Received: 25 October 2020 Accepted: 27 May 2021 DOI: 10.1111/jvim.16199 STANDARD ARTICLE Clinical features, diagnosis, and survival analysis of dogs with glioma Roberto José-Lo pez1,2 | Rodrigo Gutierrez-Quintana1 | Cristian de la Fuente2 | Edgar G. Manzanilla3,4 | Anna Suñol5 | Dolors Pi Castro2,6 | Sonia Añor2 | Daniel Sanchez-Masian7 | Francisco Fernandez-Flores7 | Emanuele Ricci7 | Katia Marioni-Henry8 | Joan Mascort5 | Lara A. Matiasek9 | Kaspar Matiasek10 | Paul M. Brennan11 | Martí Pumarola2,6 1 School of Veterinary Medicine, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, UK 2 Department of Animal Medicine and Surgery, Veterinary Faculty, Universitat Autònoma de Barcelona, Barcelona, Spain 3 School of Veterinary Medicine, University College Dublin, Dublin, Ireland 4 TEAGASC, The Irish Food and Agriculture Authority, Cork, Ireland 5 ARS Veterinaria, Barcelona, Spain 6 Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Universitat Autònoma de Barcelona, Barcelona, Spain 7 Institute of Veterinary Science, University of Liverpool, Neston, UK 8 Royal (Dick) School of Veterinary Studies and Roslin Institute, University of Edinburgh, Edinburgh, UK 9 Tierklinik Haar, Haar, Germany 10 Centre for Clinical Veterinary Medicine, Ludwig-Maximilians-Universitaet, Munich, Germany 11 Translational Neurosurgery, Centre for Clinical Brain Sciences, University of Edinburgh, Edinburgh, UK Correspondence Roberto José-Lo pez, School of Veterinary Abstract Medicine, University of Glasgow, Bearsden Background: Gliomas in dogs remain poorly understood. Road, Glasgow, G61 1QH, UK. Email: roberto.jose-lopez@glasgow.ac.uk Objectives: To characterize the clinicopathologic findings, diagnostic imaging fea- tures and survival of a large sample of dogs with glioma using the Comparative Brain Present address Anna Suñol, Royal (Dick) School of Veterinary Tumor Consortium diagnostic classification. Studies, University of Edinburgh, Animals: Ninety-one dogs with histopathological diagnosis of glioma. Edinburgh, UK Methods: Multicentric retrospective case series. Signalment, clinicopathologic find- Dolors Pi Castro, Anicura Arvivet Hospital Veterinari, Barcelona, Spain ings, diagnostic imaging characteristics, treatment, and outcome were used. Tumors Daniel Sanchez-Masian, Anderson Moores were reclassified according to the new canine glioma diagnostic scheme. Veterinary Specialists, Winchester, UK Results: No associations were found between clinicopathologic findings or survival and Lara A. Matiasek, Anicura Small Animal Clinic, tumor type or grade. However, definitive treatments provided significantly (P = .03) Babenhausen, Germany improved median survival time (84 days; 95% confidence interval [CI], 45-190) Abbreviations: CBTC, Comparative Brain Tumor Consortium; CE, contrast enhancement; CI, confidence interval; CNS, central nervous system; CSF, cerebrospinal fluid; CT, computed tomography; FLAIR, fluid-attenuation inversion recovery; GFAP, glial fibrillary acidic protein; GRE, gradient-recalled echo; HA, high-grade astrocytoma; HO, high-grade oligodendroglioma; HU, high-grade undefined glioma; LA, low-grade astrocytoma; LO, low-grade oligodendroglioma; MRI, magnetic resonance imaging; MST, median survival time; OR, odds ratio; WHO, World Health Organization. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes. © 2021 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals LLC on behalf of American College of Veterinary Internal Medicine. 1902 wileyonlinelibrary.com/journal/jvim J Vet Intern Med. 2021;35:1902–1917.  JOSE-LÓPEZ ET AL. 1903 Funding information University of Glasgow, Small Animal Hospital compared to palliative treatment (26 days; 95% CI, 11-54). On magnetic resonance imag- Fund Postgraduate Research Grant, Grant/ ing (MRI), oligodendrogliomas were associated with smooth margins and T1-weighted Award Number: 145973-02 hypointensity compared to astrocytomas (odds ratio [OR], 42.5; 95% CI, 2.42-744.97; P = .04; OR, 45.5; 95% CI, 5.78-333.33; P < .001, respectively) and undefined gliomas (OR, 84; 95% CI, 3.43-999.99; P = .02; OR, 32.3; 95% CI, 2.51-500.00; P = .008, respec- tively) and were more commonly in contact with the ventricles than astrocytomas (OR, 7.47; 95% CI, 1.03-53.95; P = .049). Tumor spread to neighboring brain structures was associated with high-grade glioma (OR, 6.02; 95% CI, 1.06-34.48; P = .04). Conclusions and Clinical Importance: Dogs with gliomas have poor outcomes, but risk factors identified in survival analysis inform prognosis and the newly identified MRI characteristics could refine diagnosis of tumor type and grade. KEYWORDS astrocytoma, dog, magnetic resonance imaging, oligodendroglioma, prognosis, tumor grade, undefined glioma 1 | I N T RO DU CT I O N criteria. However, little is known about whether histologic tumor type and grade correlates with biologic behavior in canine gliomas. To date, The incidence of brain tumors in adult dogs is 2.8% to 4.5% and, information on tumor progression and outcome after treatment is anec- although individual studies vary, gliomas represent 36% to 70% of pri- dotal.18 Since 2007, advances in molecular genetics and biology have 1-5 mary brain tumors in dogs. Consequently, glioma in dogs is increas- enhanced our understanding and subclassification of human gliomas, ingly recognized as a naturally occurring model for understanding which led in 2016 to an updated edition of the WHO brain tumor classifi- human glioma. The benefits include the size and structure of the cation.19 Subsequently, the Comparative Brain Tumor Consortium (CBTC) canine brain, the incidence of spontaneous gliomas, and the coexis- of the National Cancer Institute proposed a revised diagnostic classifica- tence with an active immune system.6,7 Nevertheless, there are many tion of canine gliomas.8 Their aim was to provide an updated canine- gaps in knowledge related to the natural biology of glioma in dogs as specific scheme for clinical and molecular data to be added into a mor- well as its molecular characteristics. phologic diagnosis, to assist with prediction of tumor behavior. Epidemiologic data on glioma in dogs indicates a median age at The aims of this study were to enhance this revised diagnostic diagnosis of 8 years, a male predilection (incidence ratio of 1.53 for classification by further characterizing the epidemiologic, clinicopath- males/females), and predominant lesion location within the fronto- ologic, diagnostic imaging, and outcome features of gliomas in dogs in olfactory, temporal, and parietal lobes of the brain.3,5,8 Over 50% of a large sample. We assess the relationship between these features 3,5,8-10 all gliomas in dogs occur in certain brachycephalic breeds and and tumor histological type and grade, based on the new diagnostic the Boston Terrier, Bulldog, and Boxer breeds have a higher preva- classification for dogs. lence of oligodendroglioma.8 On magnetic resonance imaging (MRI), intracranial gliomas in dogs are typically described as intra-axial, T1-weighted iso- to hypointense and T2-weighted iso- to 2 | M A T E R I A L S A N D M ET H O D S hyperintense mass lesions with varying degrees of contrast enhance- ment (CE).5,11-14 Oligodendrogliomas are reported to contact the brain This study was approved by the Research Ethics Committee of the surface more commonly, whereas astrocytomas have been associated School of Veterinary Medicine of the University of Glasgow with more peritumoral edema, lack of ventricular distortion, and iso- (Ref33a/17). The clinical records of dogs presented to 7 European to hyperintense T1-weighted signal.13,14 Tumors with mild to no CE, referral centers between 2005 and 2018 were retrospectively ana- absent cystic structures, and tumor location other than the thalamo- lyzed. Dogs were included if they had a histopathologic diagnosis of capsular region have been associated with low-grade gliomas.14 Grade glioma. Samples were obtained by means of surgical biopsy or at nec- and type of histologically confirmed intracranial gliomas in dogs using ropsy within 24 hours from death. these MRI features found an accuracy of 53.3% and 60% for predicted tumor grade and type, respectively.15 Standard veterinary practice in recent years has been to use the 2.1 | Morphologic diagnosis 2007 World Health Organization (WHO) human glioma classification to grade canine gliomas.16,17 This classified and graded human tumors based All samples were fixed in 10% neutral buffered formalin. Fixation times on analysis of clinical outcome and survival relative to specific pathologic varied because of the multicentric and retrospective nature of the study; 1904  JOSE-LÓPEZ ET AL. however, this was always <5 days. After fixation, transverse sections of 2.4 | Magnetic resonance imaging the brain or spinal cord were made and samples including the tumor area were routinely processed. Morphologic evaluation was performed on Magnetic resonance imaging studies were performed with scanners of 4 μm paraffin-embedded sections stained with hematoxylin and eosin. variable field strength (0.2-1.5 T). All studies included T2-weighted All gliomas were reviewed and classified by a board-certified patholo- sagittal and transverse sequences and both, precontrast and post- gist (M. Pumarola) using the CBTC diagnostic scheme.8 When available, contrast (gadopentate dimeglumine; Magnevist, Bayer Schering samples were further evaluated by immunohistochemistry for glial fibrillary Pharma AG, Berlin, Germany) T1-weighted transverse sequences. acidic protein (GFAP) (Z0334; Dako; Glostrup; Denmark; 30-minute incu- When available, additional fluid-attenuation inversion recovery bation at a dilution of 1:500), and Olig2 protein (AB 9610; Merck Mil- (FLAIR) dorsal or transverse sequences, gradient-recalled echo (GRE) lipore; Darmstadt; Germany; 30-minute incubation at a dilution of 1:100). transverse sequences, T2-weighted dorsal views, and precontrast and For both markers, slides were scored on a scale of 0 to 4 as previously postcontrast T1-weighted dorsal and sagittal views were evaluated. described.20 Immunohistochemical characterization of 16 cases (dogs Magnetic resonance imaging features were independently evalu- 10-20, 76-80; Supplementary Table 1) was reported previously.20-22 ated by 2 board-certified neurologists (R. Gutierrez-Quintana, R. José- Histopathology reports and available tissue were reviewed for fea- pez) and classified based on the consensus opinion. Both observers Lo tures of infiltrative spread consisting of invasion of adjacent central ner- were aware that lesions were gliomas but were blinded to the histo- vous system (CNS) regions as well as presence of secondary structures of pathologic type and grade and provided with standardized grading Scherer, including perineuronal and perivascular satellitosis, subpial and instructions. The MRIs were classified on the basis of 20 criteria subependymal tumor cell condensation, and invasion along white matter adapted from a recent study.14 For each criterion, observers chose 8,17,19,23 tracts. Extension through the corpus callosum into the contralat- 1 option from those specified in Table 2. Spinal cord located gliomas eral hemisphere was documented as butterfly glioma.19,24 Gliomatosis cer- were excluded from the analysis. ebri growth pattern was recorded as recently defined.19,25 Observers were provided with specific instructions for some MRI Evidence of extension into the subarachnoid space (tumor spread criteria. If the tumor margins were clear, they were divided into through the pia mater with or without proliferation within the lep- smooth and irregular whereas if they were indistinct, they were con- tomeninges), ventricular invasion or drop metastases was noted.26-29 sidered poorly defined. Each glioma was classified on the basis of sig- Penetration of the bone21 or infiltration of other non-CNS structures nal intensity relative to cortical gray matter and signal uniformity on were also recorded. Finally, postmortem examination reports were T1-weighted, T2-weighted, and FLAIR images. Interpretation was 5 reviewed for the presence of metastases elsewhere in the body. based on the majority of the tumor area. Similarly, on T1-weighted postcontrast images, degree of CE was evaluated and classified based on the pattern of the largest portion of the tumor. Peritumoral edema 2.2 | Anatomic location of tumors was graded and cystic structures noted according to a previous study.31 Subarachnoid CSF signal loss, midline shift, ventricular distor- Gliomas were allocated into 1 of 4 main anatomic locations: hemi- tion, brain herniations, and syringohydromyelia were all categorized spheric (cerebral hemispheres including deep gray matter), diencepha- lon, infratentorial, and spinal cord. Within these regions, the specific location of the largest portion of the tumor was recorded as fronto- TABLE 1 Summary of clinical signs annotation olfactory, parietal lobe (including the adjacent corpus callosum), Clinical Descriptions used in this study (total number of temporal lobe (including the piriform lobe), occipital lobe, ventricles, finding cases for each criterion) diencephalon, cerebellum, brainstem, and spinal cord segment Seizures None (35); isolated (32); cluster seizures (24) corresponding with the overlying vertebrae.8,14,30 Furthermore, any Mentation Normal (28); lethargy/disorientation (14); degree of involvement of the diencephalon was noted.5,13 depression (48) Behavior Normal (44); behavioral abnormalities (46) Posture Normal (69); head tilt (11); head turn (2); low-head 2.3 | Dog demographics, clinicopathologic data, carriage (6); kyphosis (1) and staging Gait Normal (37); ataxia, paresis, or both (53) Proprioception Normal (23); deficits (67) Demographic data (age, sex, and breed), presenting clinical signs, and Vision Normal (41); unilateral deficits (34); bilateral deficits physical and neurologic examination findings were recorded for each (15) case. Clinical signs were annotated as summarized on Table 1. Facial/nasal Normal (81); deficits (9) Cerebrospinal fluid (CSF) analysis results, when available before sensation treatment, were noted. Brainstem Yes (facial asymmetry (3); abnormal eye Reports of cases that underwent diagnostic imaging of the thorax, signs movements/position (10); anisocoria (3); pupillary light reflex deficits (3)); no (75) abdomen, or both for staging were reviewed for the presence of Hyperesthesia Yes (14); no (76) metastases.5  JOSE-LÓPEZ ET AL. 1905 TABLE 2 Standardized MRI interpretation criteria for intracranial gliomas included in this study (n = 74) MRI criteria Descriptions used in this study (total number of cases for each criterion) Origin Intra-axial (72); extra-axial (2) Margins Poorly defined (25); smooth (44); irregular (5) Shape Spherical or ovoid/elongate (47); amorphous (24); lobulated (3) Signal T2-intensity Hypointense (3); isointense (0); hyperintense (71) T2-uniformity Homogeneous (22); heterogeneous (52) T1-intensity Hypointense (60); isointense (14); hyperintense (0) T1-uniformity Homogeneous (15); heterogeneous (59) FLAIR intensity Hypointense (11); isointense (0); hyperintense (63) FLAIR uniformity Homogeneous (16); heterogeneous (58) a GRE signal voids None (25); single (6); multiple (6); diffuse (majority of tumor) (4) Degree of CE None (19); mild (20); moderate (19); severe (16) CE pattern None (19); focal (1); nonuniform (28); uniform (entire tumor) (8); partial ring (7); complete ring (11) Cystic structures None (44); cyst (16); ITFs (14) Peritumoral edema None (7); peritumoral (≤10 mm beyond tumor margins) (47); extensive (>10 mm beyond tumor margins) (20) Mass effect None (3); mild (16); moderate (29); severe (26) Subarachnoid CSF signal loss Yes (63); no (11) Midline shift Yes (54); no (20) Ventricular distortion Yes (68); no (6) Brain herniationsb None (31); subfalcine (2); transtentorial (24); foramen magnum (19) Syringohydromyelia Yes (24); no (50) Spread Adjacent brain structures None (22); butterfly glioma (3); gliomatosis cerebri growth pattern (5); freehand description (44) Brain surface contact Yes (56); no (18) Leptomeningeal CE Yes (25); no (49) Ventricular contact Yes (62); no (12) b CSF pathways None (26); subarachnoid space (25); ventricular invasion (33); drop metastases (8) Other structuresb None (60); penetration of bone (9); other (freehand description) (6) Abbreviations: CE, contrast enhancement; CSF, cerebrospinal fluid; FLAIR, fluid-attenuation inversion recovery; GRE, gradient-recalled echo; ITFs, intratumoral accumulations of fluid; MRI, magnetic resonance imaging; T1, T1-weighted; T2, T2-weighted. a GRE images were only obtained in 41 gliomas. b More than 1 criterion could be observed for a single case. independently and subsequently considered for mass effect Finally, based on the observations of published MRI predictors grading.14,32 of canine glioma type and grade (summarized in Supplementary Additionally, brain MRIs were evaluated for features of tumor Table 2), investigators predicted the grade (high or low) and type spread, categorized as specified in Table 2. For spread from the ana- (astrocytoma or oligodendroglioma) for each tumor.13-15 Accuracy tomic location containing the largest fraction of the tumor to neighbor- for predicting the lesion grade and type on MRI compared to histo- ing brain structures or distant foci, observers were instructed to provide pathologic diagnosis was performed using the consensus opinion of a freehand description of the anatomic region/s tumors were extending both observers. to. Tumor growth patterns consistent with butterfly glioma or gliomatosis cerebri were recorded.19,24,25 Leptomeningeal CE with asso- ciated FLAIR hyperintensity within the sulci was recorded as well.25,26 2.5 | Treatment and outcome When present, this was also annotated as extension into the subarach- noid space as was tumor invasion of the subarachnoid space without Treatment groups were defined as definitive if specific treatment associated meningeal MRI changes. Spread along the CSF pathways modalities including surgical resection, radiotherapy, or chemotherapy including ventricular invasion and drop metastases was noted too.26-29 were used in any combination, and palliative if corticosteroid, anti- Similarly, propagation to non-CNS structures was recorded. epileptic, or analgesic medications were the only therapeutic 1906  JOSE-LÓPEZ ET AL. interventions.33 Survival was defined for all cases from time of MRI Immunohistochemistry results for GFAP were available for diagnosis, and only for cases with survival times >1 day to exclude 60 cases with a mean immunoreactivity score of 0.32/4 (range, 0-2/4) animals euthanized at the time of diagnosis.33 on 35 oligodendrogliomas, 1.71/4 (range, 0-4/4) on 14 astrocytomas, and 1.55/4 (range, 0-4/4) on 11 high-grade undefined gliomas (HUs). Olig2 immunoreactivity results were available for 57 cases with mean 2.6 | Statistical analyses scores of 2.67/4 (range, 1-4/4) on 36 oligodendrogliomas, 1.85/4 (range, 0-4/4) on 13 astrocytomas, and 2/4 (range, 1-3/4) on 8 HUs. All analyses were carried out using SAS 9.4 (SAS Institute Inc, Information on histopathologically confirmed infiltrative spread Cary, North Carolina) and R 4.0.2 (R Foundation for Statistical was available for 75 cases. Invasion of neighboring brain structures Computing, Vienna, Austria). When not specified, alpha level for was observed in 51/75 cases including 35 oligodendrogliomas determination of significance was .05 and trends were discussed (32 high-grade and 3 low-grade), 10 high-grade astrocytomas (HAs), for alpha .1. and 6 HUs. Secondary structures of Scherer were noted in 28/75 Interobserver agreement for each MRI criterion was assessed cases: 19 oligodendrogliomas (18 high-grade and 1 low-grade), 6 HAs, using Cohen's kappa. Values of 0.81-1.00 were considered to indicate and 3 HUs. These consisted of perivascular satellitosis and subpial excellent agreement; 0.61-0.80, good agreement; 0.41-0.60, moder- tumor cell condensation (9 cases, respectively), and perineuronal ate agreement; 0.21-0.40, fair agreement; 0.01-0.20, poor agreement, satellitosis, subependymal tumor cell condensation, and white matter and 0.00, chance agreement. tract invasion (7 cases, respectively) in any combination. Three high- Agreement between histopathologic diagnosis (reference test) of grade oligodendrogliomas (HOs) presented as butterfly gliomas type and grade and classification of glioma type and grade following (Figure 1A-C) whereas gliomatosis cerebri growth pattern was seen in previously published MRI criteria13,14 was analyzed using specificity 3 HAs and 1 HU. and sensitivity for detection of astrocytoma for type and high for Ventricular invasion was confirmed in 24/75 cases (18 HOs, grade. 4 HAs, 1 HU, and the only low-grade astrocytoma [LA]). Subarachnoid Associations between tumor location, patient demographics, clini- spread with or without proliferation within the leptomeninges was cal signs, MRI criteria, and tumor type and grade were performed in noted in 22/75 gliomas (16 HOs, 4 HAs, and 2 HUs). Drop metastases 2 stages, first univariate analysis, and then multivariable analysis. Uni- were found in 9 HOs and 1 HA. Additionally, 1 HA presented 2 inde- variate analysis was done using contingency tables and Fisher's exact pendent foci (Figure 1D-G). test. Variables with P value <.25 were used for the multivariable Information on spread to non-CNS structures and extraneural model. For multivariable analysis, forward stepwise logistic regression metastases was available in 50 cases. Penetration of the cribriform was used. A P-value to enter the model of <.20 and to remain of <.05 plate and infiltration of the ethmoturbinates and nasal mucosa was was used. Final logistic regression model fit was evaluated using the confirmed in 3 HOs whereas osteolytic calvarial invasion was Hosmer-Lemeshow goodness-of-fit test. noted on a previously reported HA.21 Pituitary gland infiltration Associations between all risk factors and hazard of death was per- (Figure 1H-J) was found in 1 of each of HA, HO, and low-grade formed for dogs with survival of >1 day and available MRI for revision oligodendroglioma (LO). No metastases were found elsewhere in using univariable followed by multivariable Cox proportional hazard the body. modeling. All risk factors with a P value <.25 in the univariate analysis were used in the multivariable analysis. A forward stepwise manual approach was used to build the multivariable model. Risk factors were 3.2 | Location of gliomas within the CNS kept in the final model after assessing for confounding with a P value of .05. The proportional hazards assumption was tested by visual There were 69/91 (76%) hemispheric gliomas, 14/91 (15%) dience- inspection of log-minus-log survival plots and statistical assessment of phalic, 5/91 (6%) infratentorial, and 3/91 (3%) in the spinal cord. residuals. Geographic distribution within the brain of type and grade of glioma is summarized in Figure 2A. Within the cerebral hemispheres (Figure 2B), 29/69 tumors (42%) had a fronto-olfactory location, 3 | RESULTS 25/69 (36%) were in the temporal lobe, 11/69 (16%) in the parietal lobe, and 4/69 (6%) were primarily intraventricular. All of the latter 3.1 | Morphologic diagnosis were located within the lateral ventricles and consisted of HOs. In addition to the 14 primarily diencephalic gliomas, another 16 gliomas Ninety-one dogs with histopathological diagnosis of glioma were involved the diencephalon to some extent (diencephalic involve- identified. Ten cases were identified by surgical biopsy and 81 cases ment by tumor type and grade is presented in Table 3). Infratentorial were diagnosed at necropsy. Of these, 68.1% were classified as gliomas included 4 HAs (brainstem, 3; cerebellum, 1) and 1 HO oligodendroglioma (52 high-grade and 10 low-grade), 18.7% as astro- (brainstem). Spinal cord gliomas consisted of 2 HOs located at the cytoma (16 high-grade and 1 low-grade), and 13.2% as undefined level of C1 and T11 vertebrae, respectively, and 1 LO extending glioma (12, all high-grade). from C7 to T5 vertebral bodies.  JOSE-LÓPEZ ET AL. 1907 F I G U R E 1 Transverse (A) and dorsal (B) T1-weighted postcontrast magnetic resonance (MR) images of a ring-enhancing high-grade oligodendroglioma (HO) showing the typical features of butterfly glioma with extensive involvement of the corpus callosum leading to bihemispheric spread. Photomicrograph of the same HO as in (A) and (B) occupying the lumen of the lateral ventricle and infiltrating the cingulate gyrus (top of the figure) and the corpus callosum (bottom of the figure) (C). The inset shows neoplastic cells invading the corpus callosum towards the contralateral ventricle (the oval represents an area of higher neoplastic cell density). HE stain. Scale bar = 1 mm (inset 200 μm). Transverse fluid-attenuation inversion recovery MR image at the level of the tentorium cerebelli demonstrating 2 heterogeneously hyperintense independent foci of a high-grade astrocytoma (D). Dorsal T2-weighted images obtained at the level of the red (E) and the yellow (F) dotted lines show the largest tumor in the cerebellum and a second focus in the right occipital lobe, respectively. Both foci were characterized by large sized anisokaryotic cell populations with scant cytoplasm growing in a solid pattern (G). A mitotic figure is present (arrow) as well as glomeruloid-like vessels (arrowheads). HE stain. Scale bar = 50 μm. Midsagittal T2-weighted images of a poorly defined, heterogeneously hyperintense HO extending from the fronto-olfactory area to the diencephalon (H). Note the enlarged and hyperintense pituitary gland (arrow). Severe nonuniform enhancement of the pituitary gland was noted on T1-weighted postcontrast images whereas this was mild for the intra-axial tumor. Dorsal view of the unfixed base of the neurocranium in the same dog as (H) demonstrating an expanded pituitary gland (asterisk) (I). Photomicrograph of the pituitary gland of the dog in (H) and (I) revealing severe infiltration of the hypophyseal lobules by the neoplastic cell population (J). HE stain. Scale bar = 500 μm 3.3 | Demographics, clinicopathologic features, castrated males, 21/91 (23%) intact females, and 19/91 (21%) spayed and staging females. The ratio for all males/females was 1.28, and 1.9 for intact males/females. Seventy-eight percent of the cases (71/91) belonged Median age of dogs at diagnosis of glioma was 7.9 years (range, to brachycephalic dog breeds including 40/91 (44%) Boxers, 26/91 1.5-13.0 years). There were 40/91 (44%) intact males, 11/91 (12%) (29%) Bulldogs (French, 23; English, 1; and not otherwise specified, 2), 1908  JOSE-LÓPEZ ET AL. F I G U R E 2 Anatomic distribution of intracranial gliomas in 88 dogs classified according to the Comparative Brain Tumor Consortium diagnostic scheme (A). Specific location of 65 hemispheric gliomas (B). Note there were an additional 4 high-grade oligodendrogliomas primarily located within the lateral ventricles 3/91 (3%) Dogues de Bordeaux, and 2/91 (2%) Staffordshire Bull Ter- Magnetic resonance imaging features of spinal cord gliomas con- riers. The breeds of the remaining 22% of dogs are captured on Sup- sisted of T2-weighted hyperintense, T1-weighted isointense focal, or plementary Table 1. diffuse mass lesions with none to severe CE located intramedullary Time between onset of clinical signs and presentation ranged except for the HO at C1 that appeared intradural extramedullary between <1 and 180 days (median 14 days) for all gliomas. The main (Figures 3A-C). presenting complaint in 62% (56/91) of the cases was seizures either The MRI features of intracranial gliomas are captured in Table 2. isolated (32 dogs) or in clusters (24). Of these, 5/56 (9%) presented All intracranial gliomas in this study were intra-axial except for a HA with a normal interictal neurologic examination and seizures as the (Figures 3D-F) and a previously reported case of leptomeningeal only clinical sign. In addition to seizures, the most common presenting oligodendrogliomatosis reclassified as HO.22 Spread to adjacent brain neurologic signs included proprioceptive deficits (67/90 cases, 74%), structures was detected in 52/74 (70%) gliomas and further charac- mentation changes (62/90 cases, 69%), gait abnormalities (53/90 terized as butterfly glioma in 3 HOs and as gliomatosis cerebri growth cases, 59%), visual deficits (49/90 cases, 54%), and behavioral changes pattern in 4 HAs and 1 HU. Extension outside the CNS including pen- (46/90 cases, 51%). The remaining clinical signs are outlined in etration into the bone and invasion of the pituitary gland was Table 1 and most common neurologic signs by tumor type and grade suspected in 9/74 and 6/74 gliomas, respectively. The MRI findings are presented in Table 3. by tumor type and grade are presented in Table 5. Cerebrospinal fluid was collected in 23 dogs. The results are sum- Supplementary Figure 1 shows the interobserver agreement for marized in Table 4. These were within reference limits in 31.6% all the MRI variables studied. Kappa values were excellent or good in (6/19), consistent with albuminocytologic dissociation in 36.8% (7/19) most cases except for predicted tumor grade and spread to bone and with an elevated total nucleated cell count in 34.8% (8/23) cases. structures that showed moderate agreement, and poor for predicted Mixed cell pleocytosis was the most common cytologic abnormality. tumor type. No neoplastic cells were detected. Thoracic radiographs (40 cases) Evaluation of previously published criteria13,14 for prediction of and CT (8), as well as abdominal ultrasonography (40 cases) or CT (3), intracranial glioma type and grade demonstrated a sensitivity failed to reveal abnormalities suggestive of metastatic disease. of 58.8% (95% confidence interval [CI], 35.4%-82.2%) and specificity of 68.8% (95% CI, 55.6%-81.8%) for the diagnosis of astrocytoma, and a sensitivity of 67.2% (95% CI, 55.2%-79.3%) and specificity 3.4 | Magnetic resonance imaging 57.1% (95% CI, 20.5%-93.8%) for the diagnosis of high-grade. Magnetic resonance imaging was obtained for diagnosis in 87 dogs. The remaining 4 dogs were euthanized on presentation at their owners 3.5 | Associations among tumor location, request in view of the clinical suspicion and severity of clinical signs. demographics, clinical signs, MRI features, and glioma The MRI studies of 77 cases were available for review, including 74 brain type and grade and 3 spinal cord gliomas. Median time between MRI and postmortem examination was 1 day (range, <1-1104 days). All brain studies included The preliminary univariate analysis (Tables 3 and 5) showed associa- FLAIR sequences whereas GRE images were attained on 41. tions between glioma type and presence of facial or nasal sensation  JOSE-LÓPEZ ET AL. 1909 T A B L E 3 Univariate analysis of tumor location, patient demographics, and clinical features of 91 gliomas based on type and grade. Table shows percentage of each criterion within each type or grade (total number of cases for each criterion) Type Grade Astrocytoma Oligodendroglioma Undefined High Low (n = 17) (n = 62) (n = 12) P value (n = 80) (n = 11) P value Location Diencephalon 11.8 (2) 14.5 (9) 25.0 (3) .07 13.8 (11) 27.3 (3) .28 Hemispheric 64.7 (11) 79.0 (49) 75.0 (9) 77.5 (62) 63.6 (7) Infratentorial 23.5 (4) 1.6 (1) 0 (0) 6.3 (5) 0 (0) Spinal cord 0 (0) 4.9 (3) 0 (0) 2.5 (2) 9.1 (1) Diencephalic involvement 29.4 (5) 33.9 (21) 33.3 (4) .99 33.8 (27) 27.3 (3) .99 Demographics Age (>96 months) 53.0 (9) 46.8 (29) 58.3 (7) .73 50.0 (37) 45.5 (4) .78 Sex F 11.8 (2) 25.8 (16) 25.0 (3) .86 26.3 (21) 0 (0) .06 FN 29.4 (5) 19.4 (12) 16.7 (2) 20.0 (16) 27.3 (3) M 41.2 (7) 43.6 (27) 50.0 (6) 40.0 (32) 72.7 (8) MN 17.7 (3) 11.3 (7) 8.33 (1) 13.8 (11) 0 (0) Boxer breed 41.2 (7) 46.8 (29) 33.3 (4) .66 43.8 (35) 45.5 (5) .99 Bulldog breed 35.3 (6) 25.8 (16) 33.3 (4) .68 31.3 (25) 9.1 (1) .17 Boxer's phylogenetic clade34 82.4 (14) 79.0 (49) 66.7 (8) .64 78.7 (63) 72.7 (8) .7 Clinical features Duration of signs (>3 days) 86.7 (15) 81.4 (50) 81.8 (10) .99 79.7 (64) 100.0 (11) .2 Seizures None 41.2 (7) 37.1 (23) 41.7 (5) .87 38.8 (31) 36.4 (4) .99 Isolated 29.4 (5) 38.7 (24) 25.0 (3) 35.0 (28) 36.4 (4) Cluster seizures 29.4 (5) 24.2 (15) 33.3 (4) 26.3 (21) 27.3 (3) Mentation Normal 11.8 (2) 36.1 (22) 33.3 (4) .21 27.9 (22) 54.6 (6) .08 Lethargy/disorientation 11.8 (2) 14.8 (9) 25.0 (3) 15.2 (12) 18.2 (2) Depression 76.5 (13) 49.2 (30) 41.7 (5) 57.0 (45) 27.3 (3) Behavioral abnormalities 58.8 (10) 47.5 (29) 58.3 (7) .62 51.9 (41) 45.5 (5) .76 Kyphosis/low-head carriage 5.9 (1) 8.2 (5) 8.3 (1) .99 7.6 (6) 9.1 (1) .99 Head tilt or turn 29.4 (5) 11.5 (7) 8.3 (1) .21 16.5 (13) 0 (0) .36 Gait abnormalities 47.1 (8) 62.3 (39) 50.0 (6) .42 58.2 (47) 54.6 (6) .99 Proprioceptive deficits 70.6 (12) 70.5 (44) 91.7 (11) .36 74.7 (60) 63.6 (7) .48 Vision Normal 64.7 (11) 44.3 (27) 25.0 (3) .29 43.0 (34) 63.6 (7) .27 Unilateral deficits 29.4 (5) 37.7 (23) 50.0 (6) 38.0 (30) 36.4 (4) Bilateral deficits 5.9 (1) 18.0 (11) 25.0 (3) 19.0 (15) 0 (0) Facial/nasal sensation deficits 11.8 (2) 4.9 (3) 33.3 (4) .01 10.1 (8) 9.1 (1) .99 Facial asymmetry 0 (0) 4.9 (3) 0 (0) .99 3.8 (3) 0 (0) .99 Abnormal eye movements/position 11.8 (2) 13.1 (8) 0 (0) .52 11.4 (9) 9.1 (1) .99 Anisocoria/PLR deficits 11.8 (2) 6.6 (4) 0 (0) .55 7.6 (6) 0 (0) .99 Hyperesthesia 17.7 (3) 14.8 (9) 16.7 (2) .91 17.7 (14) 0 (0) .2 Abbreviations: F, female; FN, female neutered; M, male; MN, male neutered; PLR, pupillary light reflex. 1910  JOSE-LÓPEZ ET AL. TABLE 4 CSF analysis results by tumor type and grade in dogs with glioma5 Tumor Mean TP type and Elevated TP Elevated Normal concentration and Mean TNCC and gradea concentrationb TNCCc ACDd CSFa range (mg/dL) range (cells/μL) Differential cytology LO (3) 1 (2) 1 (3) 1 (2) 1 (2) 23.3 (20-26.5) 28 (1-81) Eosinophilic pleocytosis, 1e HO (13) 8 (11) 3 (13) 6 (11) 3 (11) 143.0 (10.1-780) 38 (0-460) Increased percentage of neutrophils (62%), 1f; Mixed cell pleocytosis, 3g HA (4) 3 (4) 3 (4) 0 (4) 1 (4) 206.1 (22.5-736) 15 (0-27) Mixed cell pleocytosis, 3 HU (3) 1 (2) 1 (3) 0 (2) 1 (2) 75.1 (20-130.2) 10 (0-27) Neutrophilic pleocytosis, 1h Abbreviations: ACD, albuminocytologic dissociation; CSF, cerebrospinal fluid; HA, high-grade astrocytoma; HO, high-grade oligodendroglioma; HU, high- grade undefined glioma; LO, low-grade oligodendroglioma; TNCC, total nucleated cell count; TP, total protein. a Number of cases in which CSF information was available. b Number of cases with >25 mg/dL TP concentration (number of cases with available information). c Number of cases with >5 cells/μL (number of cases with available information). d Number of cases with >25 mg/dL TP concentration and <5 cells/μL (number of cases with available information). e Defined as CSF with >50% eosinophils. f Defined as CSF >2% nondegenerated neutrophils.35 g Defined as CSF with a mixture of mostly lymphocytes and large mononuclear cells and >20% contribution of neutrophils and, occasionally, eosinophils.35 h Defined as CSF with >75% neutrophils. F I G U R E 3 Midsagittal (A) and transverse (B) T2-weighted magnetic resonance (MR) images of a heterogeneously hyperintense high-grade oligodendroglioma at the level of C1 vertebra (arrow) identified as intradural-extramedullary by both observers. Note the mass extension into the foramen magnum (arrowhead). Photomicrograph of the same tumor as (A) and (B) demonstrating a highly cellular proliferation in direct contact with the thickened pia mater (arrow) in the dorsal aspect of the spinal cord (C). Note the severe hemorrhages ventrally where the mass is compressing the spinal cord parenchyma (asterisk). HE stain. Scale bar = 500 μm. Transverse T1-weighted postcontrast MR image of an extra- axial nonuniformly contrast-enhancing high-grade astrocytoma ventral to the mesencephalon (arrow) (D). Formalin-fixed transverse section of the brain at the level of (D) demonstrating the extra-axial appearance of the tumor (arrow) (E). Photomicrograph of the paraffin-embedded tumor in (D) and (E) showing a highly cellular proliferation spreading through the subarachnoid space ventral to the mesencephalon (asterisk) with multiple foci of hemorrhage (arrowheads) and necrosis (arrow) (F). HE stain. Scale bar = 500 μm  JOSE-LÓPEZ ET AL. 1911 T A B L E 5 Univariate analysis of MRI criteria of 74 intracranial gliomas based on type and grade. Table shows percentage of each criterion within each type or grade (total number of cases for each criterion) Type Grade Astrocytoma Oligodendroglioma Undefined (n = 17) (n = 49) (n = 8) P value High (n = 67) Low (n = 7) P value Margins Smooth 47.1 (8) 69.4 (34) 25.0 (2) .02 56.7 (38) 85.7 (6) .53 Irregular 11.8 (2) 2.0 (1) 25.0 (2) 7.5 (5) 0 (0) Poorly defined 41.2 (7) 28.6 (14) 50.0 (4) 35.8 (24) 14.3 (1) Shape Amorphous 29.4 (5) 28.6 (14) 62.5 (5) .09 34.3 (23) 14.3 (1) .24 Lobulated 5.9 (1) 2.0 (1) 12.5 (1) 3.0 (2) 14.3 (1) Spherical or ovoid/elongate 64.7 (11) 69.4 (34) 25.0 (2) 62.7 (42) 71.4 (5) Signal T2-hypointensitya 11.8 (2) 2.0 (1) 0 (0) .26 4.5 (3) 0 (0) .99 T2-homogeneity 29.4 (5) 32.7 (16) 12.5 (1) .63 28.4 (19) 42.9 (3) .42 T1-hypointensityb 47.1 (8) 95.9 (47) 62.5 (5) <.001 80.6 (54) 85.7 (6) .99 T1-homogeneity 29.4 (5) 14.3 (7) 37.5 (3) .15 19.4 (13) 28.6 (2) .62 FLAIR hypointensityc 11.8 (2) 16.3 (8) 12.5 (1) .99 14.9 (10) 14.3 (1) .99 FLAIR homogeneity 29.4 (5) 16.3 (8) 37.5 (3) .23 22.4 (15) 14.3 (1) .99 GRE signal voidsd 50.0 (5) 37.0 (10) 25.0 (1) .7 41.0 (16) 0 (0) .51 Contrast enhancement Moderate to severee 58.8 (10) 44.9 (22) 37.5 (3) .56 49.3 (33) 28.6 (2) .43 CE pattern No CE 17.7 (3) 27.5 (14) 25.0 (2) .42 22.1 (15) 50.0 (4) .37 Partial or complete ring 23.5 (4) 27.5 (14) 0 (0) 25.0 (17) 12.5 (1) Other patterns 58.8 (10) 42.9 (21) 75.0 (6) 50.7 (34) 37.5 (3) Tumor characteristics Cystic structures 41.2 (7) 42.9 (21) 25.0 (2) .72 40.3 (27) 42.9 (3) .99 Peritumoral edema None 5.9 (1) 12.2 (6) 0 (0) .28 9.0 (6) 14.3 (1) .84 Peritumoral 47.1 (8) 67.4 (33) 75.0 (6) 64.2 (43) 57.1 (4) Extensive 47.1 (8) 20.4 (10) 25.0 (2) 26.9 (18) 28.6 (2) Mass effect None 11.8 (2) 2.0 (1) 0 (0) .11 4.5 (3) 0 (0) .11 Mild 17.7 (3) 26.5 (13) 0 (0) 20.9 (14) 28.6 (2) Moderate 23.5 (4) 44.9 (22) 37.5 (3) 35.8 (24) 71.4 (5) Severe 47.1 (8) 26.5 (13) 62.5 (5) 38.8 (26) 0 (0) Subarachnoid CSF 88.2 (15) 81.6 (40) 100.0 (8) .62 85.1 (57) 85.7 (6) .99 signal loss Midline shift 64.7 (11) 73.5 (36) 87.5 (7) .52 71.6 (48) 85.7 (6) .66 Ventricular distortion 88.2 (15) 91.8 (45) 100.0 (8) .82 91.0 (61) 100.0 (7) .99 Brain herniations None 35.3 (6) 44.9 (22) 37.5 (3) .09 41.8 (28) 42.8 (3) .21 Transtentorial or subfalcine 41.2 (7) 34.7 (17) 0 (0) 29.9 (20) 57.1 (4) Foramen magnum 23.5 (4) 20.4 (10) 62.5 (5) 28.4 (19) 0 (0) SHM 35.3 (6) 30.6 (15) 37.5 (3) .8 35.8 (24) 0 (0) .09 Spread Adjacent brain structures 58.8 (10) 69.4 (34) 100.0 (8) .09 74.6 (50) 28.6 (2) .02 (Continues) 1912  JOSE-LÓPEZ ET AL. TABLE 5 (Continued) Type Grade Astrocytoma Oligodendroglioma Undefined (n = 17) (n = 49) (n = 8) P value High (n = 67) Low (n = 7) P value Brain surface contact 76.5 (13) 77.6 (38) 62.5 (5) .65 74.6 (50) 85.7 (6) .99 Leptomeningeal CE 47.1 (8) 28.6 (14) 37.5 (3) .34 35.8 (24) 14.3 (1) .26 Ventricular contact 64.7 (11) 87.8 (43) 100.0 (8) .05 83.6 (56) 85.7 (6) .99 CSF pathways Subarachnoid space 47.1 (8) 28.6 (14) 37.5 (3) .34 35.8 (24) 14.3 (1) .41 Ventricular invasion 23.5 (4) 49.0 (24) 62.5 (5) .09 46.3 (31) 28.6 (2) .45 Drop metastases 0 (0) 16.3 (8) 0 (0) .16 11.9 (8) 0 (0) .99 Other structures Pituitary gland 5.9 (1) 8.2 (4) 12.5 (1) .82 7.5 (5) 14.3 (1) .46 Penetration of bone 17.7 (3) 12.2 (6) 0 (0) .56 13.4 (9) 0 (0) .59 Abbreviations: CE, contrast enhancement; CSF, cerebrospinal fluid; FLAIR, fluid-attenuation inversion recovery; GRE, gradient-recalled echo; MRI, magnetic resonance imaging; SHM, syringohydromyelia; T1, T1-weighted; T2, T2-weighted. a As opposed to T2-weighted hyperintensity seen in 88.2% (15) astrocytomas, 98% (48) oligodendrogliomas, and all undefined gliomas; 95.5% (64) high- grade gliomas and all low-grade gliomas. b As opposed to T1-weighted isointensity seen in 52.9% (9) astrocytomas, 4.1% (2) oligodendrogliomas, and 37.5% (3) undefined gliomas; 19.4% (13) high- grade gliomas and 14.3% (1) low-grade gliomas. c As opposed to FLAIR hyperintensity seen in 88.2% (15) astrocytomas, 83.7% (41) oligodendrogliomas, and 87.5% (7) undefined gliomas; 85.1% (57) high- grade gliomas and 85.7% (6) low-grade gliomas. d Percentages and total number of cases with signal voids out of 41 gliomas where GRE images were obtained. e As opposed to none to mild CE seen in 41.2% (7) astrocytomas, 55.1% (27) oligodendrogliomas, and 62.5% (5) undefined gliomas; 50.7% (34) high-grade gliomas and 71.4% (5) low-grade gliomas. F I G U R E 4 Transverse T2-weighted magnetic resonance (MR) image of a high-grade oligodendroglioma (HO) in the right frontal lobe (arrow) with smooth margins and in close contact with the lateral ventricle (asterisk) (A). Transverse T1-weighted image of a hypointense to gray matter HO in the right temporal lobe (arrow) (B). Dorsal T2-weighted MR image showing a high-grade undefined glioma extending from the frontal (arrow) to the temporal lobe (arrowhead) (C). Involvement of the parietal lobe and hippocampus were also noted and gliomatosis cerebri growth pattern was subsequently confirmed on histopathology deficits (P = .01), tumor margins (P = .02) on MRI, T1-weighted signal (Figure 4A) than astrocytomas (odds ratio [OR], 42.5; 95% CI, 2.42- intensity (P < .001), and ventricular contact (P = .04). 744.97; P = .04) or undefined gliomas (OR, 84; 95% CI, 3.43-999.99; Multivariable logistic regression analysis indicated that P = .02). Oligodendrogliomas were also associated with T1-weighted oligodendrogliomas were more often associated with smooth margins hypointensity (Figure 4B) in comparison with astrocytomas (OR, 45.5;  JOSE-LÓPEZ ET AL. 1913 95% CI, 5.78-333.33; P < .001) or undefined gliomas (OR, 32.3; 95% CI, 2.51-500.00; P = .008). Finally, oligodendrogliomas were more often in contact with the ventricles (Figure 4A) than astrocytomas (OR, 7.47; 95% CI, 1.03-53.95; P = .049). For tumor grade, the univariate analysis (Tables 3 and 5) showed associations with spread to adjacent brain structures (P = .02). The multivariable logistic regression analysis indicated that spread to neighboring brain structures (Figure 4C) was more often associated with high-grade gliomas (OR, 6.02; 95% CI, 1.06-34.48; P = .04). 3.6 | Treatment and outcome Survival time was available for all 87 dogs undergoing MRI for diagno- sis. Forty-two dogs survived <1 day (euthanized at diagnosis) and F I G U R E 5 Kaplan-Meier survival curve for definitively and were excluded from further analysis to remove cases without intent palliatively treated dogs with intracranial glioma that survived >1 day (n = 45). Dogs receiving definitive treatment survived significantly to treat. Of the 45 dogs that survived >1 day, 22 received palliative longer (P = .03) than did dogs with palliative treatment. Survival time treatment (10 HOs, 6 HAs, 3 HUs, and 3 LOs), and 23 received defini- represents the time from diagnosis to death or euthanasia. Shadow tive treatment (11 HOs, 5 HUs, 3 HAs, 3 LOs, and 1 LA). Treatment areas represent 95% confidence intervals modalities and survival data are outlined in Supplementary Table 1. T A B L E 6 Multivariable Cox Clinical or MRI variable P value Hazard ratio 95% confidence interval proportional hazard ratios for the survival analysis of 36 dogs surviving >1 day after Seizures <.001 0.044 0.011-0.173 imaging diagnosis and available MRI Marginsa study for evaluation Irregular .001 31.69 4.23-237.28 Poorly defined .003 7.57 2.04-22.18 Drop metastases <.001 53.15 6.25-452.22 T2-heterogeneity .001 6.93 2.21-21.73 Abbreviations: MRI, magnetic resonance imaging; T2-heterogeneity, T2-weighted heterogeneity. a Reference category: smooth margins. F I G U R E 6 Transverse T2-weighted magnetic resonance (MR) image of a left temporal lobe high-grade oligodendroglioma (arrowheads) with poorly defined margins and heterogeneously hyperintense signal extending to the parietal lobe and invading the adjacent lateral ventricle (A). Transverse T1-weighted postcontrast image at the same level demonstrating moderate non-uniform contrast enhancement of the parietal portion of the tumor (arrow) (B). Transverse T1-weighted postcontrast image of the same dog demonstrating a contrast-enhancing drop metastasis in the right cerebellum (arrowhead), just dorsal to the lateral aperture of the fourth ventricle (C) 1914  JOSE-LÓPEZ ET AL. Among the >1-day survivors, definitive treatments were associated owners.39 Definitive treatment modalities contribute to seizure con- with significantly (P = .03) longer survival (median survival time trol in human patients and this could have influenced the improved [MST], 84 days; 95% CI, 45-190) than palliative treatment (MST, survival of these cases in our study; however, there were no associa- 26 days; 95% CI, 11-54) (Figure 5). tions between seizures at onset and treatment modality. Univariate survival analysis is presented in Supplementary Table 3. Although definitive diagnosis of glioma requires histopathologic The multivariable Cox proportional hazards model included seizures analysis of tumor tissue, biopsy or resection of tumors is not always (hazards ratio [HR] 95% CI, 0.01-0.17) as a lower risk factor, and irreg- possible. Therefore, the ability to predict intracranial glioma type and ular (HR 95% CI, 4.2-237.3) or poorly defined (HR 95% CI, 2.0-22.2) grade based on MRI characteristics has recently been investigated in margins, T2-weighted heterogeneous signal (HR 95% CI, 2.21-21.7), dogs.13-15 However, currently published MRI findings for predicting and drop metastases (HR 95% CI, 6.3-452.2) as risk factors for shorter grade and tumor type have proven inaccurate.15 We further analyzed survival (Table 6; Figure 6). No confounding risk factors or interactions those MRI predictors in our larger sample and confirmed their low were found and no evidence of violation of the proportional hazard sensitivity and specificity. Additionally, the low agreement for assumption was found. predicted tumor type and grade using those features indicated high interobserver variability. These might have been related to the overlap of evaluated MRI features between tumor types and grades in our 4 | DISCUSSION population and the high variability of described predictors within each tumor. Our study provides predictive information for the clinical diagnosis of We assessed 74 brain MRI studies for additional indicators of gli- glioma type and grade as well as their prognostic outcome for oma type and grade and found that oligodendrogliomas were associ- affected dogs. New MRI characteristics for differentiation of glioma ated with smooth margins and T1-weighted hypointensity compared type and grade are identified, and clinical and imaging findings are to astrocytomas and undefined gliomas as well as more commonly in associated with survival. Additionally, we describe previously contact with the ventricles than astrocytomas. Tumor spread to unreported features of these CNS tumors. neighboring brain structures was the only finding associated with Glioma type and grade relative frequency in this study is consis- high-grade diagnosis. These are in contrast with reported associations tent with data from prior literature, indicating that oligodendroglial among peritumoral edema and astrocytomas, brain surface contact and high-grade tumors are the most prevalent in dogs.3,8,36 We found and oligodendrogliomas, and CE and cystic structures with tumor no associations between glioma location and tumor type or grade; grade.13,14 Conversely to previous reports,13,14 ventricular contact 8,13 however, consistent with recent studies, our data suggest that gli- alone, independent of distortion, was more common in omas in dogs, like their human counterparts,30 arise mainly in the oligodendrogliomas than astrocytomas. The only similarity between fronto-olfactory, temporal, and parietal regions of the cerebral hemi- this and prior studies was the significantly low likelihood of spheres. The reported increased likelihood of histologic involvement T1-weighted hypointensity in astrocytomas,14 which was shared by of the diencephalon in astrocytomas in dogs was not observed in our undefined gliomas in this series. Although we used comparable meth- cohort.5,13 All gliomas in dogs reported to be primarily intraventricular odology to a previous report,14 differing results might stem from to date have been HOs.8,26,37 inclusion of undefined gliomas in our series and the potentially magni- Almost 80% of the gliomas included herein occurred in brachyce- fied significance of individual outliers not representative of the gen- phalic dog breeds belonging to the same phylogenetic clade,34 eral population in that study,14 where statistics were performed using 3,5,8-10,13 supporting glioma pronounced predilection for these breeds. 155 MRI observations from 5 investigators in 31 gliomas, instead of However, contrasting other reports,8 the Boxers and Bulldogs in our the consensus opinion used herein. study did not show a higher prevalence of oligodendroglioma, Interobserver agreement for individual MRI features associated although this might have been related to smaller case numbers in our with glioma type and grade in our study was good to excellent; how- sample. No associations were found between sex and tumor type or ever, despite reported 89% sensitivity for MRI diagnosis of neoplastic grade, but the recently reported male sex predilection8 was confirmed brain disease and 93.7% specificity and 84.4% sensitivity for diagnosis 30 by our data and resembles the situation in humans. of glioma in dogs, other differential diagnoses with overlapping imag- The most common signs of neurologic dysfunction in this series ing characteristics still need to be considered.12,40 included proprioceptive deficits, mentation changes, and seizures. Differently from our observations, CE on MRI and in particular, Over 60% of the cases were diagnosed after an inaugural seizure, ring-enhancement, is usually associated with higher grade human glio- resembling previous studies in dogs4,5,38 and mirroring the situation in mas.41-43 However, human LAs and approximately 50% of LOs can 17,39 humans. While no presenting clinical sign was associated with gli- show CE44-47 and its absence does not exclude high-grade gli- oma type or grade, seizures at onset was identified as a favorable oma.44,48,49 On the other hand, human oligodendrogliomas are usually prognostic factor for duration of survival in both low- and high-grade well demarcated and largely T1-hypointense,50,51 and glioblastomas gliomas. Like in human gliomas, new-onset seizures might represent might extend widely to adjacent brain structures,52 resembling our an early warning sign for the presence of a brain tumor as other more observations on canine oligodendrogliomas and high-grade gliomas, subtle signs of neurologic disease in dogs might go unnoticed by their respectively.  JOSE-LÓPEZ ET AL. 1915 Novel features of gliomas in dogs encountered in this study to euthanize rather than natural death, and erroneous or incomplete included histopathologically confirmed butterfly growth pattern in record-keeping. Thus, the authors invite researchers to contribute to 3 HOs and pituitary gland infiltration in different glioma types. the creation of a mutually accessible international multicenter data- Reported human and canine butterfly gliomas to date are HAs; how- base to better enable evidence-based research in this field. ever, our findings indicate that, as in gliomatosis cerebri, this repre- Additional limitations of the current study include the fact that sents a pattern of spread rather than a distinct nosologic entity.19,24,53 necropsy samples might not be representative of the original tumor Extension of glioma to the pituitary gland had not been described in phenotype, which could change with tumor progression or be dogs and has only been confirmed once in humans.54 Other infre- influenced by treatments administered. Also, in cases where surgical quent features reported here include penetration into the bone and biopsies were obtained, these might have failed to reflect the overall multifocal distribution. Multifocality as a result of CSF drop metasta- histologic features of the glioma sampled hampering accurate classifi- 26-28 ses occurs in oligodendrogliomas in dogs ; however, truly multiple, cation. Finally, tumor typing and grading was established by a single independent gliomas have only been described in 1 dog and are a rare pathologist which could represent a potential source of bias in view of presentation in humans.55,56 Additionally, we confirmed CSF seeding the moderate agreement for glioma type and grade observed between in a HA in 1 dog, a dissemination pathway exhibited by less than 2% the multiple pathologists contributing to the CBTC classification.8 53 of human glioblastomas. Finally, extra-axial appearance of astrocy- In conclusion, smooth margins, T1-weighted hypointensity and toma had only been reported in a dog,15 and is an infrequent observa- ventricular contact on MRI diagnosed intracranial gliomas, could allow tion in humans.57,58 differentiation between oligodendrogliomas and other glial tumor Data regarding treatment and survival in dogs with histopatholog- types in dogs. High-grade glioma should be suspected if spread over ically confirmed glioma is scarce and yet to be added to the new clas- more than 1 brain region is observed. Definitive therapies appear to sification. The survival analysis presented here is the most complete improve survival time. New onset seizures are associated with a more to date; however, it has limitations inherent to the low case numbers favorable prognosis, whereas MRI observed irregular or poorly and results should be interpreted accordingly. The multivariable analy- defined tumor margins, T2-weighted heterogeneity and drop metasta- sis of available clinical and MRI findings identified prognostic factors ses are negative prognostic indicators. No associations were found affecting survival in intracranial gliomas in dogs for the first time. In between survival and tumor location or the CBTC morphologic addition to the favorable association between new-onset seizures and classification.8 survival, a series of MRI features were identified as negative prognos- tic indicators. Poorly defined and irregular tumor margins associated ACKNOWLEDG MENT death hazard was over 7 and 31 times as high as that associated with This work was supported by a grant from the University of Glasgow, smooth margins, respectively. Tumor T2-weighted heterogeneous sig- Small Animal Hospital Fund. nal and observation of drop metastases were also associated with shorter survival. These prognostic factors differ from those reported CONFLICT OF INTEREST DECLARATION for human gliomas; nevertheless, prognosis in humans is currently Authors declare no conflict of interest. guided by isocitrate dehydrogenase mutation and 1p/19q status,19 for which parallelisms have not been found in canine gliomas.59-63 Case OF F-LABEL ANTIMI CROBIAL DECLARATION series with palliative treatment outcome information relating to gli- Authors declare no off-label use of antimicrobials. oma type or grade are lacking. Canine studies evaluating experimental therapies in >3 intracranial gliomas with detailed information on their INSTITU TIONAL ANIMAL C AR E AND USE COMMITTEE type and grade have reported MSTs of 119 days (n = 7),64 224 days (IACUC) OR OTHER APPROVAL DECLARATION (n = 17),65 248 days (n = 8), 66 and 257 days (n = 8). 67 However, no Approved by the Research Ethics Committee of the School of Veteri- comparisons were made between differing types or grades. Similarly, nary Medicine of the University of Glasgow (Ref33a/17). a recent study reported a MST of 66 days after surgical re- section alone in 14 gliomas,68 8 of which were reclassified and HUMAN E THICS APPROVAL DECLARATION included in this study. Thus, we decided to further evaluate therapeu- Authors declare human ethics approval was not needed for this study. tic outcome in our larger sample of dogs surviving diagnosis. Despite marked variability in treatment modalities (surgery, radiotherapy, che- OR CID motherapy), MST for definitively treated cases (n = 23) was longer pez Roberto José-Lo https://orcid.org/0000-0002-0661-5562 (84 days) than for palliative treatment cases (n = 22; 26 days), Rodrigo Gutierrez-Quintana https://orcid.org/0000-0002-3570- suggesting that definitive therapies provide a significant survival ben- 2542 efit to dogs with intracranial gliomas, although this benefit appears to Sonia Añor https://orcid.org/0000-0002-1099-7698 be limited. 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