YAP/TAZ Transcriptional Coactivators Create Therapeutic Vulnerability to Verteporfin in EGFR-mutant Glioblastoma.

PURPOSE: Glioblastomas (GBMs), neoplasms derived from glia and neuroglial progenitor cells, are the most common and lethal malignant primary brain tumors diagnosed in adults, with a median survival of 14 months. GBM tumorigenicity is often driven by genetic aberrations in receptor tyrosine kinases, such as amplification and mutation of EGFR. EXPERIMENTAL DESIGN: Using a Drosophila glioma model and human patient-derived GBM stem cells and xenograft models, we genetically and pharmacologically tested whether the YAP and TAZ transcription coactivators, effectors of the Hippo pathway that promote gene expression via TEA domain (TEAD) cofactors, are key drivers of GBM tumorigenicity downstream of oncogenic EGFR signaling. RESULTS: YAP and TAZ are highly expressed in EGFR-amplified/mutant human GBMs, and their knockdown in EGFR-amplified/mutant GBM cells inhibited proliferation and elicited apoptosis. Our results indicate that YAP/TAZ-TEAD directly regulates transcription of SOX2, C-MYC, and EGFR itself to create a feedforward loop to drive survival and proliferation of human GBM cells. Moreover, the benzoporphyrin derivative verteporfin, a disruptor of YAP/TAZ-TEAD-mediated transcription, preferentially induced apoptosis of cultured patient-derived EGFR-amplified/mutant GBM cells, suppressed expression of YAP/TAZ transcriptional targets, including EGFR, and conferred significant survival benefit in an orthotopic xenograft GBM model. Our efforts led us to design and initiate a phase 0 clinical trial of Visudyne, an FDA-approved liposomal formulation of verteporfin, where we used intraoperative fluorescence to observe verteporfin uptake into tumor cells in GBM tumors in human patients. CONCLUSIONS: Together, our data suggest that verteporfin is a promising therapeutic agent for EGFR-amplified and -mutant GBM.

Use of Navigated Ultrasonic Bone Cutting Tool for En Bloc Resection of Thoracic Chondrosarcoma: Technical Report.

BACKGROUND: En bloc surgical resection with wide margins offers the best local control rates for chondrosarcoma of the spine. OBJECTIVE: To describe the surgical technique for en bloc resection of a large thoracic chondrosarcoma using image guidance for a complex osteotomy with an ultrasonic bone cutting device (Misonix, Farmingdale, New York). METHODS: A 2-stage procedure was performed for resection of a thoracic chondrosarcoma involving the T3-T7 vertebral bodies. During the first stage, a posterior approach, the ultrasonic bone cutter was precisely navigated to perform an intralaminar osteotomy as well as a multilevel split sagittal osteotomy through the vertebral bodies. In the second stage, a transthoracic approach was used to complete the en bloc resection of the specimen. Intraoperative frozen sections from the surgical margins were negative for tumor. RESULTS: The ultrasonic bone cutting device was navigated based on coregistration of the intraoperative computed tomography (CT) images and preoperative magnetic resonance imaging (MRI). Real-time navigation using coregistered images enabled identification of tumor margins within the bone and adjacent soft tissue allowing precise execution of the intralaminar and multilevel split sagittal vertebral osteotomies. Surgical video demonstrates the utility of real-time navigation to properly identify the tumor margins and guide the ultrasonic bone cutting tool during the osteotomies. CONCLUSION: We describe the use of image guidance to navigate an ultrasonic bone cutting tool for a complex en bloc resection of a multilevel thoracic spine chondrosarcoma.

Drak/STK17A Drives Neoplastic Glial Proliferation through Modulation of MRLC Signaling.

Glioblastoma (GBM) and lower grade gliomas (LGG) are the most common primary malignant brain tumors and are resistant to current therapies. Genomic analyses reveal that signature genetic lesions in GBM and LGG include copy gain and amplification of chromosome 7, amplification, mutation, and overexpression of receptor tyrosine kinases (RTK) such as EGFR, and activating mutations in components of the PI3K pathway. In Drosophila melanogaster, constitutive co-activation of RTK and PI3K signaling in glial progenitor cells recapitulates key features of human gliomas. Here we use this Drosophila glioma model to identify death-associated protein kinase (Drak), a cytoplasmic serine/threonine kinase orthologous to the human kinase STK17A, as a downstream effector of EGFR and PI3K signaling pathways. Drak was necessary for glial neoplasia, but not for normal glial proliferation and development, and Drak cooperated with EGFR to promote glial cell transformation. Drak phosphorylated Sqh, the Drosophila ortholog of nonmuscle myosin regulatory light chain (MRLC), which was necessary for transformation. Moreover, Anillin, which is a binding partner of phosphorylated Sqh, was upregulated in a Drak-dependent manner in mitotic cells and colocalized with phosphorylated Sqh in neoplastic cells undergoing mitosis and cytokinesis, consistent with their known roles in nonmuscle myosin-dependent cytokinesis. These functional relationships were conserved in human GBM. Our results indicate that Drak/STK17A, its substrate Sqh/MRLC, and the effector Anillin/ANLN regulate mitosis and cytokinesis in gliomas. This pathway may provide a new therapeutic target for gliomas.Significance: These findings reveal new insights into differential regulation of cell proliferation in malignant brain tumors, which will have a broader impact on research regarding mechanisms of oncogene cooperation and dependencies in cancer.See related commentary by Lathia, p. 1036.

Correction of Progressive Severe Cervical Kyphosis in a 21-Month-Old Patient With NF1: Surgical Technique and Review of Literature.

BACKGROUND: Severe cervical kyphosis in the setting of neurofibromatosis type 1 (NF1) is a rare manifestation of the disease in the pediatric population. Dystrophic and immature bone complicate the placement of hardware necessary for surgical correction of alignment and a review of the literature yields 4 cases of pediatric patients with NF1 requiring surgical intervention in which the youngest patient was 10 yr old. OBJECTIVE: To report the case of an 11-mo-old female with NF1 who presented with a plexiform cervical neurofibroma and focal cervical kyphosis. A comprehensive review of the literature and a detailed description of nonsurgical and surgical management for this patient population is described. METHODS: A literature review was completed for article reviewing management of pediatric patients with cervical spine injuries and NF1. The patient's chart was reviewed and the patient was followed for a year to provide adequate follow-up. Institutional Review Board (IRB)/ethics committee approval and patient consent were neither required nor sought for this study. RESULTS: The literature was reviewed, summarized, and utilized for operative planning and postoperative management. Postoperative imaging and 1-yr follow-up imaging showed anterior construct and lateral mass fusion, restoration of cervical alignment, and no neurological deficits. CONCLUSION: This is the youngest reported patient to have surgical cervical kyphosis correction in the setting of NF1. A review of the literature helped develop a long-term plan and shape a novel same-day front-back-front approach to restore alignment that will be of use to teams managing these complex patients in the future.

Beyond the World Health Organization grading of infiltrating gliomas: advances in the molecular genetics of glioma classification.

BACKGROUND: Traditional classification of diffuse infiltrating gliomas (DIGs) as World Health Organization (WHO) grades II-IV is based on histological features of a heterogeneous population of tumors with varying prognoses and treatments. Over the last decade, research efforts have resulted in a better understanding of the molecular basis of glioma formation as well as the genetic alterations commonly identified in diffuse gliomas. METHODS: A systematic review of the current literature related to advances in molecular phenotypes, mutations, and genomic analysis of gliomas was carried out using a PubMed search for these key terms. Data was studied and synthesized to generate a comprehensive review of glioma subclassification. RESULTS: This new data helps supplement the existing WHO grading scale by subtyping gliomas into specific molecular groups. The emerging molecular profile of diffuse gliomas includes the studies of gene expression and DNA methylation in different glioma subtypes. The discovery of novel mutations in isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) provides new biomarkers as points of stratification of gliomas based on prognosis and treatment response. Gliomas that harbor CpG island hypermethylator phenotypes constitute a subtype of glioma with improved survival. The difficulty of classifying oligodendroglial lineage of tumors can be aided with identification of 1p/19q codeletion. Glioblastomas (GBMs) previously described as primary or secondary can now be divided based on gene expression into proneural, mesenchymal, and classical subtypes and the identification of mutations in the promoter region of the telomerase reverse transcriptase (TERTp) have been correlated with poor prognosis in GBMs. CONCLUSIONS: Incorporation of new molecular and genomic changes into the existing WHO grading of DIGs may provide better patient prognostication as well as advance the development of patient-specific treatments and clinical trials.

Biomechanical, biochemical, and histological characterization of canine lumbar facet joint cartilage.

OBJECT: Tissue engineering appears to be a promising strategy for articular cartilage regeneration as a treatment for facet joint arthritis. Prior to the commencement of tissue engineering approaches, design criteria must be established to determine the required functional properties of the replacement tissue. As characterization of the functional properties of facet joint cartilage has not been performed previously, the objective of this study was to determine the biomechanical, biochemical, and histological properties of facet joint cartilage. METHODS: The in vitro testing was conducted using 4 lumbar spinal segments obtained from skeletally mature canines. In each specimen, articular cartilage was obtained from the superior surface of the L3-4 and L4-5 facet joints. Creep indentation was used to determine the compressive biomechanical properties, while uniaxial tensile testing yielded the Young modulus and ultimate tensile strength of the tissue. Additionally, biochemical assessments included determinations of cellularity, glycosaminoglycan (GAG) content, and collagen content, as well as enzymelinked immunosorbent assays for collagen I and II production. Finally, histological characterization included H & E staining, as well as staining for collagen and GAG distributions. RESULTS: The means +/- standard deviation values were determined. There were no differences between the 2 spinal levels for any of the assessed properties. Averaged over both levels, the thickness was 0.49 +/- 0.10 mm and the hydration was 74.7 +/- 1.7%. Additionally, the cells/wet weight (WW) ratio was 6.26 +/- 2.66 x 10(4) cells/mg and the cells/dry weight (DW) ratio was 2.51 +/- 1.21 x 10(5) cells/mg. The GAG/WW was 0.038 +/- 0.013 and the GAG/ DW was 0.149 +/- 0.049 mg/mg, while the collagen/WW was 0.168 +/- 0.026 and collagen/DW was 0.681 +/- 0.154 mg/ mg. Finally, the aggregate modulus was 554 +/- 133 kPa, the Young modulus was 10.08 +/- 8.07 MPa, and the ultimate tensile strength was 4.44 +/- 2.40 MPa. CONCLUSIONS: To the best of the authors' knowledge, this study is the first to provide a functional characterization of facet joint articular cartilage, thus providing design criteria for future tissue engineering studies.

Presynaptic establishment of the synaptic cleft extracellular matrix is required for post-synaptic differentiation.

Formation and regulation of excitatory glutamatergic synapses is essential for shaping neural circuits throughout development. In a Drosophila genetic screen for synaptogenesis mutants, we identified mind the gap (mtg), which encodes a secreted, extracellular N-glycosaminoglycan-binding protein. MTG is expressed neuronally and detected in the synaptic cleft, and is required to form the specialized transsynaptic matrix that links the presynaptic active zone with the post-synaptic glutamate receptor (GluR) domain. Null mtg embryonic mutant synapses exhibit greatly reduced GluR function, and a corresponding loss of localized GluR domains. All known post-synaptic signaling/scaffold proteins functioning upstream of GluR localization are also grossly reduced or mislocalized in mtg mutants, including the dPix-dPak-Dock cascade and the Dlg/PSD-95 scaffold. Ubiquitous or neuronally targeted mtg RNA interference (RNAi) similarly reduce post-synaptic assembly, whereas post-synaptically targeted RNAi has no effect, indicating that presynaptic MTG induces and maintains the post-synaptic pathways driving GluR domain formation. These findings suggest that MTG is secreted from the presynaptic terminal to shape the extracellular synaptic cleft domain, and that the cleft domain functions to mediate transsynaptic signals required for post-synaptic development.