Meningioma with rhabdoid features: Pathologic findings in dogs.

Rhabdoid meningioma is a rare type of meningeal neoplasm in humans. This study reports the clinical, pathological, and ultrastructural features of 4 cases of canine meningioma with rhabdoid features. The cases were female and 8 to 12 years of age. Biopsies from complete surgical resections were examined for all cases. The whole brain with tumor recurrence was collected at necropsy in 2 dogs. Histologically, the tumors consisted of discohesive sheets of oval-polygonal cells with abundant eosinophilic cytoplasm and occasional paranuclear hyaline-like inclusions. Cells were intensely immunopositive for vimentin, negative for melan A and S100 protein in all cases, and showed variable immunolabeling for cytokeratin in 2 cases. Focal glial fibrillary acidic protein (GFAP)-immunopositive cells were present in 1 case. Ultrastructurally, the rhabdoid cells in case 1 contained prominent cytoplasmic whorls of intermediate filaments, recapitulating the ultrastructural features of rhabdoid meningioma in humans. In cases 2 and 3, the meningioma cells contained interdigitating cell processes folded in a maze-like fashion resembling rhabdoid-like meningioma in humans. In case 4, the voluminous cytoplasm contained many round-to-flattened mitochondria admixed with rough endoplasmic reticulum, indicating a predominant oncocytic differentiation and not the rhabdoid differentiation suggested by light microscopy. Thus, rhabdoid morphology occurs in different types of meningiomas, and ultrastructural findings are essential for a correct diagnosis.

Prognostic histopathologic features of canine glial tumors.

Gliomas are relatively common tumors in aged dogs (especially brachycephalic breeds), and the dog is proving to be useful as a translational model for humans with brain tumors. Hitherto, there is relatively little prognostic data for canine gliomas and none on outcome related to specific histological features. Histologic sections of tumor biopsies from 33 dogs with glioma treated with surgical resection and immunotherapy and 21 whole brains obtained postmortem were reviewed. Tumors were diagnosed as astrocytic, oligodendroglial, or undefined glioma using Comparative Brain Tumor Consortium criteria. Putative features of malignancy were evaluated, namely, mitotic counts, glomeruloid vascularization, and necrosis. For biopsies, dogs with astrocytic tumors lived longer than those with oligodendroglial or undefined tumor types (median survival 743, 205, and 144 days, respectively). Dogs with low-grade gliomas lived longer than those with high-grade gliomas (median survival 734 and 194 days, respectively). Based on analysis of tumor biopsies, low mitotic counts, absence of glomeruloid vascularization, and absence of necrosis correlated with increased survival (median 293, 223, and 220 days, respectively), whereas high mitotic counts, glomeruloid vascularization, and necrosis correlated with poor survival (median 190, 170, and 154 days, respectively). Mitotic count was the only histological feature in biopsy samples that significantly correlated with survival (P < .05). Whole-brain analyses for those same histologic features had similar and more robust correlations, and were statistically significant for all features (P < .05). The small size of biopsy samples may explain differences between biopsy and whole-brain tumor data. These findings will allow more accurate prognosis for gliomas.

Naturally Occurring Canine Glioma as a Model for Novel Therapeutics.

BACKGROUND: Current animal models of glioma are limited to small animal models, which are less predictive of treatment of human disease. Canines often develop gliomas de novo, but the natural history of the disease is not well described. OBJECTIVE: We provide data for naturally occurring canine gliomas; evaluating medical and surgical therapies. METHODS: We reviewed medical records of pet dogs with a presumptive diagnosis of glioma from MRI imaging that underwent surgery as part of the Canine Brain Tumor Clinical Trials Program. Breed, age, sex, median progression-free, and overall survival times and cause of death were recorded for multivariate analysis. RESULTS: Ninety five dogs (56 male; mean age = 8.3 years) were included, but nine were excluded as final pathology was non-neoplastic. Gross total resection was reported in 81 cases based on postoperative MRI. Seventy had high-grade tumors (grade III or IV). Eighty three dogs presented with seizures, being the most common presenting clinical sign. Median survival after surgery was 723 days (95% CI 343-1103) for grade II tumors, 301 days (197-404) for grade III and 200 days (126-274) for grade IV (p = .009 Kaplan-Meier survival analysis; Log Rank test). Age (cox regression, p = .14) or sex (Kaplan-Meier test, p = .22) did not predict survival. CONCLUSIONS: This study establishes normative data for a model exploiting dogs with naturally occurring glioma, which can be used to test novel therapies prior to translation to human trials. Further work will focus on the effects of different therapies, including chemotherapy, radiation therapy, and immunotherapy.

CD8⁺ T Cell-Independent Immune-Mediated Mechanisms of Anti-Tumor Activity.

Despite the growing number of preclinical and clinical trials focused on immunotherapy for the treatment of malignant gliomas, the prognosis for this disease remains grim. Cancer immunotherapy seeks to recruit an effective immune response to eliminate tumor cells. To date, cancer vaccines have shown only limited effectiveness because of our incomplete understanding of the necessary effector cells and mechanisms that yield efficient tumor clearance. CD8+ T cell cytotoxic activity has long been proposed as the primary effector function necessary for tumor regression. However, there is increasing evidence that indicates that components of the immune system other than CD8+ T cells play important roles in tumor eradication and control. The following review should provide an understanding of the mechanisms involved in an effective antitumor response to guide future therapeutic designs. The information provided suggests an alternate means of effective tumor clearance in malignant glioma to the canonical CD8+ cytotoxic T cell mechanism.

CD8+ T cell-independent tumor regression induced by Fc-OX40L and therapeutic vaccination in a mouse model of glioma.

Despite the growing number of preclinical and clinical trials focused on immunotherapy for the treatment of malignant gliomas, the prognosis for this disease remains grim. Although some promising advances have been made, the immune response stimulated as a result of immunotherapeutic protocols has been inefficient at complete tumor elimination, primarily due to our lack of understanding of the necessary effector functions of the immune system. We previously demonstrated that a tumor lysate vaccine/Fc-OX40L therapy is capable of inducing enhanced survival and tumor elimination in the GL261 mouse glioma model. The following experiments were performed to determine the mechanism(s) of action of this therapy that elicits a potent antitumor immune response. The evidence subsequently outlined indicates a CD8(+) T cell-independent and CD4(+) T cell-, NK cell-, and B cell-dependent means of prolonged survival. CD8(+) T cell-independent tumor clearance is surprising considering the current focus of many cancer immunotherapy protocols. These results provide evidence for CD8(+) T cell-independent means of antitumor response and should lead to additional examination of the potential manipulation of this mechanism for future treatment strategies.

Victory and defeat in the induction of a therapeutic response through vaccine therapy for human and canine brain tumors: a review of the state of the art.

Anti-tumor immunotherapy using tumor lysate-based vaccines has made great advances over recent decades. Cancer vaccines aim to elicit adaptive immune responses through various pathways by providing tumor and tumor-associated antigens with an immune stimulant or adjuvant. These anti-tumor vaccines are therefore developed as personalized treatments. Utilizing tumors as a source of vaccine antigens in immunotherapy has demonstrated promising results with minimal toxicity. However, to date, researchers have failed to overcome the overpowering immune suppressive effects within the tumor microenvironment. Immune suppression occurs naturally via multiple mechanisms. These mechanisms serve an important homeostatic role restoring a normal tissue microenvironment following an inflammatory response. Due to these suppressive mechanisms and the inherent heterogeneity of tumors, it is imperative to then elicit and maintain a specific tumoricidal response if vaccine therapy or some other combination of reagents is chosen. In this review, we focus on the historical use of tumors as a source of antigens to elicit a tumoricidal response and the limitations encountered that prevent greater success in immunotherapy. We describe the advantages and disadvantages of various vaccines and their ineffectiveness due to tumor-induced immune suppression.

Comparison of peripheral leukocyte parameters in patients receiving conventionally and hypofractionated radiotherapy schemes for the treatment of newly diagnosed glioblastoma.

Introduction: Treatment for glioblastomas, aggressive and nearly uniformly fatal brain tumors, provide limited long-term success. Immunosuppression by myeloid cells in both the tumor microenvironment and systemic circulation are believed to contribute to this treatment resistance. Standard multi-modality therapy includes conventionally fractionated radiotherapy over 6 weeks; however, hypofractionated radiotherapy over 3 weeks or less may be appropriate for older patients or populations with poor performance status. Lymphocyte concentration changes have been reported in patients with glioblastoma; however, monocytes are likely a key cell type contributing to immunosuppression in glioblastoma. Peripheral monocyte concentration changes in patients receiving commonly employed radiation fractionation schemes are unknown. Methods: To determine the effect of conventionally fractionated and hypofractionated radiotherapy on complete blood cell leukocyte parameters, retrospective longitudinal concentrations were compared prior to, during, and following standard chemoradiation treatment. Results: This study is the first to report increased monocyte concentrations and decreased lymphocyte concentrations in patients treated with conventionally fractionated radiotherapy compared to hypofractionated radiotherapy. Discussion: Understanding the impact of fractionation on peripheral blood leukocytes is important to inform selection of dose fractionation schemes for patients receiving radiotherapy.

Establishment and characterization of two novel patient-derived lines from canine high-grade glioma.

High-grade glioma is an aggressive cancer that occurs naturally in pet dogs. Canine high-grade glioma (cHGG) is treated with radiation, chemotherapy or surgery, but has no curative treatment. Within the past eight years, there have been advances in our imaging and histopathology standards as well as genetic charactereization of cHGG. However, there are only three cHGG cell lines publicly available, all of which were derived from astrocytoma and established using methods involving expansion of tumour cells in vitro on plastic dishes. In order to provide more clinically relevant cell lines for studying cHGG in vitro, the goal of this study was to establish cHGG patient-derived lines, whereby cancer cells are expanded in vivo by injecting cells into immunocompromized laboratory mice. The cells are then harvested from mice and used for in vitro studies. This method is the standard in the human field and has been shown to minimize the acquisition of genetic alterations and gene expression changes from the original tumour. Through a multi-institutional collaboration, we describe our methods for establishing two novel cHGG patient-derived lines, Boo-HA and Mo-HO, from a high-grade astrocytoma and a high-grade oligodendroglioma, respectively. We compare our novel lines to G06-A, J3T-Bg, and SDT-3G (traditional cHGG cell lines) in terms of proliferation and sensitivity to radiation. We also perform whole genome sequencing and identify an NF1 truncating mutation in Mo-HO. We report the characterization and availability of these novel patient-derived lines for use by the veterinary community.

Clinical application of recombinant human bone morphogenetic protein in cats and dogs: a review of 13 cases.

Databases (2001-2008) for cases in which recombinant bone morphogenetic protein (rhBMP) was used to aid in management of orthopedic disease were reviewed and cases were categorized as non-unions, delayed unions, and cases expected to heal with difficulty. If follow-up in the medical record was < 6 mo for live animals, owners were surveyed by telephone. Thirteen cases (11 dogs, 2 cats) were identified; OP-1 (rhBMP-7) was used in 3 cases and INFUSE (rhBMP-2) in 10. Mean time from injury to rhBMP use for non- and delayed union cases was 156 d; mean time from rhBMP use to radiographic healing was 101 d. No systemic side effects were reported. All patients achieved clinical and radiographic bone union following rhBMP administration. Recombinant human BMP was used in 13 veterinary patients to successfully achieve bone union without serious deleterious effects in a variety of clinical applications.

CD200 Immune-Checkpoint Peptide Elicits an Anti-glioma Response Through the DAP10 Signaling Pathway.

Numerous therapies aimed at driving an effective anti-glioma response have been employed over the last decade; nevertheless, survival outcomes for patients remain dismal. This may be due to the expression of immune-checkpoint ligands such as PD-L1 by glioblastoma (GBM) cells which interact with their respective receptors on tumor-infiltrating effector T cells curtailing the activation of anti-GBM CD8+ T cell-mediated responses. Therefore, a combinatorial regimen to abolish immunosuppression would provide a powerful therapeutic approach against GBM. We developed a peptide ligand (CD200AR-L) that binds an uncharacterized CD200 immune-checkpoint activation receptor (CD200AR). We sought to test the hypothesis that CD200AR-L/CD200AR binding signals via he DAP10&12 pathways through in vitro studies by analyzing transcription, protein, and phosphorylation, and in vivo loss of function studies using inhibitors to select signaling molecules. We report that CD200AR-L/CD200AR binding induces an initial activation of the DAP10&12 pathways followed by a decrease in activity within 30 min, followed by reactivation via a positive feedback loop. Further in vivo studies using DAP10&12KO mice revealed that DAP10, but not DAP12, is required for tumor control. When we combined CD200AR-L with an immune-stimulatory gene therapy, in an intracranial GBM model in vivo, we observed increased median survival, and long-term survivors. These studies are the first to characterize the signaling pathway used by the CD200AR, demonstrating a novel strategy for modulating immune checkpoints for immunotherapy currently being analyzed in a phase I adult trial.

CD200 Checkpoint Reversal: A Novel Approach to Immunotherapy.

PURPOSE: Advances in immunotherapy have revolutionized care for some patients with cancer. However, current checkpoint inhibitors are associated with significant toxicity and yield poor responses for patients with central nervous system tumors, calling into question whether cancer immunotherapy can be applied to glioblastoma multiforme. We determined that targeting the CD200 activation receptors (CD200AR) of the CD200 checkpoint with a peptide inhibitor (CD200AR-L) overcomes tumor-induced immunosuppression. We have shown the clinical efficacy of the CD200AR-L in a trial in companion dogs with spontaneous high-grade glioma. Addition of the peptide to autologous tumor lysate vaccines significantly increased the median overall survival to 12.7 months relative to tumor lysate vaccines alone, 6.36 months. EXPERIMENTAL DESIGN: This study was developed to elucidate the mechanism of the CD200ARs and develop a humanized peptide inhibitor. We developed macrophage cell lines with each of four CD200ARs knocked out to determine their binding specificity and functional response. Using proteomics, we developed humanized CD200AR-L to explore their effects on cytokine/chemokine response, dendritic cell maturation and CMV pp65 antigen response in human CD14+ cells. GMP-grade peptide was further validated for activity. RESULTS: We demonstrated that the CD200AR-L specifically targets a CD200AR complex. Moreover, we developed and validated a humanized CD200AR-L for inducing chemokine response, stimulating immature dendritic cell differentiation and significantly enhanced an antigen-specific response, and determined that the use of the CD200AR-L downregulated the expression of CD200 inhibitory and PD-1 receptors. CONCLUSIONS: These results support consideration of a CD200AR-L as a novel platform for immunotherapy against multiple cancers including glioblastoma multiforme.

Evaluation of scintigraphy to assess incorporation of intercalary cortical bone grafts in sheep.

OBJECTIVE: To evaluate scintigraphy, radiography, and histomorphometric analysis for assessing incorporation of intercalary bone grafts and to compare incorporation of cortical autografts and allografts by the recipient. ANIMALS: 12 skeletally mature sheep. PROCEDURES: A 5-cm tibial defect was filled with a cortical allograft (n = 6) or autograft (6) and stabilized with an interlocking nail. Radiography, scintigraphy, and fluorochrome bone labeling were performed every 3 months for 24 months. Radiographic evaluation included grading of the host and graft union and assessment of implants and grafts. Technetium-99m-hydroxymethylene diphosphonate radionuclide uptake was measured. Sheep were euthanatized 24 months after surgery, and bone formation was evaluated via histomorphometric analysis of fluorochrome labeling. RESULTS: Complete union was detected on radiographs by 21 months in all sheep but developed earlier in sheep that received an autograft versus in those that received an allograft. Radionuclide uptake peaked at 3 months and returned to presurgical values at 12 months. Histomorphometric analysis revealed fluorochrome labeling corresponding to each time point, with most bone formation at 9 through 15 months. Scintigraphy findings did not correlate well with fluorochrome labeling of newly formed bone. CONCLUSIONS AND CLINICAL RELEVANCE: Although bone production around cortical bone grafts was detected by use of scintigraphy, this method did not provide accurate assessment of graft incorporation in sheep. Furthermore, bone produced by activated periosteum could not be distinguished scintgraphically from bone that replaced the graft. Intercalary autografts healed more rapidly and had greater incorporation into the host bone, compared with findings for allografts.

Biomechanical analysis of stacked plating techniques to stabilize distal radial fractures in small dogs.

OBJECTIVE: To evaluate the fatigue life of partially stacked and fully stacked (1.5/2.0 and 2.0/2.7 mm) veterinary cuttable plates (VCP) in a fracture gap model of the distal aspect of the radius. STUDY DESIGN: In vitro biomechanical study. METHODS: Constructs (n=4/group) were assembled for each of 8 groups using 8-hole plates (1.5/2.0 and 2.0/2.7 mm VCP) in the following configurations: unstacked; 2-hole stacked centered over the gap (COG); 4-hole stacked COG; and fully stacked. Plate(s) were secured to 2 separate polyvinylchloride pipe lengths, mounted to a mechanical testing system with a custom jig, and were loaded in axial compression for 10(6) cycles at 10 Hz or until failure at 6-60 N for the 1.5/2.0 mm VCP and 10-100 N for the 2.0/2.7 mm VCP. Differences in number of cycles, stiffness, and failure mode were recorded. RESULTS: All construct failures occurred through a screw hole adjacent to the gap. Fully stacked and 4-hole stacked 1.5/2.0 and 2.0/2.7 mm VCP withstood 10(6) cycles. Fatigue life and stiffness of the 1.5/2.0 or 2.0/2.7 mm unstacked constructs were significantly less than the other constructs. Differences were identified in stiffness among the 1.5/2.0 mm stacked constructs and in fatigue life among the 2.0/2.7 mm VCP stacked constructs. CONCLUSION: Four-hole partially stacked VCP (either 1.5/2.0 or 2.0/2.7 mm) have comparable mechanical properties to fully stacked VCP. CLINICAL RELEVANCE: Partial stacking of 2 holes of VCP on both sides of the fracture gap may provide sufficient strength for healing, but this premise must be tested in vivo.

Treatment Combining CD200 Immune Checkpoint Inhibitor and Tumor-Lysate Vaccination after Surgery for Pet Dogs with High-Grade Glioma.

Recent advances in immunotherapy have included inhibition of immune checkpoint proteins in the tumor microenvironment and tumor lysate-based vaccination strategies. We combined these approaches in pet dogs with high-grade glioma. Administration of a synthetic peptide targeting the immune checkpoint protein, CD200, enhanced the capacity of antigen-presenting cells to prime T-cells to mediate an anti-glioma response. We found that in canine spontaneous gliomas, local injection of a canine-specific, CD200-directed peptide before subcutaneous delivery of an autologous tumor lysate vaccine prolonged survival relative to a historical control treated with autologous tumor lysate alone (median survivals of 12.7 months and 6.36 months, respectively). Antigen-presenting cells and T-lymphocytes primed with this peptide suppressed their expression of the inhibitory CD200 receptor, thereby enhancing their ability to initiate immune reactions in a glioblastoma microenvironment replete with the immunosuppressive CD200 protein. These results support consideration of a CD200 ligand as a novel glioblastoma immunotherapeutic agent.

Development of a Novel Canine Model of Ischemic Stroke: Skull Base Approach with Transient Middle Cerebral Artery Occlusion.

OBJECTIVE: Although canine stroke models have several intrinsic advantages, establishing consistent and reproducible territorial stroke in these models has been challenging because of the abundance of collateral circulation. We have described a skull-base surgical approach that yields reproducible stroke volumes. METHODS: Ten male beagles were studied. In all 10 dogs, a craniectomy was performed to expose the circle of Willis. Cerebral aneurysm clips were temporarily applied to the middle cerebral artery (MCA), anterior cerebral artery (ACA), posterior cerebral artery, and/or ophthalmic artery (OA) for 1 hour, followed by cauterization of the distal MCA pial collateral vessels. Indocyanine green angiography was performed to assess the local blood flow to the intended area of infarction. The dogs' neurologic examination was evaluated, and the stroke burden was quantified using magnetic resonance imaging. RESULTS: High mortality was observed after 1-hour clip occlusion of the posterior cerebral artery, MCA, ACA, and OA (n = 4). Without coagulation of the MCA collateral vessels, 1-hour occlusion of the MCA and/or ACA and OA yielded inconsistent stroke volumes (n = 2). In contrast, after coagulation of the distal MCA pial collateral vessels, 1-hour occlusion of the MCA, ACA, and OA yielded consistent territorial stroke volumes (n = 4; average stroke volume, 9.13 ± 0.90 cm3; no surgical mortalities), with reproducible neurologic deficits. CONCLUSION: Consistent stroke volumes can be achieved in male beagles using a skull base surgical approach with temporary occlusion of the MCA, ACA, and OA when combined with cauterization of the distal MCA pial collateral vessels.

Fish oil decreases matrix metalloproteinases in knee synovia of dogs with inflammatory joint disease.

This study was designed to determine whether dietary fish oil affects the expression and activity of matrix metalloproteinases (MMP), tissue inhibitors of MMP-2 (TIMP-2) and urokinase plasminogen activator (uPA) in synovial fluid from dogs with spontaneously occurring stifle (knee) instability in a single hind limb resulting from acute cranial cruciate ligament (CCL) injury. Two groups of 12 dogs were fed diets from 1 week prior to surgery on the affected knee to 56 days post-surgery. The fish oil and control diets provided 90 and 4.5 mg, respectively, of combined eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)/kg body weight per day. Plasma and synovial fluid, from both surgical and nonsurgical knee joints, were obtained at start of the diet (-7), surgery day (0) and 7, 14, 28 and 56 days post-surgery. Plasma total EPA and DHA were significantly increased, and plasma total arachidonic acid (AA) was significantly decreased by the fish oil diet. In synovial fluid from the nonsurgical knee, fish oil treatment significantly decreased proMMP-2 expression at Days 7 and 14, and proMMP-9 expression at Day 56, and uPA activity at 28 days and significantly increased TIMP-2 expression at Days 7 and 28. There were no differences in MMP expression or activity, TIMP-2 expression and uPA activity in the surgical joint synovial fluid at any time throughout the study. These results suggest that dietary fish oil may exert beneficial effects on synovial fluid MMP and TIMP-2 equilibrium in the uninjured stifle of dogs with unilateral CCL injury.

Incidence of and risk factors for major complications or death in dogs undergoing cytoreductive surgery for treatment of suspected primary intracranial masses.

OBJECTIVE To determine incidence of and risk factors for major complications occurring in dogs within 30 days after cytoreductive surgery performed by a single pair of surgeons for treatment of suspected primary intracranial masses. DESIGN Retrospective cohort study. ANIMALS 160 client-owned dogs that underwent cytoreductive surgery for treatment of suspected primary intracranial masses between January 2009 and December 2015 at a veterinary teaching hospital. PROCEDURES Medical records were retrospectively reviewed for complications occurring within 30 days after surgery. Data (eg, signalment, clinical signs, previous treatments, preoperative neurologic examination findings, neuroanatomical location, time from onset of clinical signs to surgery, surgical approach, and histopathologic diagnosis) were analyzed for associations with death and with development of major complications other than death. RESULTS 21 (13.1%) dogs died (11 during hospitalization and 10 after discharge) and 30 (18.8%) developed major complications other than death during the first 30 days after surgery. Dogs with abnormal preoperative neurologic examination findings were more likely to develop complications or die. Dogs undergoing a suboccipital approach were more likely to die. The most common postoperative complications other than death were seizures (n = 18 [11.3%]), worsening of neurologic status (6 [3.8%]), and aspiration pneumonia (6 [3.8%]). CONCLUSIONS AND CLINICAL RELEVANCE Results of the present study provided valuable information on predisposing factors, odds of major complications or death, and incidences of major complications or death in dogs during the first 30 days after undergoing cytoreductive surgery for treatment of suspected primary intracranial masses. Careful case selection may help improve outcomes and minimize complications.

Association of magnetic resonance imaging-based preoperative tumor volume with postsurgical survival time in dogs with primary intracranial glioma.

OBJECTIVE To assess the relationship between preoperative volume of primary intracranial gliomas in dogs as determined via MRI and survival time after surgical debulking and adjunctive immunotherapy. DESIGN Retrospective cohort study. ANIMALS 47 client-owned dogs enrolled in clinical trials regarding glioma treatments. PROCEDURES Medical records of all dogs undergoing craniotomy at the University of Minnesota Veterinary Medicine Center with histologically confirmed glioma between 2008 and 2015 were retrospectively reviewed, and outcome data were collected. Preoperative T2-weighted or post-gadolinium administration T1-weighted MRI scans, performed at several referral institutions with scanners of magnet strengths ranging from 0.5 to 3.0 T, were used to measure tumor volumes as a percentage of total calvarial volume. Data were analyzed to assess the effect of each 2% fraction of tumor volume on median survival time (MST) after surgery and adjuvant treatment. RESULTS Tumor volumes ranged from 0.5% to 12.2% of total intracranial volume. Overall MST was 185 days (range, 2 to 802 days). No association was identified between preoperative tumor volume and MST. Only 3 (6%) dogs had low-grade tumors that had relatively small volumes, measuring 1.4%, 2.1%, and 4.3% of total calvarial volume. The MST for these 3 dogs (727 days) was longer than that for high-grade tumors (174 days); however, owing to the low number of dogs with low-grade tumors, no statistical comparison was performed. CONCLUSIONS AND CLINICAL RELEVANCE Preoperative tumor volume determined via MRI was neither associated with nor predictive of outcome following surgery and adjunctive treatment for dogs with glioma. Tumor grade was predictive of outcome, but unlike tumor volume that was measured with MRI, invasive biopsy was necessary to definitively diagnose tumor grade.

Genetically engineered minipigs model the major clinical features of human neurofibromatosis type 1.

Neurofibromatosis Type 1 (NF1) is a genetic disease caused by mutations in Neurofibromin 1 (NF1). NF1 patients present with a variety of clinical manifestations and are predisposed to cancer development. Many NF1 animal models have been developed, yet none display the spectrum of disease seen in patients and the translational impact of these models has been limited. We describe a minipig model that exhibits clinical hallmarks of NF1, including café au lait macules, neurofibromas, and optic pathway glioma. Spontaneous loss of heterozygosity is observed in this model, a phenomenon also described in NF1 patients. Oral administration of a mitogen-activated protein kinase/extracellular signal-regulated kinase inhibitor suppresses Ras signaling. To our knowledge, this model provides an unprecedented opportunity to study the complex biology and natural history of NF1 and could prove indispensable for development of imaging methods, biomarkers, and evaluation of safety and efficacy of NF1-targeted therapies.

Optimization of adenoviral vector-mediated transgene expression in the canine brain in vivo, and in canine glioma cells in vitro.

Expression of the immune-stimulatory molecule Fms-like tyrosine kinase 3 ligand (Flt3L) and the conditional cytotoxic enzyme herpes simplex virus type 1 thymidine kinase (HSV1-TK) provides long-term immune-mediated survival of large glioblastoma multiforme (GBM) models in rodents. A limitation for predictive testing of novel antiglioma therapies has been the lack of a glioma model in a large animal. Dogs bearing spontaneous GBM may constitute an attractive large-animal model for GBM, which so far has remained underappreciated. In preparation for a clinical trial in dogs bearing spontaneous GBMs, we tested and optimized adenovirus-mediated transgene expression with negligible toxicity in the dog brain in vivo and in canine J3T glioma cells. Expression of the marker gene beta-galactosidase (beta-Gal) was higher when driven by the murine (m) than the human (h) cytomegalovirus (CMV) promoter in the dog brain in vivo, without enhanced inflammation. In the canine brain, beta-Gal was expressed mostly in astrocytes. beta-Gal activity in J3T cells was also higher with the mCMV than the hCMV promoter driving tetracycline-dependent (TetON) transgene expression within high-capacity adenovirus vectors (HC-Ads). Dog glioma cells were efficiently transduced by HC-Ads expressing mCMV-driven HSV1-TK, which induced 90% reduction in cell viability in the presence of ganciclovir. J3T cells were also effectively transduced with HC-Ads expressing Flt3L under the control of the regulatable TetON promoter system, and as predicted, Flt3L release was stringently inducer dependent. HC-Ads encoding therapeutic transgenes under the control of regulatory sequences driven by the mCMV promoter are excellent vectors for the treatment of spontaneous GBM in dogs, which constitute an ideal preclinical animal model.