this poor prognosis is the highly invasive character of the tumor. GBM is characterized by microscopic infiltration of tumor cells throughout the brain, whereas non-neural metastases, as well as select lower grade gliomas, develop as self-contained and clearly delineated lesions. Illustrated by rodent xenograft tumor models as well as pathological human patient specimens, we present evidence that one fundamental switch between these two distinct pathologies--invasion and noninvasion--is mediated through the tumor extracellular matrix. Specifically, noninvasive lesions are associated with a rich matrix containing substantial amounts of glycosylated chondroitin sulfate proteoglycans (CSPGs), whereas glycosylated CSPGs are essentially absent from diffusely infiltrating tumors. CSPGs, acting as central organizers of the tumor microenvironment, dramatically influence resident reactive astrocytes, inducing their exodus from the tumor mass and the resultant encapsulation of noninvasive lesions. Additionally, CSPGs induce activation of tumor-associated microglia. We demonstrate that the astrogliotic capsule can directly inhibit tumor invasion, and its absence from GBM presents an environment favorable to diffuse infiltration. We also identify the leukocyte common antigen-related phosphatase receptor (PTPRF) as a putative intermediary between extracellular glycosylated CSPGs and noninvasive tumor cells. In all, we present CSPGs as critical regulators of brain tumor histopathology and help to clarify the role of the tumor microenvironment in brain tumor invasion.

Purpose: Glioblastoma is a highly malignant, invariably fatal brain tumor for which effective pharmacotherapy remains an unmet medical need., Experimental Design: Screening of a compound library of 160 synthetic and natural toxic substances identified the antihelmintic niclosamide as a previously unrecognized candidate for clinical development. Considering the cellular and interindividual heterogeneity of glioblastoma, a portfolio of short-term expanded primary human glioblastoma cells (pGBM; n = 21), common glioma lines (n = 5), and noncancer human control cells (n = 3) was applied as a discovery platform and for preclinical validation. Pharmacodynamic analysis, study of cell-cycle progression, apoptosis, cell migration, proliferation, and on the frequency of multipotent/self-renewing pGBM cells were conducted in vitro, and orthotopic xenotransplantation was used to confirm anticancer effects in vivo., Results: Niclosamide led to cytostatic, cytotoxic, and antimigratory effects, strongly reduced the frequencies of multipotent/self-renewing cells in vitro, and after exposure significantly diminished the pGBMs' malignant potential in vivo. Mechanism of action analysis revealed that niclosamide simultaneously inhibited intracellular WNT/CTNNB1-, NOTCH-, mTOR-, and NF-κB signaling cascades. Furthermore, combinatorial drug testing established that a heterozygous deletion of the NFKBIA locus in glioblastoma samples could serve as a genomic biomarker for predicting a synergistic activity of niclosamide with temozolomide, the current standard in glioblastoma therapy., Conclusions: Together, our data advocate the use of pGBMs for exploration of compound libraries to reveal unexpected leads, for example, niclosamide that might be suited for further development toward personalized clinical application., (©2013 AACR.)

Premise of the Study: Microsatellite loci were isolated and characterized from enriched genomic libraries of Artocarpus altilis (breadfruit) and tested in four Artocarpus species and one hybrid. The microsatellite markers provide new tools for further studies in Artocarpus. •, Methods and Results: A total of 25 microsatellite loci were evaluated across four Artocarpus species and one hybrid. Twenty-one microsatellite loci were evaluated on A. altilis (241), A. camansi (34), A. mariannensis (15), and A. altilis × mariannensis (64) samples. Nine of those loci plus four additional loci were evaluated on A. heterophyllus (jackfruit, 426) samples. All loci are polymorphic for at least one species. The average number of alleles ranges from two to nine within taxa. •, Conclusions: These microsatellite primers will facilitate further studies on the genetic structure and evolutionary and domestication history of Artocarpus species. They will aid in cultivar identification and establishing germplasm conservation strategies for breadfruit and jackfruit.

Cellular heterogeneity, for example, the intratumoral coexistence of cancer cells with and without stem cell characteristics, represents a potential root of therapeutic resistance and a significant challenge for modern drug development in glioblastoma (GBM). We propose here that activation of the innate immune system by stimulation of innate immune receptors involved in antiviral and antitumor responses can similarly target different malignant populations of glioma cells. We used short-term expanded patient-specific primary human GBM cells to study the stimulation of the cytosolic nucleic acid receptors melanoma differentiation-associated gene 5 (MDA5) and retinoic acid-inducible gene I (RIG-I). Specifically, we analyzed cells from the tumor core versus "residual GBM cells" derived from the tumor resection margin as well as stem cell-enriched primary cultures versus specimens without stem cell properties. A portfolio of human, nontumor neural cells was used as a control for these studies. The expression of RIG-I and MDA5 could be induced in all of these cells. Receptor stimulation with their respective ligands, p(I:C) and 3pRNA, led to in vitro evidence for an effective activation of the innate immune system. Most intriguingly, all investigated cancer cell populations additionally responded with a pronounced induction of apoptotic signaling cascades revealing a second, direct mechanism of antitumor activity. By contrast, p(I:C) and 3pRNA induced only little toxicity in human nonmalignant neural cells. Granted that the challenge of effective central nervous system (CNS) delivery can be overcome, targeting of RIG-I and MDA5 could thus become a quintessential strategy to encounter heterogeneous cancers in the sophisticated environments of the brain., (Copyright © 2013 AlphaMed Press.)

Acyl-CoA binding proteins (ACBPs) have been identified in most branches of life, and play various roles in lipid metabolism, among other functions. Plants contain multiple classes of ACBP genes. The most diverse group is the class III proteins. Tung tree (Vernicia fordii) contains two such genes, designated VfACBP3A and VfACBP3B. The two proteins are significantly different in length and sequence. Analysis of tung ACBP3 genes revealed significant evolution, suggesting relatively ancient divergence of the two genes from a common ancestor. Phylogenetic comparisons of multiple plant class III proteins suggest that this group is the most evolutionarily dynamic class of ACBP. Both tung ACBP3 genes are expressed at similar levels in most tissues tested, but ACBP3A is stronger in leaves. Three-dimensional modeling predictions confirmed the presence of the conserved four α-helix bundle acyl-CoA binding (ACB); however, other regions of these proteins likely fold much differently. Acyl-CoA binding assays revealed different affinities for different acyl-CoAs, possibly contradicting the redundancy of function suggested by the gene expression studies. Subcellular targeting of transiently-expressed plant ACBP3 proteins contradicted earlier studies, and suggested that at least some class III ACBPs may be predominantly targeted to endoplasmic reticulum membranes, with little or no targeting to the apoplast., (Published by Elsevier Ireland Ltd.)

Objectives: In this study, we analyzed the prognostic value of different MRI progression patterns for survival in patients with recurrent malignant glioma treated with the vascular endothelial growth factor antibody bevacizumab., Patients and Methods: Twenty-six adult patients with recurrent malignant glioma treated with bevacizumab or bevacizumab/irinotecan were retrospectively analyzed for the development of contrast-enhanced (T1-weighted MRI) and T2/FLAIR lesions. According to the progression pattern, patients were divided into 3 subgroups: (1) patients with primarily progressive contrast-enhanced lesions in the first MRI after initiation of therapy ('primary PD group'); (2) patients with stable or regressive enhanced lesions but progressive FLAIR lesions ('FLAIR-only PD group'), and (3) patients with stable or regressive contrast-enhanced T1 and FLAIR lesions ('no PD group')., Results: Overall survival (OS) in the 6 patients in the FLAIR-only PD group was not significantly different from the 11 patients in the no PD group (median 311 vs. 254 days, respectively). In contrast, survival in the FLAIR-only PD group was significantly better (p = 0.025) than in the primary PD group., Conclusion: FLAIR-only progression is not an independent prognostic factor negatively influencing OS in recurrent glioblastoma treated with bevacizumab and should not lead to discontinuation of bevacizumab therapy., (Copyright © 2013 S. Karger AG, Basel.)

Alterations of DNA methylation play an important role in gliomas. In a genome-wide screen, we identified a CpG-rich fragment within the 5' region of the tumor necrosis factor receptor superfamily, member 11A gene (TNFRSF11A) that showed de novo methylation in gliomas. TNFRSF11A, also known as receptor activator of NF-κB (RANK), activates several signaling pathways, such as NF-κB, JNK, ERK, p38α, and Akt/PKB. Using pyrosequencing, we detected RANK/TNFRSF11A promoter methylation in 8 (57.1%) of 14 diffuse astrocytomas, 17 (77.3%) of 22 anaplastic astrocytomas, 101 (84.2%) of 120 glioblastomas, 6 (100%) of 6 glioma cell lines, and 7 (100%) of 7 glioma stem cell-enriched glioblastoma primary cultures but not in four normal white matter tissue samples. Treatment of glioma cell lines with the demethylating agent 5-aza-2'-deoxycytidine significantly reduced the methylation level and resulted in increased RANK/TNFRSF11A mRNA expression. Overexpression of RANK/TNFRSF11A in glioblastoma cell lines leads to a significant reduction in focus formation and elevated apoptotic activity after flow cytometric analysis. Reporter assay studies of transfected glioma cells supported these results by showing the activation of signaling pathways associated with regulation of apoptosis. We conclude that RANK/TNFRSF11A is a novel and frequent target for de novo methylation in gliomas, which affects apoptotic activity and focus formation thereby contributing to the molecular pathogenesis of gliomas.