Glioblastoma is characterized by widespread genetic and transcriptional heterogeneity, yet little is known about the role of the epigenome in glioblastoma disease progression. Here, we present ...genome-scale maps of DNA methylation in matched primary and recurring glioblastoma tumors, using data from a highly annotated clinical cohort that was selected through a national patient registry. We demonstrate the feasibility of DNA methylation mapping in a large set of routinely collected FFPE samples, and we validate bisulfite sequencing as a multipurpose assay that allowed us to infer a range of different genetic, epigenetic, and transcriptional characteristics of the profiled tumor samples. On the basis of these data, we identified subtle differences between primary and recurring tumors, links between DNA methylation and the tumor microenvironment, and an association of epigenetic tumor heterogeneity with patient survival. In summary, this study establishes an open resource for dissecting DNA methylation heterogeneity in a genetically diverse and heterogeneous cancer, and it demonstrates the feasibility of integrating epigenomics, radiology, and digital pathology for a national cohort, thereby leveraging existing samples and data collected as part of routine clinical practice.
Maximal safe tumor resection remains the key prognostic factor for improved prognosis in brain tumor patients. Despite 5-aminolevulinic acid-based fluorescence guidance the neurosurgeon is, however, ...not able to visualize most low-grade gliomas (LGG) and infiltration zone of high-grade gliomas (HGG). To overcome the need for a more sensitive visualization, we investigated the potential of macroscopic, wide-field fluorescence lifetime imaging of nicotinamide adenine dinucleotide (NADH) and protoporphyrin IX (PPIX) in selected human brain tumors. For future intraoperative use, the imaging system offered a square field of view of 11 mm at 250 mm free working distance. We performed imaging of tumor tissue ex vivo, including LGG and HGG as well as brain metastases obtained from 21 patients undergoing fluorescence-guided surgery. Half of all samples showed visible fluorescence during surgery, which was associated with significant increase in PPIX fluorescence lifetime. While the PPIX lifetime was significantly different between specific tumor tissue types, the NADH lifetimes did not differ significantly among them. However, mainly necrotic areas exhibited significantly lower NADH lifetimes compared to compact tumor in HGG. Our pilot study indicates that combined fluorescence lifetime imaging of NADH/PPIX represents a sensitive tool to visualize brain tumor tissue not detectable with conventional 5-ALA fluorescence.
The factors driving therapy resistance in diffuse glioma remain poorly understood. To identify treatment-associated cellular and genetic changes, we analyzed RNA and/or DNA sequencing data from the ...temporally separated tumor pairs of 304 adult patients with isocitrate dehydrogenase (IDH)-wild-type and IDH-mutant glioma. Tumors recurred in distinct manners that were dependent on IDH mutation status and attributable to changes in histological feature composition, somatic alterations, and microenvironment interactions. Hypermutation and acquired CDKN2A deletions were associated with an increase in proliferating neoplastic cells at recurrence in both glioma subtypes, reflecting active tumor growth. IDH-wild-type tumors were more invasive at recurrence, and their neoplastic cells exhibited increased expression of neuronal signaling programs that reflected a possible role for neuronal interactions in promoting glioma progression. Mesenchymal transition was associated with the presence of a myeloid cell state defined by specific ligand-receptor interactions with neoplastic cells. Collectively, these recurrence-associated phenotypes represent potential targets to alter disease progression.
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•Longitudinal glioma evolution follows an IDH mutation-dependent trajectory•Hypermutation and CDKN2A deletions underlie increased proliferation at recurrence•Recurrent IDH-wild-type neoplastic cells up-regulate neuronal signaling programs•Mesenchymal transitions associate with distinct myeloid cell interactions
Integrating longitudinal transcriptomic and genomic data from paired diffuse glioma samples with complementary single-cell RNA-seq and multiplex immunofluorescence datasets reveals recurrence-associated genetic and microenvironmental changes that are dependent on IDH mutation status.
Intraoperative identification of anaplastic foci in diffusely infiltrating gliomas (DIG) with non-significant contrast-enhancement on MRI is indispensible to avoid histopathological undergrading and ...subsequent treatment failure. Recently, we found that 5-aminolevulinic acid (5-ALA) induced protoporphyrin IX (PpIX) fluorescence can visualize areas with increased proliferative and metabolic activity in such gliomas intraoperatively. As treatment of DIG is predominantely based on histopathological World Health Organisation (WHO) parameters, we analyzed whether PpIX fluorescence can detect anaplastic foci according to these criteria.
We prospectively included DIG patients with non-significant contrast-enhancement that received 5-ALA prior to resection. Intraoperatively, multiple samples from PpIX positive and negative intratumoral areas were collected using a modified neurosurgical microscope. In all samples, histopathological WHO criteria and proliferation rate were assessed and correlated to the PpIX fluorescence status.
A total of 215 tumor specimens were collected in 59 patients. Of 26 WHO grade III gliomas, 23 cases (85%) showed focal PpIX fluorescence, whereas 29 (91%) of 33 WHO grade II gliomas were PpIX negative. In intratumoral areas with focal PpIX fluorescence, mitotic rate, cell density, nuclear pleomorphism, and proliferation rate were significantly higher than in non-fluorescing areas. The positive predictive value of focal PpIX fluorescence for WHO grade III histology was 85%.
Our study indicates that 5-ALA induced PpIX fluorescence is a powerful marker for intraoperative identification of anaplastic foci according to the histopathological WHO criteria in DIG with non-significant contrast-enhancement. Therefore, application of 5-ALA optimizes tissue sampling for precise histopathological diagnosis independent of brain-shift.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Incomplete neurosurgical resection of brain metastases (BM) due to insufficient intraoperative visualization of tumor tissue is a major clinical challenge and might result in local recurrence. ...Recently, visible 5-aminolevulinic acid (5-ALA) induced fluorescence was first reported in patients with BM. The aim of this study was thus to investigate, for the first time systematically, the value of 5-ALA fluorescence for intraoperative visualization of BM in a large patient cohort.
Adult patients (≥ 18 years) with resection of suspected BM after preoperative 5-ALA administration were prospectively recruited at two specialized neurosurgical centers. During surgery, the fluorescence status (visible or no fluorescence); fluorescence quality (strong, vague, or none); and fluorescence homogeneity (homogeneous or heterogeneous) of each BM was investigated. Additionally, these specific fluorescence characteristics of BM were correlated with the primary tumor type and the histopathological subtype. Tumor diagnosis was established according to the current WHO 2016 criteria.
Altogether, 157 BM were surgically treated in 154 patients. Visible fluorescence was observed in 104 BM (66%), whereas fluorescence was absent in the remaining 53 cases (34%). In detail, 53 tumors (34%) showed strong fluorescence, 51 tumors (32%) showed vague fluorescence, and 53 tumors (34%) had no fluorescence. The majority of BM (84% of cases) demonstrated a heterogeneous fluorescence pattern. According to primary tumor, visible fluorescence was less frequent in BM of melanomas compared to all other tumors (p = 0.037). According to histopathological subtype, visible fluorescence was more common in BM of ductal breast cancer than all other subtypes (p = 0.008). It is of note that visible fluorescence was observed in the surrounding brain tissue after the resection of BM in 74 (67%) of 111 investigated cases as well.
In this largest series to date, visible 5-ALA fluorescence was detected in two-thirds of BM. However, the characteristic heterogeneous fluorescence pattern and frequent lack of strong fluorescence limits the use of 5-ALA in BM and thus this technique needs further improvements.
OBJECTIVE Glioblastoma (GBM) is characterized by distinct intratumoral histopathological heterogeneity with regard to variable tumor morphology, cell proliferation, and microvascularity. Maximum ...resection of a GBM results in an improved prognosis and thus represents the aim of surgery in the majority of cases. Fluorescence-guided surgery using 5-aminolevulinic acid (5-ALA) is currently widely applied for improved intraoperative tumor visualization in patients with a GBM. Three intratumoral fluorescence levels (i.e., strong, vague, or no fluorescence) can usually be distinguished during surgery. So far, however, their exact histopathological correlates and their surgical relevance have not been clarified sufficiently. Thus, the aim of this study was to systematically analyze tissue samples from newly diagnosed GBMs with different fluorescence levels according to relevant histopathological parameters. METHODS This prospective study recruited patients who underwent 5-ALA fluorescence-guided resection of a newly diagnosed radiologically suspected GBM. Each patient received 5-ALA approximately 3 hours before surgery, and a modified neurosurgical microscope was applied for intraoperative visualization of 5-ALA-induced fluorescence. During surgery, tissue samples with strong, vague, or no fluorescence were collected. For each sample, the presence of tumor tissue, quality of tissue (compact, infiltrative, or no tumor), histopathological criteria of malignancy (cell density, nuclear pleomorphism, mitotic activity, and presence of microvascular proliferation/necrosis), proliferation rate (MIB-1 labeling index LI), and microvessel density (using CD34 staining) were investigated. RESULTS Altogether, 77 patients with a newly diagnosed, histopathologically confirmed GBM were included, and 131 samples with strong fluorescence, 69 samples with vague fluorescence, and 67 samples with no fluorescence were collected. Tumor tissue was detected in all 131 (100%) of the samples with strong fluorescence and in 65 (94%) of the 69 samples with vague fluorescence. However, mostly infiltrative tumor tissue was still found in 33 (49%) of 67 samples despite their lack of fluorescence. Strong fluorescence corresponded to compact tumors in 109 (83%) of 131 samples, whereas vague fluorescence was consistent with infiltrative tumors in 44 (64%) of 69 samples. In terms of the histopathological criteria of malignancy, a significant positive correlation of all analyzed parameters comprising cell density, nuclear pleomorphism, mitotic activity, microvascular proliferation, and necrosis with the 3 fluorescence levels was observed (p < 0.001). Furthermore, the proliferation rate significantly and positively correlated with strong (MIB-1 LI 28.3%), vague (MIB-1 LI 16.7%), and no (MIB-1 LI 8.8%) fluorescence (p < 0.001). Last, a significantly higher microvessel density was detected in samples with strong fluorescence (CD34 125.5 vessels/0.25 mm
) than in those with vague (CD34 82.8 vessels/0.25 mm
) or no (CD34 68.6 vessels/0.25 mm
) fluorescence (p < 0.001). CONCLUSIONS Strong and vague 5-ALA-induced fluorescence enables visualization of intratumoral areas with specific histopathological features and thus supports neurosurgeons in improving the extent of resection in patients with a newly diagnosed GBM. Despite the lack of fluorescence, tumor tissue was still observed in approximately half of the cases. To overcome this current limitation, the promising approach of complementary spectroscopic measurement of fluorescence should be investigated further.
Adult diffuse gliomas are a diverse group of brain neoplasms that inflict a high emotional toll on patients and their families. The Cancer Genome Atlas and similar projects have provided a ...comprehensive understanding of the somatic alterations and molecular subtypes of glioma at diagnosis. However, gliomas undergo significant cellular and molecular evolution during disease progression. We review the current knowledge on the genomic and epigenetic abnormalities in primary tumors and after disease recurrence, highlight the gaps in the literature, and elaborate on the need for a new multi-institutional effort to bridge these knowledge gaps and how the Glioma Longitudinal Analysis Consortium (GLASS) aims to systemically catalog the longitudinal changes in gliomas. The GLASS initiative will provide essential insights into the evolution of glioma toward a lethal phenotype, with the potential to reveal targetable vulnerabilities and, ultimately, improved outcomes for a patient population in need.
A common coordinate space enabling comparison across individuals is vital to understanding human brain organization and individual differences. By leveraging dimensionality reduction algorithms, ...high-dimensional fMRI data can be represented in a low-dimensional space to characterize individual features. Such a representative space encodes the functional architecture of individuals and enables the observation of functional changes across time. However, determining comparable functional features across individuals in resting-state fMRI in a way that simultaneously preserves individual-specific connectivity structure can be challenging. In this work we propose scalable joint embedding to simultaneously embed multiple individual brain connectomes within a common space that allows individual representations across datasets to be aligned. Using Human Connectome Project data, we evaluated the joint embedding approach by comparing it to the previously established orthonormal alignment model. Alignment using joint embedding substantially increased the similarity of functional representations across individuals while simultaneously capturing their distinct profiles, allowing individuals to be more discriminable from each other. Additionally, we demonstrated that the common space established using resting-state fMRI provides a better overlap of task-activation across participants. Finally, in a more challenging scenario - alignment across a lifespan cohort aged from 6 to 85 - joint embedding provided a better prediction of age (r2 = 0.65) than the prior alignment model. It facilitated the characterization of functional trajectories across lifespan. Overall, these analyses establish that joint embedding can simultaneously capture individual neural representations in a common connectivity space aligning functional data across participants and populations and preserve individual specificity.
Currently, approximately 150 different brain tumour types are defined by the WHO. Recent endeavours to exploit machine learning and deep learning methods for supporting more precise diagnostics based ...on the histological tumour appearance have been hampered by the relative paucity of accessible digital histopathological datasets. While freely available datasets are relatively common in many medical specialties such as radiology and genomic medicine, there is still an unmet need regarding histopathological data. Thus, we digitized a significant portion of a large dedicated brain tumour bank based at the Division of Neuropathology and Neurochemistry of the Medical University of Vienna, covering brain tumour cases from 1995-2019. A total of 3,115 slides of 126 brain tumour types (including 47 control tissue slides) have been scanned. Additionally, complementary clinical annotations have been collected for each case. In the present manuscript, we thoroughly discuss this unique dataset and make it publicly available for potential use cases in machine learning and digital image analysis, teaching and as a reference for external validation.
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