Identification of a single-dose, low flip angle based CBV threshold for fractional tumor burden (FTB) mapping in recurrent glioblastoma.

BACKGROUND AND PURPOSE: DSC-MRI can be used to generate fractional tumor burden (FTB) maps, via application of relative CBV thresholds, to spatially differentiate glioblastoma recurrence from post treatment radiation effects (PTRE). Image-localized histopathology was previously used to validate FTB maps derived from a reference DSC-MRI protocol using preload, a moderate flip angle (MFA, 60°) and post-processing leakage correction. Recently, a DSC-MRI protocol with a low flip angle (LFA, 30°) with no preload was shown to provide leakage-corrected RCBV equivalent to the reference protocol. This study aims to identify the RCBV thresholds for the LFA protocol that generate the most accurate FTB maps, concordant with those obtained from the reference MFA protocol. MATERIALS AND METHODS: Fifty-two patients with grade IV GBM who had prior surgical resection and received chemotherapy and radiotherapy were included in the study. Two sets of DSC-MRI data were collected sequentially first using LFA protocol with no preload, which served as the preload for the subsequent MFA protocol. Standardized relative CBV maps (sRCBV) were obtained for each patient and co-registered with the anatomical post-contrast T1-weighted images. The reference MFA-based FTB maps were computed using previously published sRCBV thresholds (1.0 and 1.56). An ROC analysis was conducted to identify the optimal, voxelwise LFA sRCBV thresholds, and the sensitivity, specificity, and accuracy of the LFA-based FTB maps were computed with respect to the MFA-based reference. RESULTS: The mean sRCBV values of tumors across patients exhibited strong agreement (CCC = 0.99) between the two protocols. Using the ROC analysis, the optimal lower LFA threshold that accurately distinguishes PTRE from tumor recurrence was found to be 1.0 (sensitivity: 87.77%; specificity: 90.22%), equivalent to the ground truth. To identify aggressive tumor regions, the ROC analysis identified an upper LFA threshold of 1.37 (sensitivity: 90.87%; specificity: 91.10%) for the reference MFA threshold of 1.56. CONCLUSION: For LFA-based FTB maps, a sRCBV threshold of 1.0 and 1.37 can differentiate PTRE from recurrent tumor. FTB maps aids in surgical planning, guiding pathological diagnosis and treatment strategies in the recurrent setting. This study further confirms the reliability of single-dose LFA-based DSC-MRI. ABBREVIATIONS: LFA = low flip angle; MFA = moderate flip angle; sRCBV = standardized relative cerebral blood volume; FTB = fractional tumor burden; PTRE = post treatment radiation effects; ROC = receiver operating characteristics; CCC = concordance correlation coefficient.

Intraoperative in vivo confocal laser endomicroscopy imaging at glioma margins: can we detect tumor infiltration?

OBJECTIVE: Confocal laser endomicroscopy (CLE) is a US Food and Drug Administration-cleared intraoperative real-time fluorescence-based cellular resolution imaging technology that has been shown to image brain tumor histoarchitecture rapidly in vivo during neuro-oncological surgical procedures. An important goal for successful intraoperative implementation is in vivo use at the margins of infiltrating gliomas. However, CLE use at glioma margins has not been well studied. METHODS: Matching in vivo CLE images and tissue biopsies acquired at glioma margin regions of interest (ROIs) were collected from 2 institutions. All images were reviewed by 4 neuropathologists experienced in CLE. A scoring system based on the pathological features was implemented to score CLE and H&E images from each ROI on a scale from 0 to 5. Based on the H&E scores, all ROIs were divided into a low tumor probability (LTP) group (scores 0-2) and a high tumor probability (HTP) group (scores 3-5). The concordance between CLE and H&E scores regarding tumor probability was determined. The intraclass correlation coefficient (ICC) and diagnostic performance were calculated. RESULTS: Fifty-six glioma margin ROIs were included for analysis. Interrater reliability of the scoring system was excellent when used for H&E images (ICC [95% CI] 0.91 [0.86-0.94]) and moderate when used for CLE images (ICC [95% CI] 0.69 [0.40-0.83]). The ICCs (95% CIs) of the LTP group (0.68 [0.40-0.83]) and HTP group (0.68 [0.39-0.83]) did not differ significantly. The concordance between CLE and H&E scores was 61.6%. The sensitivity and specificity values of the scoring system were 79% and 37%. The positive predictive value (PPV) and negative predictive value were 65% and 53%, respectively. Concordance, sensitivity, and PPV were greater in the HTP group than in the LTP group. Specificity was higher in the newly diagnosed group than in the recurrent group. CONCLUSIONS: CLE may detect tumor infiltration at glioma margins. However, it is not currently dependable, especially in scenarios where low probability of tumor infiltration is expected. The proposed scoring system has excellent intrinsic interrater reliability, but its interrater reliability is only moderate when used with CLE images. These results suggest that this technology requires further exploration as a method for consistent actionable intraoperative guidance with high dependability across the range of tumor margin scenarios. Specific-binding and/or tumor-specific fluorophores, a CLE image atlas, and a consensus guideline for image interpretation may help with the translational utility of CLE.

Intraoperative confocal laser endomicroscopy for interpretation of a sellar hemangioblastoma: illustrative case.

BACKGROUND: Intraoperative frozen sections play a critical role in surgical strategy because of their ability to provide rapid histopathological information. In cases in which intraoperative biopsy carries a significant risk of bleeding, intraoperative confocal laser endomicroscopy (CLE) can assist in decision-making. OBSERVATIONS: The authors present a rare case of a large sellar hemangioblastoma. Preoperative radiographic imaging and normal pituitary function suggested a differential diagnosis that included hemangioblastoma. The patient underwent partial preoperative embolization and a right-sided pterional craniotomy for resection of the lesion. Gross intraoperative examination revealed a highly vascular sellar lesion requiring circumferential dissection to minimize blood loss. The serious vascularity precluded intraoperative frozen section analysis, and CLE imaging was performed. CLE imaging provided excellent visualization of the remarkable vascular structure and characteristic histoarchitecture with microvasculature, intracytoplasmic vacuoles, and atypical cells consistent with hemangioblastoma. Resection and decompression of the chiasm was accomplished, and the patient was discharged with improved vision. The final histopathological diagnosis was hemangioblastoma. LESSONS: When the benefits of obtaining intraoperative frozen sections greatly outweigh the associated risks, CLE imaging can aid in decision-making. CLE imaging offers real-time, on-the-fly evaluation of intraoperative tissue without the need to biopsy a vascular lesion.

Integrated molecular and multiparametric MRI mapping of high-grade glioma identifies regional biologic signatures.

Sampling restrictions have hindered the comprehensive study of invasive non-enhancing (NE) high-grade glioma (HGG) cell populations driving tumor progression. Here, we present an integrated multi-omic analysis of spatially matched molecular and multi-parametric magnetic resonance imaging (MRI) profiling across 313 multi-regional tumor biopsies, including 111 from the NE, across 68 HGG patients. Whole exome and RNA sequencing uncover unique genomic alterations to unresectable invasive NE tumor, including subclonal events, which inform genomic models predictive of geographic evolution. Infiltrative NE tumor is alternatively enriched with tumor cells exhibiting neuronal or glycolytic/plurimetabolic cellular states, two principal transcriptomic pathway-based glioma subtypes, which respectively demonstrate abundant private mutations or enrichment in immune cell signatures. These NE phenotypes are non-invasively identified through normalized K2 imaging signatures, which discern cell size heterogeneity on dynamic susceptibility contrast (DSC)-MRI. NE tumor populations predicted to display increased cellular proliferation by mean diffusivity (MD) MRI metrics are uniquely associated with EGFR amplification and CDKN2A homozygous deletion. The biophysical mapping of infiltrative HGG potentially enables the clinical recognition of tumor subpopulations with aggressive molecular signatures driving tumor progression, thereby informing precision medicine targeting.

Genetically engineered human pituitary corticotroph tumor organoids exhibit divergent responses to glucocorticoid receptor modulators.

Cushing's disease (CD) is a serious endocrine disorder attributed to an adrenocorticotropic hormone (ACTH)-secreting pituitary neuroendocrine tumor (PitNET) that that subsequently leads to chronic hypercortisolemia. PitNET regression has been reported following treatment with the investigational selective glucocorticoid receptor (GR) modulator relacorilant, but the mechanisms behind that effect remain unknown. Human PitNET organoid models were generated from induced human pluripotent stem cells (iPSCs) or fresh tissue obtained from CD patient PitNETs (hPITOs). Genetically engineered iPSC derived organoids were used to model the development of corticotroph PitNETs expressing USP48 (iPSCUSP48) or USP8 (iPSCUSP8) somatic mutations. Organoids were treated with the GR antagonist mifepristone or the GR modulator relacorilant with or without somatostatin receptor (SSTR) agonists pasireotide or octreotide. In iPSCUSP48 and iPSCUSP8 cultures, mifepristone induced a predominant expression of SSTR2 with a concomitant increase in ACTH secretion and tumor cell proliferation. Relacorilant predominantly induced SSTR5 expression and tumor cell apoptosis with minimal ACTH induction. Hedgehog signaling mediated the induction of SSTR2 and SSTR5 in response to mifepristone and relacorilant. Relacorilant sensitized PitNET organoid responsiveness to pasireotide. Therefore, our study identified the potential therapeutic use of relacorilant in combination with somatostatin analogs and demonstrated the advantages of relacorilant over mifepristone, supporting its further development for use in the treatment of Cushing's disease patients.

Intraoperative confocal laser endomicroscopy: prospective in vivo feasibility study of a clinical-grade system for brain tumors.

OBJECTIVE: The authors evaluated the feasibility of using the first clinical-grade confocal laser endomicroscopy (CLE) system using fluorescein sodium for intraoperative in vivo imaging of brain tumors. METHODS: A CLE system cleared by the FDA was used in 30 prospectively enrolled patients with 31 brain tumors (13 gliomas, 5 meningiomas, 6 other primary tumors, 3 metastases, and 4 reactive brain tissue). A neuropathologist classified CLE images as interpretable or noninterpretable. Images were compared with corresponding frozen and permanent histology sections, with image correlation to biopsy location using neuronavigation. The specificities and sensitivities of CLE images and frozen sections were calculated using permanent histological sections as the standard for comparison. A recently developed surgical telepathology software platform was used in 11 cases to provide real-time intraoperative consultation with a neuropathologist. RESULTS: Overall, 10,713 CLE images from 335 regions of interest were acquired. The mean duration of the use of the CLE system was 7 minutes (range 3-18 minutes). Interpretable CLE images were obtained in all cases. The first interpretable image was acquired within a mean of 6 (SD 10) images and within the first 5 (SD 13) seconds of imaging; 4896 images (46%) were interpretable. Interpretable image acquisition was positively correlated with study progression, number of cases per surgeon, cumulative length of CLE time, and CLE time per case (p ≤ 0.01). The diagnostic accuracy, sensitivity, and specificity of CLE compared with frozen sections were 94%, 94%, and 100%, respectively, and the diagnostic accuracy, sensitivity, and specificity of CLE compared with permanent histological sections were 92%, 90%, and 94%, respectively. No difference was observed between lesion types for the time to first interpretable image (p = 0.35). Deeply located lesions were associated with a higher percentage of interpretable images than superficial lesions (p = 0.02). The study met the primary end points, confirming the safety and feasibility and acquisition of noninvasive digital biopsies in all cases. The study met the secondary end points for the duration of CLE use necessary to obtain interpretable images. A neuropathologist could interpret the CLE images in 29 (97%) of 30 cases. CONCLUSIONS: The clinical-grade CLE system allows in vivo, intraoperative, high-resolution cellular visualization of tissue microstructure and identification of lesional tissue patterns in real time, without the need for tissue preparation.

Larotrectinib in NTRK Fusion-Positive High-Grade Glioneuronal Tumor: A Case Report.

Glioneuronal tumors are rare central nervous system tumors with heterogeneous histological and molecular features. While the majority are low grade, a small percentage can behave aggressively. Due to the rarity of these tumors, there is no consensus on how to treat high-grade glioneuronal tumors, and they are often managed similarly to glial tumors. With the advent of molecular profiling, management decisions are increasingly determined by molecular alterations in the tumor rather than the tumor type, which can be a useful approach for tumor types that do not have robust supportive clinical trial data due to low prevalence. We present a case of an 18-year-old patient with a high-grade glioneuronal neoplasm initially treated with craniospinal irradiation, vincristine, and cyclophosphamide. He presented eight years later with a recurrent tumor and was found to be positive for MEF2D-NTRK1 fusion. He was treated with surgical resection and postoperative intensity-modulated radiation therapy (IMRT; 55.8 Gy) with concurrent temozolomide, followed by the NTRK inhibitor larotrectinib. He achieved a radiographic response, with a decrease in residual enhancement and radiographic improvement over the course of treatment. He remained in clinical and radiographic remission for six months. This demonstrates the successful treatment of a high-grade glioneuronal NTRK fusion-positive tumor with larotrectinib, which has only been previously reported once in the literature.

Development of Human Pituitary Neuroendocrine Tumor Organoids to Facilitate Effective Targeted Treatments of Cushing's Disease.

(1) Background: Cushing's disease (CD) is a serious endocrine disorder caused by an adrenocorticotropic hormone (ACTH)-secreting pituitary neuroendocrine tumor (PitNET) that stimulates the adrenal glands to overproduce cortisol. Chronic exposure to excess cortisol has detrimental effects on health, including increased stroke rates, diabetes, obesity, cognitive impairment, anxiety, depression, and death. The first-line treatment for CD is pituitary surgery. Current surgical remission rates reported in only 56% of patients depending on several criteria. The lack of specificity, poor tolerability, and low efficacy of the subsequent second-line medical therapies make CD a medical therapeutic challenge. One major limitation that hinders the development of specific medical therapies is the lack of relevant human model systems that recapitulate the cellular composition of PitNET microenvironment. (2) Methods: human pituitary tumor tissue was harvested during transsphenoidal surgery from CD patients to generate organoids (hPITOs). (3) Results: hPITOs generated from corticotroph, lactotroph, gonadotroph, and somatotroph tumors exhibited morphological diversity among the organoid lines between individual patients and amongst subtypes. The similarity in cell lineages between the organoid line and the patient's tumor was validated by comparing the neuropathology report to the expression pattern of PitNET specific markers, using spectral flow cytometry and exome sequencing. A high-throughput drug screen demonstrated patient-specific drug responses of hPITOs amongst each tumor subtype. Generation of induced pluripotent stem cells (iPSCs) from a CD patient carrying germline mutation CDH23 exhibited dysregulated cell lineage commitment. (4) Conclusions: The human pituitary neuroendocrine tumor organoids represent a novel approach in how we model complex pathologies in CD patients, which will enable effective personalized medicine for these patients.

Characterization of ex vivo and in vivo intraoperative neurosurgical confocal laser endomicroscopy imaging.

Background: The new US Food and Drug Administration-cleared fluorescein sodium (FNa)-based confocal laser endomicroscopy (CLE) imaging system allows for intraoperative on-the-fly cellular level imaging. Two feasibility studies have been completed with intraoperative use of this CLE system in ex vivo and in vivo modalities. This study quantitatively compares the image quality and diagnostic performance of ex vivo and in vivo CLE imaging. Methods: Images acquired from two prospective CLE clinical studies, one ex vivo and one in vivo, were analyzed quantitatively. Two image quality parameters - brightness and contrast - were measured using Fiji software and compared between ex vivo and in vivo images for imaging timing from FNa dose and in glioma, meningioma, and intracranial metastatic tumor cases. The diagnostic performance of the two studies was compared. Results: Overall, the in vivo images have higher brightness and contrast than the ex vivo images (p < 0.001). A weak negative correlation exists between image quality and timing of imaging after FNa dose for the ex vivo images, but not the in vivo images. In vivo images have higher image quality than ex vivo images (p < 0.001) in glioma, meningioma, and intracranial metastatic tumor cases. In vivo imaging yielded higher sensitivity and negative predictive value than ex vivo imaging. Conclusions: In our setting, in vivo CLE optical biopsy outperforms ex vivo CLE by producing higher quality images and less image deterioration, leading to better diagnostic performance. These results support the in vivo modality as the modality of choice for intraoperative CLE imaging.

Real-time intraoperative surgical telepathology using confocal laser endomicroscopy.

OBJECTIVE: Communication between neurosurgeons and pathologists is mandatory for intraoperative decision-making and optimization of resection, especially for invasive masses. Handheld confocal laser endomicroscopy (CLE) technology provides in vivo intraoperative visualization of tissue histoarchitecture at cellular resolution. The authors evaluated the feasibility of using an innovative surgical telepathology software platform (TSP) to establish real-time, on-the-fly remote communication between the neurosurgeon using CLE and the pathologist. METHODS: CLE and a TSP were integrated into the surgical workflow for 11 patients with brain masses (6 patients with gliomas, 3 with other primary tumors, 1 with metastasis, and 1 with reactive brain tissue). Neurosurgeons used CLE to generate video-flow images of the operative field that were displayed on monitors in the operating room. The pathologist simultaneously viewed video-flow CLE imaging using a digital tablet and communicated with the surgeon while physically located outside the operating room (1 pathologist was in another state, 4 were at home, and 6 were elsewhere in the hospital). Interpretations of the still CLE images and video-flow CLE imaging were compared with the findings on the corresponding frozen and permanent H&E histology sections. RESULTS: Overall, 24 optical biopsies were acquired with mean ± SD 2 ± 1 optical biopsies per case. The mean duration of CLE system use was 1 ± 0.3 minutes/case and 0.25 ± 0.23 seconds/optical biopsy. The first image with identifiable histopathological features was acquired within 6 ± 0.1 seconds. Frozen sections were processed within 23 ± 2.8 minutes, which was significantly longer than CLE usage (p < 0.001). Video-flow CLE was used to correctly interpret tissue histoarchitecture in 96% of optical biopsies, which was substantially higher than the accuracy of using still CLE images (63%) (p = 0.005). CONCLUSIONS: When CLE is employed in tandem with a TSP, neurosurgeons and pathologists can view and interpret CLE images remotely and in real time without the need to biopsy tissue. A TSP allowed neurosurgeons to receive real-time feedback on the optically interrogated tissue microstructure, thereby improving cross-functional communication and intraoperative decision-making and resulting in significant workflow advantages over the use of frozen section analysis.

Introduction of In Vivo Confocal Laser Endomicroscopy and Real-Time Telepathology for Remote Intraoperative Neurosurgery-Pathology Consultation.

BACKGROUND: Precise communication between neurosurgeons and pathologists is crucial for optimizing patient care, especially for intraoperative diagnoses. Confocal laser endomicroscopy (CLE) combined with a telepathology software platform (TSP) provides a novel venue for neurosurgeons and pathologists to review CLE images and converse intraoperatively in real-time. OBJECTIVE: To describe the feasibility of integrating CLE and a TSP in the surgical workflow for real-time review of in vivo digital fluorescence tissue imaging in 3 patients with intracranial tumors. METHODS: Although the neurosurgeon used the CLE probe to generate fluorescence images of histoarchitecture within the operative field that were displayed on monitors in the operating room, the pathologist simultaneously remotely viewed the CLE images. The neurosurgeon and pathologist discussed in real-time the histological structures of intraoperative imaging locations. RESULTS: The neurosurgeon placed the CLE probe at various locations on and around the tumor, in the surgical resection bed, and on surrounding brain tissue with communication through the TSP. The neurosurgeon oriented the pathologist to the location of the CLE, and the pathologist and neurosurgeon discussed the CLE images in real-time. The TSP and CLE were integrated successfully and rapidly in the operating room in all 3 cases. No patient had perioperative complications. CONCLUSION: Two novel digital neurosurgical cellular imaging technologies were combined with intraoperative neurosurgeon-pathologist communication to guide the identification of abnormal histoarchitectural tissue features in real-time. CLE with the TSP may allow rapid decision-making during tumor resection that may hold significant advantages over the frozen section process and surgical workflow in general.

Information theory approaches to improve glioma diagnostic workflows in surgical neuropathology.

AIMS: Resource-strained healthcare ecosystems often struggle with the adoption of the World Health Organization (WHO) recommendations for the classification of central nervous system (CNS) tumors. The generation of robust clinical diagnostic aids and the advancement of simple solutions to inform investment strategies in surgical neuropathology would improve patient care in these settings. METHODS: We used simple information theory calculations on a brain cancer simulation model and real-world data sets to compare contributions of clinical, histologic, immunohistochemical, and molecular information. An image noise assay was generated to compare the efficiencies of different image segmentation methods in H&E and Olig2 stained images obtained from digital slides. An auto-adjustable image analysis workflow was generated and compared with neuropathologists for p53 positivity quantification. Finally, the density of extracted features of the nuclei, p53 positivity quantification, and combined ATRX/age feature was used to generate a predictive model for 1p/19q codeletion in IDH-mutant tumors. RESULTS: Information theory calculations can be performed on open access platforms and provide significant insight into linear and nonlinear associations between diagnostic biomarkers. Age, p53, and ATRX status have significant information for the diagnosis of IDH-mutant tumors. The predictive models may facilitate the reduction of false-positive 1p/19q codeletion by fluorescence in situ hybridization (FISH) testing. CONCLUSIONS: We posit that this approach provides an improvement on the cIMPACT-NOW workflow recommendations for IDH-mutant tumors and a framework for future resource and testing allocation.

Novel role for tumor suppressor gene, liver kinase B1, in epithelial-mesenchymal transition leading to chronic lung allograft dysfunction.

Epithelial-mesenchymal transition (EMT) has been implicated to play a role in chronic lung allograft dysfunction (CLAD). Liver kinase B1 (LKB1), a tumor suppressor gene, can regulate EMT. However, its role in CLAD development following lung transplantation remains unknown. Using qRT-PCR, biopsies from lung transplant recipients with bronchiolitis obliterans syndrome (BOS) demonstrated significant downregulation of LKB1 (p = .0001), compared to stable biopsies. To determine the role of LKB1 in EMT development, we analyzed EMT in human bronchial epithelial cell line BEAS-2B. Knockdown of LKB1 by siRNA significantly dysregulated mesenchymal markers expression in BEAS-2B cells. Following incubation of human primary bronchial epithelial cell or BEAS-2B cells with exosomes isolated from BOS or stable lung transplant recipients, LKB1 expression was inhibited when incubated with BOS-exosome. Incubation with BOS-exosomes also decreased LKB1 expression and induced EMT markers in air-liquid interface culture method. Our results provide novel evidence that exosomes released from transplanted lungs undergoing chronic rejection are associated with inactivated tumor suppressor gene LKB1 and this loss induces EMT leading to the pathogenesis of CLAD following human lung transplantation.

Intracranial mesenchymal tumors with FET-CREB fusion are composed of at least two epigenetic subgroups distinct from meningioma and extracranial sarcomas.

'Intracranial mesenchymal tumor, FET-CREB fusion-positive' occurs primarily in children and young adults and has previously been termed intracranial angiomatoid fibrous histiocytoma (AFH) or intracranial myxoid mesenchymal tumor (IMMT). Here we performed genome-wide DNA methylation array profiling of 20 primary intracranial mesenchymal tumors with FET-CREB fusion to further study their ontology. These tumors resolved into two distinct epigenetic subgroups that were both divergent from all other analyzed intracranial neoplasms and soft tissue sarcomas, including meningioma, clear cell sarcoma of soft tissue (CCS), and AFH of extracranial soft tissue. The first subgroup (Group A, 16 tumors) clustered nearest to but independent of solitary fibrous tumor and AFH of extracranial soft tissue, whereas the second epigenetic subgroup (Group B, 4 tumors) clustered nearest to but independent of CCS and also lacked expression of melanocytic markers (HMB45, Melan A, or MITF) characteristic of CCS. Group A tumors most often occurred in adolescence or early adulthood, arose throughout the neuroaxis, and contained mostly EWSR1-ATF1 and EWSR1-CREB1 fusions. Group B tumors arose most often in early childhood, were located along the cerebral convexities or spinal cord, and demonstrated an enrichment for tumors with CREM as the fusion partner (either EWSR1-CREM or FUS-CREM). Group A tumors more often demonstrated stellate/spindle cell morphology and hemangioma-like vasculature, whereas Group B tumors more often demonstrated round cell or epithelioid/rhabdoid morphology without hemangioma-like vasculature, although robust comparison of these clinical and histologic features requires future study. Patients with Group B tumors had inferior progression-free survival relative to Group A tumors (median 4.5 vs. 49 months, p = 0.001). Together, these findings confirm that intracranial AFH-like neoplasms and IMMT represent histologic variants of a single tumor type ('intracranial mesenchymal tumor, FET-CREB fusion-positive') that is distinct from meningioma and extracranial sarcomas. Additionally, epigenomic evaluation may provide important prognostic subtyping for this unique tumor entity.

High-Dose Fluorescein Reveals Unusual Confocal Endomicroscope Imaging of Low-Grade Glioma.

Background: Fluorescence-guided brain tumor surgery using fluorescein sodium (FNa) for contrast is effective in high-grade gliomas. However, the effectiveness of this technique for visualizing noncontrast-enhancing and low-grade gliomas is unknown. This report is the first documented case of the concurrent use of wide-field fluorescence-guided surgery and confocal laser endomicroscopy (CLE) with high-dose FNa (40 mg/kg) for intraoperative visualization of tumor tissue cellularity in a nonenhancing glioma. Case Description: A patient underwent fluorescence-guided surgery for a left frontal lobe mass without contrast enhancement on magnetic resonance imaging. The patient received 40 mg/kg FNa intravenously at the induction of anesthesia. Surgery was performed under visualization with a Yellow 560 filter and white-light wide-field imaging. Intraoperative CLE produced high-quality images of the lesion 1.5 h after FNa injection. Frozen-section analysis demonstrated findings comparable to those of intraoperative CLE visualization and consistent with World Health Organization (WHO) glioma grades II-III. The patient recovered without complications. Analysis of the permanent histologic sections identified the tumor as an anaplastic oligodendroglioma, IDH-mutant, 1p/19q co-deleted, consistent with WHO grade III because of discrete foci of hypercellularity and increased mitotic figures, but large regions of the lesion were low grade. Conclusions: The use of high-dose FNa in this patient with a nonenhancing borderline low-grade/high-grade glioma produced actionable wide-field fluorescence imaging using the operating microscope and improved CLE visualization of tumor cellularity. Higher doses of FNa for intraoperative CLE imaging and possible simultaneous wide-field fluorescence surgical guidance in nonenhancing gliomas merit further investigation.

Analysis of morphological characteristics of IDH-mutant/wildtype brain tumors using whole-lesion phenotype analysis.

BACKGROUND: Although IDH-mutant tumors aggregate to the frontotemporal regions, the clustering pattern of IDH-wildtype tumors is less clear. As voxel-based lesion-symptom mapping (VLSM) has several limitations for solid lesion mapping, a new technique, whole-lesion phenotype analysis (WLPA), is developed. We utilize WLPA to assess spatial clustering of tumors with IDH mutation from The Cancer Genome Atlas and The Cancer Imaging Archive. METHODS: The degree of tumor clustering segmented from T1 weighted images is measured to every other tumor by a function of lesion similarity to each other via the Hausdorff distance. Each tumor is ranked according to the degree to which its neighboring tumors show identical phenotypes, and through a permutation technique, significant tumors are determined. VLSM was applied through a previously described method. RESULTS: A total of 244 patients of mixed-grade gliomas (WHO II-IV) are analyzed, of which 150 were IDH-wildtype and 139 were glioblastomas. VLSM identifies frontal lobe regions that are more likely associated with the presence of IDH mutation but no regions where IDH-wildtype was more likely to be present. WLPA identifies both IDH-mutant and -wildtype tumors exhibit statistically significant spatial clustering. CONCLUSION: WLPA may provide additional statistical power when compared with VLSM without making several potentially erroneous assumptions. WLPA identifies tumors most likely to exhibit particular phenotypes, rather than producing anatomical maps, and may be used in conjunction with VLSM to understand the relationship between tumor morphology and biologically relevant tumor phenotypes.

Molecular Imaging of Glucose Metabolism for Intraoperative Fluorescence Guidance During Glioma Surgery.

PURPOSE: This study evaluated the use of molecular imaging of fluorescent glucose analog 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG) as a discriminatory marker for intraoperative tumor border identification in a murine glioma model. PROCEDURES: 2-NBDG was assessed in GL261 and U251 orthotopic tumor-bearing mice. Intraoperative fluorescence of topical and intravenous 2-NBDG in normal and tumor regions was assessed with an operating microscope, handheld confocal laser scanning endomicroscope (CLE), and benchtop confocal laser scanning microscope (LSM). Additionally, 2-NBDG fluorescence in tumors was compared with 5-aminolevulinic acid-induced protoporphyrin IX fluorescence. RESULTS: Intravenously administered 2-NBDG was detectable in brain tumor and absent in contralateral normal brain parenchyma on wide-field operating microscope imaging. Intraoperative and benchtop CLE showed preferential 2-NBDG accumulation in the cytoplasm of glioma cells (mean [SD] tumor-to-background ratio of 2.76 [0.43]). Topically administered 2-NBDG did not create sufficient tumor-background contrast for wide-field operating microscope imaging or under benchtop LSM (mean [SD] tumor-to-background ratio 1.42 [0.72]). However, topical 2-NBDG did create sufficient contrast to evaluate cellular tissue architecture and differentiate tumor cells from normal brain parenchyma. Protoporphyrin IX imaging resulted in a more specific delineation of gross tumor margins than intravenous or topical 2-NBDG and a significantly higher tumor-to-normal-brain fluorescence intensity ratio. CONCLUSION: After intravenous administration, 2-NBDG selectively accumulated in the experimental brain tumors and provided bright contrast under wide-field fluorescence imaging with a clinical-grade operating microscope. Topical 2-NBDG was able to create a sufficient contrast to differentiate tumor from normal brain cells on the basis of visualization of cellular architecture with CLE. 5-Aminolevulinic acid demonstrated superior specificity in outlining tumor margins and significantly higher tumor background contrast. Given the nontoxicity of 2-NBDG, its use as a topical molecular marker for noninvasive in vivo intraoperative microscopy is encouraging and warrants further clinical evaluation.

Uncertainty quantification in the radiogenomics modeling of EGFR amplification in glioblastoma.

Radiogenomics uses machine-learning (ML) to directly connect the morphologic and physiological appearance of tumors on clinical imaging with underlying genomic features. Despite extensive growth in the area of radiogenomics across many cancers, and its potential role in advancing clinical decision making, no published studies have directly addressed uncertainty in these model predictions. We developed a radiogenomics ML model to quantify uncertainty using transductive Gaussian Processes (GP) and a unique dataset of 95 image-localized biopsies with spatially matched MRI from 25 untreated Glioblastoma (GBM) patients. The model generated predictions for regional EGFR amplification status (a common and important target in GBM) to resolve the intratumoral genetic heterogeneity across each individual tumor-a key factor for future personalized therapeutic paradigms. The model used probability distributions for each sample prediction to quantify uncertainty, and used transductive learning to reduce the overall uncertainty. We compared predictive accuracy and uncertainty of the transductive learning GP model against a standard GP model using leave-one-patient-out cross validation. Additionally, we used a separate dataset containing 24 image-localized biopsies from 7 high-grade glioma patients to validate the model. Predictive uncertainty informed the likelihood of achieving an accurate sample prediction. When stratifying predictions based on uncertainty, we observed substantially higher performance in the group cohort (75% accuracy, n = 95) and amongst sample predictions with the lowest uncertainty (83% accuracy, n = 72) compared to predictions with higher uncertainty (48% accuracy, n = 23), due largely to data interpolation (rather than extrapolation). On the separate validation set, our model achieved 78% accuracy amongst the sample predictions with lowest uncertainty. We present a novel approach to quantify radiogenomics uncertainty to enhance model performance and clinical interpretability. This should help integrate more reliable radiogenomics models for improved medical decision-making.

Intracranial mesenchymal tumor with FET-CREB fusion-A unifying diagnosis for the spectrum of intracranial myxoid mesenchymal tumors and angiomatoid fibrous histiocytoma-like neoplasms.

Intracranial mesenchymal tumors with FET-CREB fusions are a recently described group of neoplasms in children and young adults characterized by fusion of a FET family gene (usually EWSR1, but rarely FUS) to a CREB family transcription factor (ATF1, CREB1, or CREM), and have been variously termed intracranial angiomatoid fibrous histiocytoma or intracranial myxoid mesenchymal tumor. The clinical outcomes, histologic features, and genomic landscape are not well defined. Here, we studied 20 patients with intracranial mesenchymal tumors proven to harbor FET-CREB fusion by next-generation sequencing (NGS). The 16 female and four male patients had a median age of 14 years (range 4-70). Tumors were uniformly extra-axial or intraventricular and located at the cerebral convexities (n = 7), falx (2), lateral ventricles (4), tentorium (2), cerebellopontine angle (4), and spinal cord (1). NGS demonstrated that eight tumors harbored EWSR1-ATF1 fusion, seven had EWSR1-CREB1, four had EWSR1-CREM, and one had FUS-CREM. Tumors were uniformly well circumscribed and typically contrast enhancing with solid and cystic growth. Tumors with EWSR1-CREB1 fusions more often featured stellate/spindle cell morphology, mucin-rich stroma, and hemangioma-like vasculature compared to tumors with EWSR1-ATF1 fusions that most often featured sheets of epithelioid cells with mucin-poor collagenous stroma. These tumors demonstrated polyphenotypic immunoprofiles with frequent positivity for desmin, EMA, CD99, MUC4, and synaptophysin, but absence of SSTR2A, myogenin, and HMB45 expression. There was a propensity for local recurrence with a median progression-free survival of 12 months and a median overall survival of greater than 60 months, with three patients succumbing to disease (all with EWSR1-ATF1 fusions). In combination with prior case series, this study provides further insight into intracranial mesenchymal tumors with FET-CREB fusion, which represent a distinct group of CNS tumors encompassing both intracranial myxoid mesenchymal tumor and angiomatoid fibrous histiocytoma-like neoplasms.