In Vivo PET Detection of Lung Micrometastasis in Mice by Targeting Endothelial VCAM-1 Using a Dual-Contrast PET/MRI Probe.

Current clinical diagnostic imaging methods for lung metastases are sensitive only to large tumours (1-2 mm cross-sectional diameter), and early detection can dramatically improve treatment. We have previously demonstrated that an antibody-targeted MRI contrast agent based on microparticles of iron oxide (MPIO; 1 µm diameter) enables the imaging of endothelial vascular cell adhesion molecule-1 (VCAM-1). Using a mouse model of lung metastasis, upregulation of endothelial VCAM-1 expression was demonstrated in micrometastasis-associated vessels but not in normal lung tissue, and binding of VCAM-MPIO to these vessels was evident histologically. Owing to the lack of proton MRI signals in the lungs, we modified the VCAM-MPIO to include zirconium-89 (89Zr, t1/2 = 78.4 h) in order to allow the in vivo detection of lung metastases by positron emission tomography (PET). Using this new agent (89Zr-DFO-VCAM-MPIO), it was possible to detect the presence of micrometastases within the lung in vivo from ca. 140 µm in diameter. Histological analysis combined with autoradiography confirmed the specific binding of the agent to the VCAM-1 expressing vasculature at the sites of pulmonary micrometastases. By retaining the original VCAM-MPIO as the basis for this new molecular contrast agent, we have created a dual-modality (PET/MRI) agent for the concurrent detection of lung and brain micrometastases.

Targeted opening of the blood-brain barrier using VCAM-1 functionalised microbubbles and "whole brain" ultrasound.

Metastatic tumours in the brain now represent one of the leading causes of death from cancer. Current treatments are largely ineffective owing to the combination of late diagnosis and poor delivery of therapies across the blood-brain barrier (BBB). Conjugating magnetic resonance imaging (MRI) contrast agents with a monoclonal antibody for VCAM-1 (anti-VCAM1) has been shown to enable detection of micrometastases, two to three orders of magnitude smaller in volume than those currently detectable clinically. The aim of this study was to exploit this targeting approach to enable localised and temporary BBB opening at the site of early-stage metastases using functionalised microbubbles and ultrasound. Methods: Microbubbles functionalised with anti-VCAM1 were synthesised and shown to bind to VCAM-1-expressing cells in vitro. Experiments were then conducted in vivo in a unilateral breast cancer brain metastasis mouse model using Gadolinium-DTPA (Gd-DTPA) enhanced MRI to detect BBB opening. Following injection of Gd-DTPA and targeted microbubbles, the whole brain volume was simultaneously exposed to ultrasound (0.5 MHz, 10% duty cycle, 0.7 MPa peak negative pressure, 2 min treatment time). T1-weighted MRI was then performed to identify BBB opening, followed by histological confirmation via immunoglobulin G (IgG) immunohistochemistry. Results: In mice treated with targeted microbubbles and ultrasound, statistically significantly greater extravasation of Gd-DTPA and IgG was observed in the left tumour-bearing hemisphere compared to the right hemisphere 5 min after treatment. No acute adverse effects were observed. There was no investigation of longer term bioeffects owing to the nature of the study. Conclusion: The results demonstrate the feasibility of using targeted microbubbles in combination with low intensity ultrasound to localise opening of the BBB to metastatic sites in the brain. This approach has potential application in the treatment of metastatic tumours whose location cannot be established a priori with conventional imaging methods.

Inflammatory bowel disease induces pathological α-synuclein aggregation in the human gut and brain.

AIMS: According to Braak's hypothesis, it is plausible that Parkinson's disease (PD) originates in the enteric nervous system (ENS) and spreads to the brain through the vagus nerve. In this work, we studied whether inflammatory bowel diseases (IBDs) in humans can progress with the emergence of pathogenic α-synuclein (α-syn) in the gastrointestinal tract and midbrain dopaminergic neurons. METHODS: We have analysed the gut and the ventral midbrain from subjects previously diagnosed with IBD and form a DSS-based rat model of gut inflammation in terms of α-syn pathology. RESULTS: Our data support the existence of pathogenic α-syn in both the gut and the brain, thus reinforcing the potential role of the ENS as a contributing factor in PD aetiology. Additionally, we have analysed the effect of a DSS-based rat model of gut inflammation to demonstrate (i) the appearance of P-α-syn inclusions in both Auerbach's and Meissner's plexuses (gut), (ii) an increase in α-syn expression in the ventral mesencephalon (brain) and (iii) the degeneration of nigral dopaminergic neurons, which all are considered classical hallmarks in PD. CONCLUSION: These results strongly support the plausibility of Braak's hypothesis and emphasise the significance of peripheral inflammation and the gut-brain axis in initiating α-syn aggregation and transport to the substantia nigra, resulting in neurodegeneration.

An open resource combining multi-contrast MRI and microscopy in the macaque brain.

Understanding brain structure and function often requires combining data across different modalities and scales to link microscale cellular structures to macroscale features of whole brain organisation. Here we introduce the BigMac dataset, a resource combining in vivo MRI, extensive postmortem MRI and multi-contrast microscopy for multimodal characterisation of a single whole macaque brain. The data spans modalities (MRI and microscopy), tissue states (in vivo and postmortem), and four orders of spatial magnitude, from microscopy images with micrometre or sub-micrometre resolution, to MRI signals on the order of millimetres. Crucially, the MRI and microscopy images are carefully co-registered together to facilitate quantitative multimodal analyses. Here we detail the acquisition, curation, and first release of the data, that together make BigMac a unique, openly-disseminated resource available to researchers worldwide. Further, we demonstrate example analyses and opportunities afforded by the data, including improvement of connectivity estimates from ultra-high angular resolution diffusion MRI, neuroanatomical insight provided by polarised light imaging and myelin-stained histology, and the joint analysis of MRI and microscopy data for reconstruction of the microscopy-inspired connectome. All data and code are made openly available.

Imaging angiogenesis in an intracerebrally induced model of brain macrometastasis using αv ß3 -targeted iron oxide microparticles.

Brain metastasis is responsible for a large proportion of cancer mortality, and there are currently no effective treatments. Moreover, the impact of treatments, particularly antiangiogenic therapeutics, is difficult to ascertain using current magnetic resonance imaging (MRI) methods. Imaging of the angiogenic vasculature has been successfully carried out in solid tumours using microparticles of iron oxide (MPIO) conjugated to a Arg-Gly-Asp peptide (RGD) targeting integrin αv ß3 . The aim of this study was to determine whether RGD-MPIO could be used to identify angiogenic blood vessels in brain metastases in vivo. A mouse model of intracerebrally implanted brain macrometastasis was established through intracerebral injection of 4T1-GFP cells. T2 *-weighted imaging was used to visualise MPIO-induced hypointense voxels in vivo, and Prussian blue staining was used to visualise MPIO and endogenous iron histologically ex vivo. The RGD-MPIO showed target-specific binding in vivo, but the sensitivity of the agent for visualising angiogenic vessels per se was reduced by the presence of endogenous iron-laden macrophages in larger metastases, resulting in pre-existing hypointense areas within the tumour. Further, our data suggest that peptide-targeted MPIO, but not antibody-targeted MPIO, are taken up by perivascular macrophages within the macrometastatic microenvironment, resulting in additional nonspecific contrast. While pre-MPIO imaging will circumvent the issues surrounding pre-existing hypointensities and enable detection of specific contrast, our preliminary findings suggest that the use of antibodies rather than peptides as the targeting ligand may represent a preferable route forward for new angiogenesis-targeted molecular MRI agents.

Inhibition of Anti-Inflammatory Macrophage Phenotype Reduces Tumour Growth in Mouse Models of Brain Metastasis.

Breast cancer brain metastasis is a significant clinical problem and carries a poor prognosis. Although it is well-established that macrophages are a primary component of the brain metastasis microenvironment, the role of blood-derived macrophages (BDM) and brain-resident microglia in the progression of brain metastases remains uncertain. The aim of this study, therefore, was to determine the role, specifically, of pro- and anti-inflammatory BDM and microglial phenotypes on metastasis progression. Initial in vitro studies demonstrated decreased migration of EO771 metastatic breast cancer cells in the presence of pro-inflammatory, but not anti-inflammatory, stimulated RAW 264.7 macrophages. In vivo, suppression of the anti-inflammatory BDM phenotype, specifically, via myeloid knock out of Krüppel-like Factor 4 (KLF4) significantly reduced EO771 tumour growth in the brains of C57BL/6 mice. Further, pharmacological inhibition of the anti-inflammatory BDM and/or microglial phenotypes, via either Colony Stimulating Factor 1 Receptor (CSF-1R) or STAT6 pathways, significantly decreased tumour burden in two different syngeneic mouse models of breast cancer brain metastasis. These findings suggest that switching BDM and microglia towards a more pro-inflammatory phenotype may be an effective therapeutic strategy in brain metastasis.

Quantitative chemical exchange saturation transfer imaging of nuclear overhauser effects in acute ischemic stroke.

PURPOSE: In chemical exchange saturation transfer imaging, saturation effects between - 2 to - 5 ppm (nuclear Overhauser effects, NOEs) have been shown to exhibit contrast in preclinical stroke models. Our previous work on NOEs in human stroke used an analysis model that combined NOEs and semisolid MT; however their combination might feasibly have reduced sensitivity to changes in NOEs. The aim of this study was to explore the information a 4-pool Bloch-McConnell model provides about the NOE contribution in ischemic stroke, contrasting that with an intentionally approximate 3-pool model. METHODS: MRI data from 12 patients presenting with ischemic stroke were retrospectively analyzed, as well as from six animals induced with an ischemic lesion. Two Bloch-McConnell models (4 pools, and a 3-pool approximation) were compared for their ability to distinguish pathological tissue in acute stroke. The association of NOEs with pH was also explored, using pH phantoms that mimic the intracellular environment of naïve mouse brain. RESULTS: The 4-pool measure of NOEs exhibited a different association with tissue outcome compared to 3-pool approximation in the ischemic core and in tissue that underwent delayed infarction. In the ischemic core, the 4-pool measure was elevated in patient white matter ( 1.20±0.20 ) and in animals ( 1.27±0.20 ). In the naïve brain pH phantoms, significant positive correlation between the NOE and pH was observed. CONCLUSION: Associations of NOEs with tissue pathology were found using the 4-pool metric that were not observed using the 3-pool approximation. The 4-pool model more adequately captured in vivo changes in NOEs and revealed trends depending on tissue pathology in stroke.

VCAM-1-targeted MRI Improves Detection of the Tumor-brain Interface.

PURPOSE: Despite optimal local therapy, tumor cell invasion into normal brain parenchyma frequently results in recurrence in patients with solid tumors. The aim of this study was to determine whether microvascular inflammation can be targeted to better delineate the tumor-brain interface through vascular cell adhesion molecule-1 (VCAM-1)-targeted MRI. EXPERIMENTAL DESIGN: Intracerebral xenograft rat models of MDA231Br-GFP (breast cancer) brain metastasis and U87MG (glioblastoma) were used to histologically examine the tumor-brain interface and to test the efficacy of VCAM-1-targeted MRI in detecting this region. Human biopsy samples of the brain metastasis and glioblastoma margins were examined for endothelial VCAM-1 expression. RESULTS: The interface between tumor and surrounding normal brain tissue exhibited elevated endothelial VCAM-1 expression and increased microvessel density. Tumor proliferation and stemness markers were also significantly upregulated at the tumor rim in the brain metastasis model. T2*-weighted MRI, following intravenous administration of VCAM-MPIO, highlighted the tumor-brain interface of both tumor models more extensively than gadolinium-DTPA-enhanced T1-weighted MRI. Sites of VCAM-MPIO binding, evident as hypointense signals on MR images, correlated spatially with endothelial VCAM-1 upregulation and bound VCAM-MPIO beads detected histologically. These findings were further validated in an orthotopic medulloblastoma model. Finally, the tumor-brain interface in human brain metastasis and glioblastoma samples was similarly characterized by microvascular inflammation, extending beyond the region detectable using conventional MRI. CONCLUSIONS: This work illustrates the potential of VCAM-1-targeted MRI for improved delineation of the tumor-brain interface in both primary and secondary brain tumors.

Metabolomic Biomarkers in Blood Samples Identify Cancers in a Mixed Population of Patients with Nonspecific Symptoms.

PURPOSE: Early diagnosis of cancer is critical for improving patient outcomes, but cancers may be hard to diagnose if patients present with nonspecific signs and symptoms. We have previously shown that nuclear magnetic resonance (NMR) metabolomics analysis can detect cancer in animal models and distinguish between differing metastatic disease burdens. Here, we hypothesized that biomarkers within the blood metabolome could identify cancers within a mixed population of patients referred from primary care with nonspecific symptoms, the so-called "low-risk, but not no-risk" patient group, as well as distinguishing between those with and without metastatic disease. EXPERIMENTAL DESIGN: Patients (n = 304 comprising modeling, n = 192, and test, n = 92) were recruited from 2017 to 2018 from the Oxfordshire Suspected CANcer (SCAN) pathway, a multidisciplinary diagnostic center (MDC) referral pathway for patients with nonspecific signs and symptoms. Blood was collected and analyzed by NMR metabolomics. Orthogonal partial least squares discriminatory analysis (OPLS-DA) models separated patients, based upon diagnoses received from the MDC assessment, within 62 days of initial appointment. RESULTS: Area under the ROC curve for identifying patients with solid tumors in the independent test set was 0.83 [95% confidence interval (CI): 0.72-0.95]. Maximum sensitivity and specificity were 94% (95% CI: 73-99) and 82% (95% CI: 75-87), respectively. We could also identify patients with metastatic disease in the cohort of patients with cancer with sensitivity and specificity of 94% (95% CI: 72-99) and 88% (95% CI: 53-98), respectively. CONCLUSIONS: For a mixed group of patients referred from primary care with nonspecific signs and symptoms, NMR-based metabolomics can assist their diagnosis, and may differentiate both those with malignancies and those with and without metastatic disease. See related commentary by Van Tine and Lyssiotis, p. 1477.

Selective blood-brain barrier permeabilization of brain metastases by a type 1 receptor-selective tumor necrosis factor mutein.

BACKGROUND: Metastasis to the brain is a major challenge with poor prognosis. The blood-brain barrier (BBB) is a significant impediment to effective treatment, being intact during the early stages of tumor development and heterogeneously permeable at later stages. Intravenous injection of tumor necrosis factor (TNF) selectively induces BBB permeabilization at sites of brain micrometastasis, in a TNF type 1 receptor (TNFR1)-dependent manner. Here, to enable clinical translation, we have developed a TNFR1-selective agonist variant of human TNF that induces BBB permeabilization, while minimizing potential toxicity. METHODS: A library of human TNF muteins (mutTNF) was generated and assessed for binding specificity to mouse and human TNFR1/2, endothelial permeabilizing activity in vitro, potential immunogenicity, and circulatory half-life. The permeabilizing ability of the most promising variant was assessed in vivo in a model of brain metastasis. RESULTS: The primary mutTNF variant showed similar affinity for human TNFR1 than wild-type human TNF, similar affinity for mouse TNFR1 as wild-type mouse TNF, undetectable binding to human/mouse TNFR2, low potential immunogenicity, and permeabilization of an endothelial monolayer. Circulatory half-life was similar to mouse/human TNF and BBB permeabilization was induced selectively at sites of micrometastases in vivo, with a time window of ≥24 hours and enabling delivery of agents within a therapeutically relevant range (0.5-150 kDa), including the clinically approved therapy, trastuzumab. CONCLUSIONS: We have developed a clinically translatable mutTNF that selectively opens the BBB at micrometastatic sites, while leaving the rest of the cerebrovasculature intact. This approach will open a window for brain metastasis treatment that currently does not exist.

Radioimmunotherapy for Brain Metastases: The Potential for Inflammation as a Target of Choice.

Brain metastases (BM) are frequently detected during the follow-up of patients with malignant tumors, particularly in those with advanced disease. Despite a major progress in systemic anti-cancer treatments, the average overall survival of these patients remains limited (6 months from diagnosis). Also, cognitive decline is regularly reported especially in patients treated with whole brain external beam radiotherapy (WBRT), due to the absorbed radiation dose in healthy brain tissue. New targeted therapies, for an earlier and/or more specific treatment of the tumor and its microenvironment, are needed. Radioimmunotherapy (RIT), a combination of a radionuclide to a specific antibody, appears to be a promising tool. Inflammation, which is involved in multiple steps, including the early phase, of BM development is attractive as a relevant target for RIT. This review will focus on the (1) early biomarkers of inflammation in BM pertinent for RIT, (2) state of the art studies on RIT for BM, and (3) the importance of dosimetry to RIT in BM. These two last points will be addressed in comparison to the conventional EBRT treatment, particularly with respect to the balance between tumor control and healthy tissue complications. Finally, because new diagnostic imaging techniques show a potential for the detection of BM at an early stage of the disease, we focus particularly on this therapeutic window.

Multimodal evaluation of hypoxia in brain metastases of lung cancer and interest of hypoxia image-guided radiotherapy.

Lung cancer patients frequently develop brain metastases (BM). Despite aggressive treatment including neurosurgery and external-radiotherapy, overall survival remains poor. There is a pressing need to further characterize factors in the microenvironment of BM that may confer resistance to radiotherapy (RT), such as hypoxia. Here, hypoxia was first evaluated in 28 biopsies from patients with non­small cell lung cancer (NSCLC) BM, using CA-IX immunostaining. Hypoxia characterization (pimonidazole, CA-IX and HIF-1α) was also performed in different preclinical NSCLC BM models induced either by intracerebral injection of tumor cells (H2030-Br3M, H1915) into the cortex and striatum, or intracardial injection of tumor cells (H2030-Br3M). Additionally, [18F]-FMISO-PET and oxygen-saturation-mapping-MRI (SatO2-MRI) were carried out in the intracerebral BM models to further characterize tumor hypoxia and evaluate the potential of Hypoxia-image-guided-RT (HIGRT). The effect of RT on proliferation of BM ([18F]-FLT-PET), tumor volume and overall survival was determined. We showed that hypoxia is a major yet heterogeneous feature of BM from lung cancer both preclinically and clinically. HIGRT, based on hypoxia heterogeneity observed between cortical and striatal metastases in the intracerebrally induced models, showed significant potential for tumor control and animal survival. These results collectively highlight hypoxia as a hallmark of BM from lung cancer and the value of HIGRT in better controlling tumor growth.

Study of common quantification methods of amide proton transfer magnetic resonance imaging for ischemic stroke detection.

PURPOSE: To assess the correlation and differences between common amide proton transfer (APT) quantification methods in the diagnosis of ischemic stroke. METHODS: Five APT quantification methods, including asymmetry analysis and its variants as well as two Lorentzian model-based methods, were applied to data acquired from six rats that underwent middle cerebral artery occlusion scanned at 9.4T. Diffusion and perfusion-weighted images, and water relaxation time maps were also acquired to study the relationship of these conventional imaging modalities with the different APT quantification methods. RESULTS: The APT ischemic area estimates had varying sizes (Jaccard index: 0.544 ≤ J ≤ 0.971) and had varying correlations in their distributions (Pearson correlation coefficient: 0.104 ≤ r ≤ 0.995), revealing discrepancies in the quantified ischemic areas. The Lorentzian methods produced the highest contrast-to-noise ratios (CNRs; 1.427 ≤ CNR ≤ 2.002), but generated APT ischemic areas that were comparable in size to the cerebral blood flow (CBF) deficit areas; asymmetry analysis and its variants produced APT ischemic areas that were smaller than the CBF deficit areas but larger than the apparent diffusion coefficient deficit areas, though having lower CNRs (0.561 ≤ CNR ≤ 1.083). CONCLUSION: There is a need to further investigate the accuracy and correlation of each quantification method with the pathophysiology using a larger scale multi-imaging modality and multi-time-point clinical study. Future studies should include the magnetization transfer ratio asymmetry results alongside the findings of the study to facilitate the comparison of results between different centers and also the published literature.

A novel molecular magnetic resonance imaging agent targeting activated leukocyte cell adhesion molecule as demonstrated in mouse brain metastasis models.

Molecular magnetic resonance imaging (MRI) allows visualization of biological processes at the molecular level. Upregulation of endothelial ALCAM (activated leukocyte cell adhesion molecule) is a key element for leukocyte recruitment in neurological disease. The aim of this study, therefore, was to develop a novel molecular MRI contrast agent, by conjugating anti-ALCAM antibodies to microparticles of iron oxide (MPIO), for detection of endothelial ALCAM expression in vivo. Binding specificity of ALCAM-MPIO was demonstrated in vitro under static and flow conditions. Subsequently, in a proof-of-concept study, mouse models of brain metastasis were induced by intracardial injection of brain-tropic human breast carcinoma, lung adenocarcinoma or melanoma cells to upregulate endothelial ALCAM. At selected time-points, mice were injected intravenously with ALCAM-MPIO, and ALCAM-MPIO induced hypointensities were observed on T2*-weighted images in all three models. Post-gadolinium MRI confirmed an intact blood-brain barrier, indicating endoluminal binding. Correlation between endothelial ALCAM expression and ALCAM-MPIO binding was confirmed histologically. Statistical analysis indicated high sensitivity (80-90%) and specificity (79-83%) for detection of endothelial ALCAM in vivo with ALCAM-MPIO. Given reports of endothelial ALCAM upregulation in numerous neurological diseases, this advance in our ability to image ALCAM in vivo may yield substantial improvements for both diagnosis and targeted therapy.

Microbubbles Containing Lysolipid Enhance Ultrasound-Mediated Blood-Brain Barrier Breakdown In Vivo.

Ultrasound and microbubbles (MBs) offer a noninvasive method of temporarily enhancing blood-brain barrier (BBB) permeability to therapeutics. To reduce off-target effects, it is desirable to minimize the ultrasound pressures required. It has been shown that a new formulation of MBs containing lysolipids (Lyso-MBs) can increase the cellular uptake of a model drug in vitro. The aim of this study is to investigate whether Lyso-MBs can also enhance BBB permeability in vivo. Female BALB/c mice are injected with either Lyso-MBs or control MBs and gadolinium-DTPA (Gd-DTPA) and exposed to ultrasound (500 kHz, 1 Hz pulse repetition frequency, 1 ms pulse length, peak-negative pressures 160-480 kPa) for 2 min. BBB permeabilization is measured via magnetic resonance imaging (7.0 T) of Gd-DTPA extravasation and subsequent histological examination of brain tissue to assess serum immunoglobulin G (IgG) extravasation (n = 8 per group). An approximately twofold enhancement in BBB permeability is produced by the Lyso-MBs at the highest ultrasound pressure compared with the control. These findings indicate that modifying the composition of phospholipid-shelled MBs has the potential to improve the efficiency of BBB opening, without increasing the ultrasound pressure amplitude required. This is particularly relevant for delivery of therapeutics deep within the brain.

STAT3-Mediated Astrocyte Reactivity Associated with Brain Metastasis Contributes to Neurovascular Dysfunction.

Astrocytes are thought to play a pivotal role in coupling neural activity and cerebral blood flow. However, it has been shown that astrocytes undergo morphologic changes in response to brain metastasis, switching to a reactive phenotype, which has the potential to significantly compromise cerebrovascular function and contribute to the neurological sequelae associated with brain metastasis. Given that STAT3 is a key regulator of astrocyte reactivity, we aimed here to determine the impact of STAT3-mediated astrocyte reactivity on neurovascular function in brain metastasis. Rat models of brain metastasis and ciliary neurotrophic factor were used to induce astrocyte reactivity. Multimodal imaging, electrophysiology, and IHC were performed to determine the relationship between reactive astrocytes and changes in the cerebrovascular response to electrical and physiological stimuli. Subsequently, the STAT3 pathway in astrocytes was inhibited with WP1066 to determine the role of STAT3-mediated astrocyte reactivity, specifically, in brain metastasis. Astrocyte reactivity associated with brain metastases impaired cerebrovascular responses to stimuli at both the cellular and functional level and disrupted astrocyte-endothelial interactions in both animal models and human brain metastasis samples. Inhibition of STAT3-mediated astrocyte reactivity in rats with brain metastases restored cerebrovascular function, as shown by in vivo imaging, and limited cerebrovascular changes associated with tumor growth. Together these findings suggest that inhibiting STAT3-mediated astrocyte reactivity may confer significant improvements in neurological outcome for patients with brain metastases and could potentially be tested in other brain tumors. SIGNIFICANCE: These findings demonstrate that selectively targeting STAT3-mediated astrocyte reactivity ameliorates the cerebrovascular dysfunction associated with brain metastasis, providing a potential therapeutic avenue for improved patient outcome.

Brain Metastasis Cell Lines Panel: A Public Resource of Organotropic Cell Lines.

Spread of cancer to the brain remains an unmet clinical need in spite of the increasing number of cases among patients with lung, breast cancer, and melanoma most notably. Although research on brain metastasis was considered a minor aspect in the past due to its untreatable nature and invariable lethality, nowadays, limited but encouraging examples have questioned this statement, making it more attractive for basic and clinical researchers. Evidences of its own biological identity (i.e., specific microenvironment) and particular therapeutic requirements (i.e., presence of blood-brain barrier, blood-tumor barrier, molecular differences with the primary tumor) are thought to be critical aspects that must be functionally exploited using preclinical models. We present the coordinated effort of 19 laboratories to compile comprehensive information related to brain metastasis experimental models. Each laboratory has provided details on the cancer cell lines they have generated or characterized as being capable of forming metastatic colonies in the brain, as well as principle methodologies of brain metastasis research. The Brain Metastasis Cell Lines Panel (BrMPanel) represents the first of its class and includes information about the cell line, how tropism to the brain was established, and the behavior of each model in vivo. These and other aspects described are intended to assist investigators in choosing the most suitable cell line for research on brain metastasis. The main goal of this effort is to facilitate research on this unmet clinical need, to improve models through a collaborative environment, and to promote the exchange of information on these valuable resources.

White matter structure and myelin-related gene expression alterations with experience in adult rats.

White matter (WM) plasticity during adulthood is a recently described phenomenon by which experience can shape brain structure. It has been observed in humans using diffusion tensor imaging (DTI) and myelination has been suggested as a possible mechanism. Here, we set out to identify molecular and cellular changes associated with WM plasticity measured by DTI. We combined DTI, immunohistochemistry and mRNA expression analysis and examined the effects of somatosensory experience in adult rats. First, we observed experience-induced DTI differences in WM and in grey matter structure. C-Fos mRNA expression, a marker of cortical activity, in the barrel cortex correlated with the MRI WM metrics, indicating that molecular correlates of cortical activity relate to macroscale measures of WM structure. Analysis of myelin-related genes revealed higher myelin basic protein (MBP) mRNA expression. Higher MBP protein expression was also found via immunohistochemistry in WM. Finally, unbiased RNA sequencing analysis identified 134 differentially expressed genes encoding proteins involved in functions related to cell proliferation and differentiation, regulation of myelination and neuronal activity modulation. In conclusion, macroscale measures of WM plasticity are supported by both molecular and cellular evidence and confirm that myelination is one of the underlying mechanisms.

Improving Delineation of True Tumor Volume With Multimodal MRI in a Rat Model of Brain Metastasis.

PURPOSE: Brain metastases are almost universally lethal with short median survival times. Despite this, they are often potentially curable, with therapy failing only because of local relapse. One key reason relapse occurs is because treatment planning did not delineate metastasis margins sufficiently or accurately, allowing residual tumor to regrow. The aim of this study was to determine the extent to which multimodal magnetic resonance imaging (MRI), with a simple and automated analysis pipeline, could improve upon current clinical practice of single-modality, independent-observer tumor delineation. METHODS AND MATERIALS: We used a single rat model of brain metastasis (ENU1564 breast carcinoma cells in BD-IX rats), with and without radiation therapy. Multimodal MRI data were acquired using sequences either in current clinical use or in clinical trial and included postgadolinium T1-weighted images and maps of blood flow, blood volume, T1 and T2 relaxation times, and apparent diffusion coefficient. RESULTS: In all cases, independent observers underestimated the true size of metastases from single-modality gadolinium-enhanced MRI (85 ± 36 µL vs 131 ± 40 µL histologic measurement), although multimodal MRI more accurately delineated tumor volume (132 ± 41 µL). Multimodal MRI offered increased sensitivity compared with independent observer for detecting metastasis (0.82 vs 0.61, respectively), with only a slight decrease in specificity (0.86 vs 0.98). Blood flow maps conferred the greatest improvements in margin detection for late-stage metastases after radiation therapy. Gadolinium-enhanced T1-weighted images conferred the greatest increase in accuracy of detection for smaller metastases. CONCLUSIONS: These findings suggest that multimodal MRI of brain metastases could significantly improve the visualization of brain metastasis margins, beyond current clinical practice, with the potential to decrease relapse rates and increase patient survival. This finding now needs validation in additional tumor models or clinical cohorts.