Facilitating Repeat Intracarotid Injections in Mouse Models by a Novel Injection Site Repair Technique.

Given recent advances in the delivery of novel antitumor therapeutics using endovascular selective intraarterial delivery methods in neuro-oncology, there is an urgent need to develop methods for intracarotid injections in mouse models, including methods to repair the carotid artery in mice after injection to allow for subsequent injections. We developed a method of intracarotid injection in a mouse model to deliver therapeutics into the internal carotid artery (ICA) with two alternative procedures. During injection, the needle is inserted into the common carotid artery (CCA) after tying a suture around the external carotid artery (ECA) and injected therapeutics are delivered into the ICA. Following injection, the common carotid artery (CCA) can be ligated, which limits the number of intracarotid injections to one. The alternative procedure described in this article includes a modification where intracarotid artery injection is followed by injection site repair of the CCA, which restores blood flow within the CCA and avoids the complication of cerebral ischemia seen in some mouse models. We also compared the delivery of bone marrow-derived human mesenchymal stem cells (BM-hMSCs) to intracranial tumors when delivered through intracarotid injection with and without injection site repair following the injection. Delivery of BM-hMSCs does not differ significantly between the methods. Our results demonstrate that injection site repair of the CCA allows for repeat injections through the same artery and does not impair the delivery and distribution of injected material, thus providing a model with greater flexibility that more closely emulates intracarotid injection in humans.

The surgical intervention for pineal region tumors.

Histological and molecular characterization is essential for the diagnosis of pediatric brain tumors. In the pineal region tumors, it is necessary to remove a sufficient tumor volume to make a diagnosis. However, surgery in this region is challenging due to its deep anatomical location and surrounded by critical structures and complex venous system. Knowledge of the anatomy and function of the pineal region and tumor histological types is imperative for the successful management of pineal region tumors. This article describes surgical approaches to pineal tumors, focusing on the occipital transtentorial approach and adding the author's experience to what has been known in the literature. Recent innovations have made this approach more popular and can be applied to occipital fossa lesions.

Utility of real-time polymerase chain reaction for the assessment of CDKN2A homozygous deletion in adult-type IDH-mutant astrocytoma.

The World Health Organization Classification of Tumors of the Central Nervous System 5th Edition (WHO CNS5) introduced a newly defined astrocytoma, IDH-mutant grade 4, for adult diffuse glioma classification. One of the diagnostic criteria is the presence of a CDKN2A/B homozygous deletion (HD). Here, we report a robust and cost-effective quantitative polymerase chain reaction (qPCR)-based test for assessing CDKN2A HD. A TaqMan copy number assay was performed using a probe located within CDKN2A. The linear correlation between the Ct values and relative CDKN2A copy number was confirmed using a serial mixture of DNA from normal blood and U87MG cells. The qPCR assay was performed in 109 IDH-mutant astrocytomas, including 14 tumors with CDKN2A HD, verified either by multiplex ligation-dependent probe amplification (MLPA) or CytoScan HD microarray platforms. Receiver operating characteristic curve analysis indicated that a cutoff value of 0.85 yielded optimal sensitivity (100%) and specificity (99.0%) for determining CDKN2A HD. The assay applies to DNA extracted from frozen or formalin-fixed paraffin-embedded tissue samples. Survival was significantly shorter in patients with than in those without CDKN2A HD, assessed by either MLPA/CytoScan or qPCR. Thus, our qPCR method is clinically applicable for astrocytoma grading and prognostication, compatible with the WHO CNS5.

KLC1-ROS1 Fusion Exerts Oncogenic Properties of Glioma Cells via Specific Activation of JAK-STAT Pathway.

Here, we investigated the detailed molecular oncogenic mechanisms of a novel receptor tyrosine kinase (RTK) fusion, KLC1-ROS1, with an adapter molecule, KLC1, and an RTK, ROS1, discovered in pediatric glioma, and we explored a novel therapeutic target for glioma that possesses oncogenic RTK fusion. When wild-type ROS1 and KLC1-ROS1 fusions were stably expressed in the human glioma cell lines A172 and U343MG, immunoblotting revealed that KLC1-ROS1 fusion specifically activated the JAK2-STAT3 pathway, a major RTK downstream signaling pathway, when compared with wild-type ROS1. Immunoprecipitation of the fractionated cell lysates revealed a more abundant association of the KLC1-ROS1 fusion with JAK2 than that observed for wild-type ROS1 in the cytosolic fraction. A mutagenesis study of the KLC1-ROS1 fusion protein demonstrated the fundamental roles of both the KLC1 and ROS1 domains in the constitutive activation of KLC1-ROS1 fusion. Additionally, in vitro assays demonstrated that KLC1-ROS1 fusion upregulated cell proliferation, invasion, and chemoresistance when compared to wild-type ROS1. Combination treatment with the chemotherapeutic agent temozolomide and an inhibitor of ROS1, JAK2, or a downstream target of STAT3, demonstrated antitumor effects against KLC1-ROS1 fusion-expressing glioma cells. Our results demonstrate that KLC1-ROS1 fusion exerts oncogenic activity through serum-independent constitutive activation, resulting in specific activation of the JAK-STAT pathway. Our data suggested that molecules other than RTKs may serve as novel therapeutic targets for RTK fusion in gliomas.

Characterization of patient-derived bone marrow human mesenchymal stem cells as oncolytic virus carriers for the treatment of glioblastoma.

OBJECTIVE: Delta-24-RGD is an oncolytic adenovirus that is capable of replicating in and killing human glioma cells. Although intratumoral delivery of Delta-24-RGD can be effective, systemic delivery would improve its clinical application. Bone marrow-derived human mesenchymal stem cells (BM-hMSCs) obtained from healthy donors have been investigated as virus carriers. However, it is unclear whether BM-hMSCs can be derived from glioma patients previously treated with marrow-toxic chemotherapy or whether such BM-hMSCs can deliver oncolytic viruses effectively. Herein, the authors undertook a prospective clinical trial to determine the feasibility of obtaining BM-hMSCs from patients with recurrent malignant glioma who were previously exposed to marrow-toxic chemotherapy. METHODS: The authors enrolled 5 consecutive patients who had been treated with radiation therapy and chemotherapy. BM aspirates were obtained from the iliac crest and were cultured to obtain BM-hMSCs. RESULTS: The patient-derived BM-hMSCs (PD-BM-hMSCs) had a morphology similar to that of healthy donor-derived BM-hMSCs (HD-BM-hMSCs). Flow cytometry revealed that all 5 cell lines expressed canonical MSC surface markers. Importantly, these cultures could be made to differentiate into osteocytes, adipocytes, and chondrocytes. In all cases, the PD-BM-hMSCs homed to intracranial glioma xenografts in mice after intracarotid delivery as effectively as HD-BM-hMSCs. The PD-BM-hMSCs loaded with Delta-24-RGD (PD-BM-MSC-D24) effectively eradicated human gliomas in vitro. In in vivo studies, intravascular administration of PD-BM-MSC-D24 increased the survival of mice harboring U87MG gliomas. CONCLUSIONS: The authors conclude that BM-hMSCs can be acquired from patients previously treated with marrow-toxic chemotherapy and that these PD-BM-hMSCs are effective carriers for oncolytic viruses.

Case of penetrating brain injury caused by a ventriculoperitoneal shunting procedure.

Brain injury with ventricle puncture is a well-known complication of ventriculoperitoneal (VP) shunting. However, parenchymal injuries caused by a shunt tunneller are rare. Herein, we present a case of penetrating brain injury caused by a shunt tunneller during VP shunting. An 83-year-old woman with brainstem glioma underwent VP shunting to control hydrocephalus due to tumour growth. She underwent brainstem tumour biopsy with a lateral suboccipital approach. After the shunting, CT showed a linear haematoma in the left occipital lobe far from the site of the ventricular puncture. MRI revealed a small contusion in the left cerebellar hemisphere, disconnection of the left tentorial membrane and linear haematoma on a straight line. These facts suggested that the shunt tunneller had penetrated the skull through the craniotomy of the posterior fossa. This is a rare complication of VP shunting, with limited cases reported in the literature.

A case of solitary subependymal giant cell astrocytoma with histopathological anaplasia and TSC2 gene alteration.

We report a case of subependymal giant cell astrocytoma (SEGA) with anaplastic histological features in a 3-year-old girl. She had no clinical manifestations of tuberous sclerosis complex (TSC) and no relevant family history. A few cases have been reported in which patients with SEGA had no other clinical manifestations of TSC (solitary SEGA). Genetic analysis using a blood sample from the patient showed no germline alterations in TSC1 or TSC2 genes, while the tumor tissue exhibited loss of heterozygosity (LOH) in TSC2. SEGAs are benign, slowly growing tumors that rarely have significant mitotic activity. However, histopathological examination in the present case revealed high mitotic activity and necrosis besides the typical large plump cells arranged in sheets. This may be the first genetically proven case of a solitary SEGA with histopathological anaplastic features. In this report, we reviewed solitary SEGAs and histopathological malignancy in SEGA.

Microcatheter delivery of neurotherapeutics: compatibility with mesenchymal stem cells.

OBJECTIVE: Bone marrow-derived human mesenchymal stem cells (BM-hMSCs) have been used in clinical trials for the treatment of several neurological disorders. MSCs have been explored as a delivery modality for targeted viral therapeutic agents in the treatment of intracranial pathologies. Delta-24-RGD, a tumor-selective oncolytic adenovirus designed to target malignant glioma cells, has been shown to be effective in animal models and in a recent clinical trial. However, the most efficient strategy for delivering oncolytic therapies remains unclear. BM-hMSCs have been shown to home toward glioma xenografts after intracarotid delivery. The feasibility of selective intraarterial infusion of BM-hMSCs loaded with Delta-24-RGD (BM-hMSC-Delta-24) to deliver the virus to the tumor is being investigated. To evaluate the feasibility of endovascular intraarterial delivery, the authors tested in vitro the compatibility of BM-hMSC-Delta-24 with a variety of commercially available, clinically common microcatheters. METHODS: BM-hMSCs were cultured, transfected with Delta-24-RGD, and resuspended in 1% human serum albumin. The solution was then injected via 4 common neuroendovascular microcatheters of different inner diameters (Marathon, Echelon-14, Marksman, and SL-10). Cell count and viability after injection through the microcatheters were assessed, including tests of injection velocity and catheter configuration. Transwell assays were performed with the injected cells to test the efficacy of BM-hMSC-Delta-24 activity against U87 glioma cells. BM-hMSC-Delta-24 compatibility was also tested with common neuroendovascular medications: Omnipaque, verapamil, and heparin. RESULTS: The preinfusion BM-hMSC-Delta-24 cell count was 1.2 × 105 cells/ml, with 98.7% viability. There was no significant difference in postinfusion cell count or viability for any of the catheters. Increasing the injection velocity from 1.0 ml/min to 73.2 ml/min, or modifying the catheter shape from straight to tortuous, did not significantly reduce cell count or viability. Cell count and viability remained stable for up to 5 hours when the cell solution was stored on ice. Mixing BM-hMSC-Delta-24 with clinical concentrations of Omnipaque, verapamil, and heparin prior to infusion did not alter cell count or viability. Transwell experiments demonstrated that the antiglioma activity of BM-hMSC-Delta-24 was maintained after infusion. CONCLUSIONS: BM-hMSC-Delta-24 is compatible with a wide variety of microcatheters and medications commonly used in neuroendovascular therapy. Stem cell viability and viral agent activity do not appear to be affected by catheter configuration or injection velocity. Commercially available microcatheters can be used to deliver stem cell neurotherapeutics via intraarterial routes.

A rare presentation of a pediatric neurenteric cyst as an intra-axial pontine lesion: A case report with a 5-year follow-up.

BACKGROUND: Intracranial neurenteric cysts are rare, benign, and slow-growing tumors. However, we encountered a pediatric case that the cyst expansion occurred in a short period of time resulting in rapid deterioration of the patient's symptoms. CASE DESCRIPTION: A previously healthy 7-year-old girl had a week history of dysarthric speech and diplopia along with headaches. Her magnetic resonance images (MRI) showed an abnormal cystic mass in her brainstem. Her symptoms were deteriorated for 1 month and her second MRI revealed an enlargement of the cystic lesion. The tumor biopsy and cyst drainage were carried out and histopathological examination of the cyst wall showed columnar epithelium containing ciliated cells. The final diagnosis of her tumor was neurenteric cyst. CONCLUSION: We report a pediatric case of a neurenteric cyst in the brainstem, which expanded in a short period, and review this rare entity.

A Comprehensive Method for Detecting Fusion Genes in Paediatric Brain Tumours.

BACKGROUND: Fusion genes driving tumourigenesis have drawn the attention of researchers and oncologists. Despite the importance of such molecular alterations, there are no comprehensive reproducible methods for detecting fusion genes. MATERIALS AND METHODS: Nineteen paediatric brain tumours of five types, namely pilocytic astrocytoma, oligodendroglioma, anaplastic astrocytoma, glioblastoma and, ganglioglioma, were examined to detect fusion genes using a pyrosequencing-based method following RNA isolation, cDNA synthesis and real-time polymerase chain reaction. RESULTS: Our method successfully detected KIAA1549-v-raf murine sarcoma viral oncogene homolog B1 (BRAF) fusion in 14 out of 19 patients suffering from five types of paediatric brain tumours providing information on fusion breakpoints within 2 h. CONCLUSION: A comprehensive method for detecting fusion genes in paediatric brain tumours was evaluated. This method identified KIAA1549-BRAF fusion variants quickly. Our results may help researchers interested in the role of fusion genes in tumourigenesis.

Glioblastoma stem cell-derived exosomes induce M2 macrophages and PD-L1 expression on human monocytes.

Exosomes can mediate a dynamic method of communication between malignancies, including those sequestered in the central nervous system and the immune system. We sought to determine whether exosomes from glioblastoma (GBM)-derived stem cells (GSCs) can induce immunosuppression. We report that GSC-derived exosomes (GDEs) have a predilection for monocytes, the precursor to macrophages. The GDEs traverse the monocyte cytoplasm, cause a reorganization of the actin cytoskeleton, and skew monocytes toward the immune suppresive M2 phenotype, including programmed death-ligand 1 (PD-L1) expression. Mass spectrometry analysis demonstrated that the GDEs contain a variety of components, including members of the signal transducer and activator of transcription 3 (STAT3) pathway that functionally mediate this immune suppressive switch. Western blot analysis revealed that upregulation of PD-L1 in GSC exosome-treated monocytes and GBM-patient-infiltrating CD14+ cells predominantly correlates with increased phosphorylation of STAT3, and in some cases, with phosphorylated p70S6 kinase and Erk1/2. Cumulatively, these data indicate that GDEs are secreted GBM-released factors that are potent modulators of the GBM-associated immunosuppressive microenvironment.

Mesenchymal stem cells as natural biofactories for exosomes carrying miR-124a in the treatment of gliomas.

Background: MicroRNAs (miRs) are promising new therapeutics for glioblastoma. However, which miRs are most effective against glioblastomas and how these miRs should be delivered are major unanswered problems. Methods: To identify potent antiglioma miRs, we selected 8 miRs based on a literature search and screened them against a panel of glioma stem cell (GSC) lines, representing all of the glioblastoma subtypes defined by The Cancer Genome Atlas. To address delivery, we tested the hypothesis that ex vivo cultured bone marrow-derived mesenchymal stem cells (MSCs) can package miRs into exosomes and that these engineered exosomes can systemically deliver antiglioma miRs to glioblastomas. Results: Of the screened miRs, we identified miR-124a as the most effective antiglioma agent against GSCs. We then transduced MSCs with lentivirus vectors containing miR-124a and isolated vesicles from the medium. Electron microscopy, western blotting, and Nanosight proved that the isolated vesicles were exosomes. Quantitative PCR documented that these exosomes contained high levels of miR-124a, which was not present in control exosomes. In vitro treatment of GSCs with exosomes containing miR-124a (Exo-miR124) resulted in a significant reduction in viability and clonogenicity of GSCs compared with controls. In vivo treatment of mice harboring intracranial GSC267 with systemically delivered Exo-miR124 resulted in 50% of animals living long term. No evidence of tumor was present on histological analysis of the survivors. Mechanistic studies showed that miR-124a acts by silencing Forkhead box (FOX)A2, resulting in aberrant intracellular lipid accumulation. Conclusion: MSCs can be used as natural biofactories to produce Exo-miR124, which is an effective antiglioma agent worthy of further clinical evaluation.

Mesenchymal stromal cells for the delivery of oncolytic viruses in gliomas.

Mesenchymal stromal cells (MSCs) are a type of adult stem cell that has been exploited for the treatment of a variety of diseases, including cancer. In particular, MSCs have been studied extensively for their ability to treat glioblastoma (GBM), the most common and deadly form of brain cancer in adults. MSCs are attractive therapeutics because they can be obtained relatively easily from patients, are capable of being expanded numerically in vitro, can be easily engineered and are inherently capable of homing to tumors, making them ideal vehicles for delivering biological antitumoral agents. Oncolytic viruses are promising biological therapeutic agents that have been used in the treatment of GBMs, and MSCs are currently being explored as a means of delivering these viruses. Here we review the role of MSCs in the treatment of GBMs, focusing on the intersection of MSCs and oncolytic viruses.

Carotid cavernous sinus fistula caused by dental implant-associated infection.

A 61-year-old woman presented with painful ophthalmoplegia, Tolosa-Hunt syndrome. The patient had undergone a placement of dental implant 5 months before the presentation and had a local maxillary sinusitis 1 month later. She had not been aware of any preceding head trauma or infection. On examination, the patient showed serious right oculomotor nerve paresis and retro-orbital pain. Blood examination showed normal findings. Magnetic resonance imaging identified abnormal structure in the right cavernous sinus with flow void signals. Angiography revealed a carotid cavernous sinus fistula fed by the intracavernous branches of the internal carotid artery on both sides, right internal maxillary and middle meningeal arteries, and left ascending pharyngeal artery. The patient underwent coil embolization via both external carotid arteries. We assumed that local maxillary sinusitis caused by dental implant might spread hematogenously into the sphenoid and cavernous sinuses and formed a carotid cavernous sinus fistula, which presented with Tolosa-Hunt syndrome. Implant-associated infection has to be managed promptly with adequate manner before it spreads.

Arteriovenous fistula arising from the persistent primitive olfactory artery with dual supply from the bilateral anterior ethmoidal arteries.

A 59-year-old male presented with generalized seizure. The patient had not been aware of any traumatic head injuries or preceding infection, and had no contributory medical history. On admission, he was alert and well oriented, without neurological impairment or headache. He was afebrile and blood examination showed no abnormal findings. Computed tomography revealed an irregular intracerebral hematoma, 3 x 1.5 cm in diameter, in the left rectal gyrus. Cerebral angiography showed an arteriovenous fistula (AVF) in the anterior cranial fossa supplied only by the persistent primitive olfactory artery (PPOA) originating from the anterior cerebral artery, forming a shunt to an ascending cortical vein, and drained by the superior sagittal sinus. The patient underwent endovascular obliteration of the AVF via the transarterial route. Immediately after successful isolation, angiography showed that the bilateral anterior ethmoidal arteries supplied the AVF. The feeding branches from the left anterior ethmoidal artery were completely occluded via the ophthalmic artery, but introduction of the catheter into the right ophthalmic artery markedly decreased the stump pressure. Follow-up angiography performed at 3 and 8 weeks following embolization showed spontaneous resolution of the residual AVF without findings of recanalization or new abnormal channels. AVF arising in the anterior cranial fossa may be associated with an unusual pattern of the blood supply when including the PPOA.

[Simultaneous presentation of Tolosa-Hunt syndrome and oculomotor nerve palsy due to the nonruptured internal carotid-posterior communicating artery aneurysm: a case report].

A 45-year-old female developed mild dysesthesia and swelling, followed by ptosisand trigeminal pain, in the right side of the face. Her past medical history was unremarkable, and she had not been aware of any infectious sign. A local otolaryngologist administered glucocorticoid therapy that resolved the face pain, but the ptosis persisted. Neurological examination found complete right oculomotor nerve paresis and mild sensory loss in the first and second segments of the right trigeminal nerve. Blood examination found no abnormalities. Neuroimaging revealed a saccular aneurysm at the branching site of the posterior communicating artery, projecting posteriorly and adjacent to the dorsum sellae, without other intracranial abnormalities. Cerebral angiography demonstrated poor opacification of the superior ophthalmic vein and cavernous sinus on the right side. The patient underwent coil embolization under a diagnosis of symptomatic aneurysm, but her oculomotor neuropathy was only partially improved. We thought that the impairment of the oculomotor function by inflammatory reaction in the cavernous sinus and mechanical compression by the aneurysm had already persisted for too long for post-treatment recovery. We think that the simultaneous occurrence of Tolosa-Hunt syndrome and oculomotor nerve palsy may have resulted because trigeminal neuralgia had increased the blood pressure to induce rapid growth of the preexisting aneurysm, or the inflammatory reaction in the cavernous sinus had promoted the growth of the aneurysm, or that the association was by chance.