Bone Tumor In Dogs Research Articles

A Comparative Study on Radiosensitivity of Canine Osteosarcoma Cell Lines Subjected to Spatially Fractionated Radiotherapy.

Canine appendicular osteosarcoma (OSCA) is a highly aggressive cancer, constituting 85% of all bone tumors in dogs, predominantly affecting larger breeds and exhibiting a high metastatic rate. This disease also shares many genomic similarities with human osteosarcomas, making it an ideal comparative model for treatment discovery. In this study, we characterized the radiobiological properties of several OSCA cell lines when subjected to spatially fractionated radiation therapy (SFRT) and chemotherapy. Specifically, we focused on lower (peak) doses from SFRT ranging from 1 to 10 Gy. These canine OSCA cell lines serve as useful models for osteosarcoma research that can be utilized to find translational treatments for both canine and human patients. This study reaffirms established clinical wisdom regarding the notoriously radioresistant profile of osteosarcomas but additionally offers compelling evidence supporting SFRT as a promising treatment option that could be used in conjunction with other cytotoxic agents.

Köpek ve kedilerde primer kemik tümörleri: 98 vaka

The aim of this study is to contribute to the current literature by determining the distribution of bone tumors in dogs and cats by breed, age, gender and location. Bone tumors are more common in dogs than cats, and the most common primary bone tumor in both species is osteosarcoma. The biopsy and necropsy reports of the Ankara University, Faculty of Veterinary Medicine, Department of Pathology , were retrospectively studied examining for cases of primary bone tumors in dogs and cats. This study's period encompassed from 2001 through 2020 (20 years). A total of 98 bone tumors were detected 70 in dogs and 28 in cats. Of the cases in dogs, 13 (18.57%) were necropsy, while 57 (81.43%) were biopsy. Sex distribution of bone tumors is in female dogs (n:28, 40%) and in male dogs (n:38, 54.29%). Also, females (n:17, 60.7%) and males (n:11, 39.3%) were observed in cats. in dogs, although 57.14% (n=40) were purebreds and 31.43% (n=22) were mongrel breeds, in cats, 75% of them were mongrel (n=21) and 7.1% (n=2) were purebred. While locations of tumors in dogs were appendicular (65.71%, n=46), axial (30%, n=21), locations of tumors in cats were appendicular (50%, n=14), axial (39.29%, n=11) and both appendicular and axial (10.71%, n:3). While 13 benign (18.57%) and 57 malignant (81.43%) tumors were observed in dogs, 4 benign (14.29%) and 24 malignant (85.71%) tumors were observed in cats. The data were analyzed in the SPSS program and no significant relationship was detected between the data (P>0.05). This study would contribute and conduce the comparative oncology for dogs and cats.

Investigating cell death responses associated with histotripsy ablation of canine osteosarcoma

Background Osteosarcoma (OS) is the most frequently occurring primary bone tumor in dogs and people and innovative treatment options are profoundly needed. Histotripsy is an emerging tumor ablation modality, and it is essential for the clinical translation of histotripsy to gain knowledge about the outcome of nonablated tumor cells that could remain postablation. The objective of this study was to characterize the cell death genetic signature and proliferation response of canine OS cells post a near complete histotripsy ablation (96% ± 1.5) and to evaluate genetic cell death signatures associated with histotripsy ablation and OS in vivo. Methods In the current study, we ablated three canine OS cell lines with a histotripsy dose that resulted in near complete ablation to allow for a viable tumor cell population for downstream analyses. To assess the in vivo cell death genetic signature, we characterized cell death genetic signature in histotripsy-ablated canine OS tumors collected 24-h postablation. Results Differential gene expression changes observed in the 4% viable D17 and D418 cells, and histotripsy-ablated OS tumor samples, but not in Abrams cells, were associated with immunogenic cell death (ICD). The 4% viable OS cells demonstrated significantly reduced proliferation, compared to control OS cells, in vitro. Conclusion Histotripsy ablation of OS cell lines leads to direct and potentially indirect cell death as evident by, reduced proliferation in remaining viable OS cells and cell death genetic signatures suggestive of ICD both in vitro and in vivo.

Overall and relapse-free survival of dogs with appendicular osteosarcoma after limb-sparing bio-implantation treatment

Introduction. Canine osteosarcoma is a most common bone tumor in dogs, usually involving long tubular bones lesion. Limb-sparing treatment method allows to save extremity functionality and sufficiently improve quality of life in comparison with amputation.Aim. To analyze the survival rates and local results after limb-sparing treatment of canine osteosarcoma using bio-implantation – de-immunized allogenic bone populated with recipient stromal mesenchymal cell preparation.Materials and methods. Group of 25 dogs underwent a combined treatment including cisplatinum chemotherapy (60–70 mg/m2 intravenously infusion at 14–21 day intervals, up to 4 cycles) and wide segmental resection of the affected bone with a bioimplant defect replacement. Overall survival, progression-free survival and local recurrence frequency in the group were analyzed.Results. Overall survival median was 321 day, and progression-free median was 222 days. 4 (16,6 %) cases of local relapse were recorded, 2 of which were confirmed by radiography and histological examination, and 2 other by radiography only.Conclusion. Efficiency of the canine appendicular osteosarcoma combined limb-sparing treatment is comparable with amputation practice results (according to scientific literature) while foregoing method is preferable due to higher life quality of patients.

PEG-liposomal doxorubicin as a potential agent for canine metastatic osteosarcoma - in vitro and ex ovo studies.

Appendicular osteosarcoma (OSA) is a highly aggressive and metastatic primary bone tumour in dogs. Standard therapy is amputation and adjuvant chemotherapy (e.g. with doxorubicin). Liposomal drug delivery may augment therapeutic efficacy and reduce negative side effects. Polyethylene glycol (PEG)-liposomal doxorubicin treats human metastatic cancers effectively. The study aimed was to evaluate PEG-liposomal doxorubicin's inhibitory effect on canine metastatic proliferation and migration in vitro. It also aimed to appraise the drug's extravasation inhibition in vivo using the human medicine-proven chick embryo chorioallantoic membrane ex ovo model. The canine D-17 OSA cell line was cultured and inoculated with decreasing concentrations of PEG-liposomal doxorubicin and conventional doxorubicin in a 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) test of cell viability, proliferation and cytotoxicity. Flow cytometry with Annexin V and Draq 7 staining confirmed the MTT test results, indicating dead, early and late apoptotic, and live cells. The inhibitory effect of the two preparations on cancer cell migration was investigated with a wound-healing assay. Culture plates seeded with cells were prepared. The cell monolayer was scratched and images of cells migrating to the scratch were captured at 0 h, 12 h and 24 h. Also, embryos were removed from three-day-incubated fertilised chicken eggs. On the 12th day, labelled D-17 cells were injected into each embryo. Embryos in one group received 100 μL of phosphate-buffered saline as controls, those in another group 30 μg/mL of PEG-liposomal doxorubicin, and those in the last group 6 μg/mL of conventional doxorubicin. The effectiveness of the intravascular administration of the D-17 cells was confirmed under a microscope. PEG-liposomal doxorubicin inhibited the migration of canine OSA cells more effectively than conventional doxorubicin (P ≤ 0.05). The ex ovo model showed that both drugs had similar impacts on canine metastatic OSA. The liposomal form of the drug may be considered a potentially effective compound in canine metastatic OSA; nevertheless, further in vivo studies are essential to confirm this hypothesis.

Evaluating immunological outcomes associated with histotripsy treatment in canine osteosarcoma: a comparative human oncology model

Abstract Osteosarcoma (OS), the most commonly occurring primary bone tumor in dogs and humans, has deficient treatment options and dismal survival outcomes. Spontaneously occurring OS in dogs shares numerous similarities with humans and has greater prevalence, allowing the dog to serve as an informative comparative oncology model. The objective of this study was to investigate the non-thermal focused ultrasound technique histotripsy as an innovative treatment option for OS. Histotripsy has the potential to both ablate the primary tumor and stimulate an antitumor immune response to mitigate metastatic disease, the leading cause of death in OS. To investigate the immunological outcomes associated with histotripsy and OS, we delivered histotripsy to client-owned canine OS patients (n=29) and utilized an in-vitro model for cross species comparisons between human, dog, and mouse. Histotripsy ablation led to a significantly greater expression of activation markers CD80 and CD62L on circulating monocytes, at 1- and 5-days post histotripsy (DPH), respectively. We observed a significantly greater production of TGFβ in circulating monocytes at 3 DPH compared to baseline. On an independent patient basis, we observed a greater population of CD5 +CD4 +and CD5 +CD8 +cells within the treated region of the tumor compared to untreated region and changes in genetic signature also indicated immune activation. Across species in-vitro we observed macrophage activation (CD80, CD62L, and CD86), and upregulation of genes associated with immune cell chemotaxis and pro-inflammatory response. Our results indicate that histotripsy ablation leads to immune activation and has strong potential to target both primary OS and metastatic disease. NIH (1R21 EB030182-01), Focused Ultrasound Foundation (PLL7GBVK and FUSF-RAP-823R1), and American Kennel Club (Canine Health Foundation No. 02773)

Histotripsy Ablation of Spontaneously Occurring Canine Bone Tumors In Vivo.

Osteosarcoma (OS) is a devastating primary bone tumor in dogs and humans with limited non-surgical treatment options. As the first completely non-invasive and non-thermal ablation technique, histotripsy has the potential to significantly improve the standard of care for patients with primary bone tumors. Standard of care treatment for primary appendicular OS involves surgical resection via either limb amputation or limb-salvage surgery for suitable candidates. Biological similarities between canine and human OS make the dog an informative comparative oncology research model to advance treatment options for primary OS. Evaluating histotripsy for ablating spontaneous canine primary OS will build a foundation upon which histotripsy can be translated clinically into a standard of care therapy for canine and human OS. Five dogs with suspected spontaneous OS were treated with a 500 kHz histotripsy system guided by real-time ultrasound image guidance. Spherical ablation volumes within each tumor (1.25-3 cm in diameter) were treated with single cycle histotripsy pulses applied at a pulse repetition frequency of 500 Hz and a dose of 500 pulses/point. Tumor ablation was successfully identified grossly and histologically within the targeted treatment regions of all subjects. Histotripsy treatments were well-tolerated amongst all patients with no significant clinical adverse effects. Conclusion & Significance: Histotripsy safely and effectively ablated the targeted treatment volumes in all subjects, demonstrating its potential to serve as a non-invasive treatment modality for primary bone tumors.

Canine Idiopathic Arteriopathy, Appendicular Bone Infarcts, and Neoplastic Transformation of Bone Infarcts in 108 Dogs (Canis lupus familiaris).

Osteosarcoma (OSA) is the most common primary bone tumor in both dogs and humans. The dog is an important research model for OSA, yet dogs have much higher prevalence of bone tumors than do humans, a disparity that has yet to be explained. Neoplastic transformation of cells within or adjacent to bone infarcts into primary bone tumors has been described in humans but only sparsely characterized in the veterinary literature. In this study, 653 cases of canine bone infarcts were received through a referral veterinary osteopathology service over a 14-y period. We identified an idiopathic disorder affecting the nutrient artery, termed canine idiopathic arteriopathy (CIA), which to our knowledge has no direct counterpart in human medicine. This disorder was documented alongside ischemic necrosis of the medullary cavity in 114 bone infarcts in 108 dogs. We hypothesize that CIA precipitated an ischemic environment, resulting in development of a bone infarct down- stream of the abnormal artery. In 52% (59 of 114) of cases, bone infarcts demonstrated evidence of repair (termed reparative bone infarcts [RBI]), while in 48% (55 of 114) of infarcts, a bone tumor was also present, including pleomorphic sarcoma, OSA, fibrosarcoma, and chondrosarcoma. In some cases, a spectrum of tumors was present. We hypothesize that the ischemic infarct environment provoked bone marrow mesenchymal stem cells (MSCs) to attempt repair of the stroma, and in approximately half of cases, MSCs underwent neoplastic transformation (BINT) to produce tumors. The most common sites of bone infarcts were the distal femur, distal radius, proximal humerus, and distal tibia, coinciding with common sites of canine OSA. The authors propose that CIA leading to bone infarcts and infarct-derived tumors, in combination with possible underdiagnosis of canine bone infarcts and misdiagnosis of some RBI as neoplasia, may contribute to the higher reported proportion of bone tumors in dogs compared with humans.

The mitotic regulator polo-like kinase 1 as a potential therapeutic target for c-Myc-overexpressing canine osteosarcomas.

Osteosarcoma is the most common primary malignant bone tumour in dogs, characterized by a locally aggressive and highly metastatic behaviour. Despite the current standards of care, most dogs succumb to the disease, indicating the need for novel treatment strategies. Polo-like kinase 1 (PLK1) is dysregulated in a variety of human cancer types, including osteosarcoma, and induces c-Myc accumulation. The crosstalk between the two molecules coordinates cell proliferation, differentiation, self-renewal and apoptosis. Therefore, PLK1 has recently emerged as a potential therapeutic target, mainly in tumours overexpressing c-Myc. BI 2536 is a selective PLK1 inhibitor promoting mitotic arrest and apoptosis in a variety of cancer cells. This research aimed at evaluating PLK1 and c-Myc protein expression in 53 appendicular canine osteosarcoma (cOSA) samples and the in vitro effects of BI 2536 on a c-Myc and PLK1-overexpressing cOSA cell line (D17). PLK1 and c-Myc expression in cOSA samples showed no correlation with clinicopathological data. However, c-Myc overexpression was associated with a significantly reduced overall survival (p= .003). Western Blot and RT-qPCR assays revealed that D17 expressed high protein and transcript levels of both PLK1 and MYC. When treated with BI 2536 (range 2.5-15 nM) for 24 h, D17 showed a substantial decrease in cell growth, inducing apoptosis and G2 /M cell cycle arrest. Interestingly, under BI 2536 treatment, D17 showed decreased c-Myc protein levels. Consistent with human OSA, these preliminary data outline the prognostic value of c-Myc expression in cOSA and highlight the potential role of PLK1 as an antiproliferative therapeutic target for tumours overexpressing c-Myc.

Chondroblastic osteosarcoma in a feline: a case report

Osteosarcomas are noteworthy as one of the main malignant bone tumors in dogs and cats, comprising an important cause of limb amputation in these species. Literature reports for felines are, however, scarce. Therefore, the aim of the present study was to characterize the clinical and histopathological findings of a chondroblastic osteosarcoma case in a female cat. The chondroblastic osteosarcoma diagnosis was based on radiographic, cytological, histopathological and immune-histochemical findings. Clinical, radiographic, and cytological findings suggested a presumptive osteosarcoma diagnosis, while histopathological and immune-histochemical findings confirmed a chondroblastic osteosarcoma diagnosis.

The use of alkaline phosphatase and runx2 to distinguish osteosarcoma from other common malignant primary bone tumors in dogs.

In dogs, primary bone tumors can be difficult to distinguish with histopathology. Of those tumors, osteosarcoma (OSA) is the most common and aggressive. In this study, 4 immunohistochemistry markers-alkaline phosphatase (ALP), osteonectin (ON), osteopontin (OP), and runx2-were evaluated for their ability to distinguish OSA from other primary bone tumors. The 42 formalin-fixed, paraffin-embedded, primary canine bone tumors included 15 OSAs, 8 chondrosarcomas, 11 fibrosarcomas, and 8 histiocytic sarcomas. All 4 antibodies were highly sensitive for detection of osteosarcoma. ALP was the most sensitive at 100% and runx2 the most specific at 78%. Running ALP and runx2 in series resulted in a sensitivity of 87% and a specificity of 85%. This combination of immunomarkers resulted in a diagnostic panel for distinguishing osteosarcoma from other primary bone tumors.

Extrato etanólico de açafrão (Curcuma longa L.) reduz apoptose e promove proliferação de células de osteossarcoma canino

Abstract Curcuma longa L., also known as turmeric, has been widely studied for its various therapeutic properties, including antineoplastic action. The ethanolic extract of the plant contains several phenolic compounds, especially curcumin. Osteosarcoma is a predominant bone tumor in dogs and humans, characterized by high metastatic potential and an unfavorable prognosis. The aim of this study was to investigate the effects of turmeric ethanol extract on canine osteosarcoma cells from established culture. The cells were cultured and treated with different curcumin concentrations (0, 10 μM, 20 μM, 50 μM, 100 μM, and 1000 μM) and exposure times (24h, 48h, and 72h). We first performed tetrazolium reduction technique (MTT) assay and calculated IC50. An immunocytochemistry assay was performed after extract treatment to verify the expression of mutated p53 and therefore study the proliferative potential of malignant cells; Bcl-2 and Ki-67 were used to assess apoptosis and the degree of malignancy, respectively. The extract enhanced the proliferation of canine osteosarcoma cells, reaching 3,819.74% at 50 μM of curcumin. The extract also significantly altered the expression of mutated p53 and Ki-67 proteins but not that of Bcl-2, suggesting that it did not induce this antiapoptotic pathway. Overall, these results are prerequisite to better understanding how natural compounds such as turmeric ethanolic extract affect cell proliferation and could be used to treat various diseases.

Genome-Wide Analyses for Osteosarcoma in Leonberger Dogs Reveal the CDKN2A/B Gene Locus as a Major Risk Locus.

Dogs represent a unique spontaneous cancer model. Osteosarcoma (OSA) is the most common primary bone tumor in dogs (OMIA 001441-9615), and strongly resembles human forms of OSA. Several large- to giant-sized dog breeds, including the Leonberger, have a greatly increased risk of developing OSA. We performed genome-wide association analysis with high-density imputed SNP genotype data from 273 Leonberger cases with a median age of 8.1 [3.1–13.5] years and 365 controls older than eight years. This analysis revealed significant associations at the CDKN2A/B gene locus on canine chromosome 11, mirroring previous findings in other dog breeds, such as the greyhound, that also show an elevated risk for OSA. Heritability (h2SNP) was determined to be 20.6% (SE = 0.08; p-value = 5.7 × 10−4) based on a breed prevalence of 20%. The 2563 SNPs across the genome accounted for nearly all the h2SNP of OSA, with 2183 SNPs of small effect, 316 SNPs of moderate effect, and 64 SNPs of large effect. As with many other cancers it is likely that regulatory, non-coding variants underlie the increased risk for cancer development. Our findings confirm a complex genetic basis of OSA, moderate heritability, and the crucial role of the CDKN2A/B locus leading to strong cancer predisposition in dogs. It will ultimately be interesting to study and compare the known genetic loci associated with canine OSA in human OSA.

Genomic and Transcriptomic Characterization of Canine Osteosarcoma Cell Lines: A Valuable Resource in Translational Medicine.

Osteosarcoma (OSA) represents the most common primary bone tumor in dogs and is characterized by a highly aggressive behavior. Cell lines represent one of the most suitable and reproducible pre-clinical models, and therefore the knowledge of their molecular landscape is mandatory to investigate oncogenic mechanisms and drug response. The present study aims at determining variants, putative driver genes, and gene expression aberrations by integrating whole-exome and RNA sequencing. For this purpose, eight canine OSA cell lines and one matched pair of primary tumor and normal tissue were analyzed. Overall, cell lines revealed a mean tumor mutational burden of 9.6 mutations/Mb (range 3.9–16.8). Several known oncogenes and tumor suppressor genes, such as ALK, MYC, and MET, were prioritized as having a likely role in canine OSA. Mutations in eight genes, previously described as human OSA drivers and including TP53, PTCH1, MED12, and PI3KCA, were retrieved in our cell lines. When variants were cross-referenced with human OSA driver mutations, the E273K mutation of TP53 was identified in the Wall cell line and tumor sample. The transcriptome profiling detected two possible p53 inactivation mechanisms in the Wall cell line on the one hand, and in D17 and D22 on the other. Moreover, MET overexpression, potentially leading to MAPK/ERK pathway activation, was observed in D17 and D22 cell lines. In conclusion, our data provide the molecular characterization of a large number of canine OSA cell lines, allowing future investigations on potential therapeutic targets and associated biomarkers. Notably, the Wall cell line represents a valuable model to empower prospective in vitro studies both in human and in dogs, since the TP53 driver mutation was maintained during cell line establishment and was widely reported as a mutation hotspot in several human cancers.

Molecular Mechanisms of Canine Osteosarcoma Metastasis.

Osteosarcoma (OSA) represents the most common bone tumor in dogs. The malignancy is highly aggressive, and most of the dogs die due to metastasis, especially to the lungs. The metastatic process is complex and consists of several main steps. Assessment of the molecular mechanisms of metastasis requires in vitro and especially in vivo studies for a full evaluation of the process. The molecular and biological resemblance of canine OSA to its human counterpart enables the utilization of dogs as a spontaneous model of this disease in humans. The aim of the present review article is to summarize the knowledge of genes and proteins, including p63, signal transducer and activator of transcription 3 (STAT3), Snail2, ezrin, phosphorylated ezrin-radixin-moesin (p-ERM), hepatocyte growth factor-scatter factor (HGF-SF), epidermal growth factor receptor (EGFR), miR-9, and miR-34a, that are proven, by in vitro and/or in vivo studies, to be potentially involved in the metastatic cascade of canine OSA. The determination of molecular targets of metastatic disease may enhance the development of new therapeutic strategies.

A Holistic Approach to Bone Tumors in Dogs and Cats: Radiographic and Histologic Correlation.

The skeletal system is a common site for neoplasia in dogs and cats, and primary bone tumors may develop from any of the mesenchymal tissues present in bone. Imaging and histopathology are routinely used in the diagnosis of bone tumors, and the 2 techniques are highly complementary. While imaging may be highly suggestive of a specific diagnosis and treatment may be instituted based on this, definitive diagnosis requires histopathology of either incisional or excisional biopsies or an amputation specimen. However, there are a number of diagnostic dilemmas when the pathologist interprets bone biopsy samples, such as distinguishing reactive bone and tumor bone, fracture callus and tumor bone, different benign fibro-osseous lesions, and different types of bone sarcoma. This review outlines the characteristic radiographic and histologic changes associated with these diagnostic problems to aid in resolving them. When a holistic approach is taken to evaluation of the signalment, history, and clinical, radiologic, and microscopic features, a diagnosis may be possible. The pathologist is greatly assisted in the interpretation of bone samples by having access to imaging and should routinely request either the images or the imaging reports if they are not received from submitting veterinarians.

Curcumina promove apoptose extrínseca em células de osteossarcoma canino

O osteossarcoma canino é o tumor ósseo mais comum em cães. Ele apresenta intensa capacidade metastásica e a sobrevida do paciente é baixa nessa doença. A curcumina, o composto mais importante derivado da planta Curcuma longa L., tem sido amplamente estudada e mostrou efeitos antineoplásicos consideráveis contra vários tumores. Este estudo tem por objetivo identificar a ativação de proteínas específicas de vias de apoptose, sobrevivência tumoral e mau prognóstico dessa doença em células OSC da linhagem D-17. Para isso, as células foram cultivadas e tratadas com a curcumina nas concentrações de 20μM, 50 μM e 100 μM em lâminas, que foram preparadas e fixadas. Posteriormente, foi realizada a técnica de imucitoquímica, com os anticorpos anti-caspase3, anti-JNK, anti-AMPK, anti-p53, anti-AKT e anti-mTOR. Foi observado que a curcumina ativou em células de osteossarcoma canino in vitro as proteínas de morte celular caspase-3, JNK e AMPK, reduziu a expressão da proteína p53 mutada e não alterou as proteínas AKT e mTOR. Assim, verificou-se que a curcumina promove apoptose extrínseca mediada por caspase, JNK e cAMP/AMPK em células de osteossarcoma canino. Além disso, tem o potencial de melhorar o prognóstico tumoral dessa doença por inativação da p53 mutada. No entanto, ela não interfere na expressão de AKT/mTOR, relacionado a proliferação e sobrevivência tumoral. Tais resultados servirão de base para estudos futuros que analisem o efeito da curcumina in vivo nessa doença.

Safety evaluation of the canine osteosarcoma vaccine, live Listeria vector.

Canine osteosarcoma (OSA) is an aggressive bone tumour in dogs. Standard‐of‐care treatment typically results in relatively short survival times; thus, alternative treatments are needed to confer a survival advantage. It has been shown that OSA is an immunogenic tumour, suggesting that immune modulation may result in superior outcomes. A cryopreserved, Listeria‐based OSA vaccine was recently developed and an initial study in dogs reported prolonged survival for patients receiving the vaccine in conjunction with standard‐of‐care. The goal of the current observational study was to report on the safety of the lyophilized formulation of this vaccine (the canine OSA vaccine, live Listeria vector [COV‐LLV]) in a group of dogs previously diagnosed with OSA. Forty‐nine (49) dogs received the COV‐LLV and were included for analysis. Adverse events (AEs) noted during and after vaccinations were recorded. The AEs observed were typically mild and self‐limiting, with nausea, lethargy and fever being most common. Four dogs (8%) cultured positive for Listeria (three infections including an amputation site abscess, septic stifle joint and bacterial cystitis; and one dog whose lungs cultured Listeria‐positive on necropsy within 24 hours of COV‐LLV administration). These cases join the previously reported Listeria‐positive thoracic abscess that developed in a canine following use of COV‐LLV. Although uncommon, it is important to realize this clinically significant AE is possible in patients treated with live therapeutic Listeria vaccines. As Listeria is zoonotic, caution is required not only for the patient receiving the vaccine, but also for the health care workers and family caring for the patient.

Abstract B59: Validation of an exosomal osteosarcoma-associated gene signature in dogs with osteosarcoma

Abstract Introduction: Osteosarcoma, the most common primary bone tumor in dogs, has a guarded prognosis. A major hurdle in developing more effective therapies is the lack of osteosarcoma-specific biomarkers to predict risk, prognosis, or therapeutic response. Exosomes are secreted microvesicles emerging as powerful diagnostic tools. However, wide clinical use of exosomes is precluded by the difficulty in identifying diseased cargo from the vastly larger background of normal cargo. We developed a method to distinguish tumor-derived exosome cargo from normal background, allowing for identification of an osteosarcoma-specific gene signature. Methods: Serum exosomes were enriched from osteosarcoma xenografts and control mice. Enriched genes associated with canine osteosarcoma were identified with RNA-sequencing. From identified candidates, we defined an osteosarcoma-associated gene signature. qRT-PCR amplification validated the gene signature in serum exosomes from clinical canine cases. Machine learning algorithms classified patients into disease groups based on this gene signature. Results: We identified an osteosarcoma-associated signature consisting of five mRNAs (SKA2, NEU1, PAF1, PSMG2, and NOB1). Serum exosomes were isolated from dogs in clinical groups, including “healthy,” “osteosarcoma,” “other bone tumor,” or “non-neoplastic disease.” Machine learning classified samples, with 82% and 86% of untrained samples predicted as osteosarcoma by CN2 and RF models, respectively. Post-treatment samples “misclassified” as non-osteosarcoma were associated with longer remissions, potentially from a lack of remaining osteosarcoma cells. Conclusions: We identified a gene signature associated with canine osteosarcoma. This gene signature was validated by qRT-PCR with serum exosomes from patients with osteosarcoma, as well as used to train artificial intelligence to correctly classify canine patients according to disease group with up to 93.8% accuracy. Clinical Significance: This study combines a bioinformatics approach to biomarker discovery with machine learning to correctly identify osteosarcoma in canine patients. These results set the stage for future discoveries to inform cancer risk, diagnosis, prognosis, and response to therapy. Citation Format: Kelly M. Makielski, Alicia J. Donnelly, Milcah C. Scott, Hirotaka Tomiyasu, Ali Khammanivong, William C. Kisseberth, Jaime F. Modiano. Validation of an exosomal osteosarcoma-associated gene signature in dogs with osteosarcoma [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B59.

MiR-106B-25 Cluster expression: a comparative human and canine osteosarcoma study

BackgroundOsteosarcoma (OS) is the most common primary malignant bone tumour in dogs and human beings, characterised by similar genetic and clinical features. With the aim to define similarities and differences...