Long-term assessment of risk factors for canine tumors registered in Xi’an, China

Tumors are one of the leading causes to death in pet dogs among diseases. The tumor incidence of pet dogs has been increasing, raising widespread concern.The tumor incidence of pet dogs has been increasing, raising widespread concern. In this study, retrospective analysis was performed with 246 tumor cases registered in Xi’an Animal Hospital, Northwest A&F University from 2009 to 2018. Correlations of sex, age and breed with tumor incidences were evaluated. The results showed that reproductive system tumors occupied the highest proportion (39.84%), followed by cutaneous tumors (28.05%), digestive tumors (18.70%) and ocular tumor (4.47%). Among the reproductive system tumors, breast tumors are the most common tumor in female pet dogs, especially for Pekingese (11.43%). Female dogs with high susceptibility to breast tumors were at the ages of 6–18 years old. As far as cutaneous tumors were concerned, the male pet dogs at all ages, particularly Golden Retrievers (17.39%), showed a high incidence. By contrast, male Samoyed aged from 4 to 13 years had the highest incidence (15.22%) of digestive tumors. In addition, pet dogs with ocular tumors mainly happened at the ages of 0–1 years and 6–13 years. Collectively, our findings are significant to develop effective measures of medical surveillance for pet dogs’ health and will provide insights for comparative oncology.

Intraoperative Ultrasound Applications in Intracranial Surgery

The methods and use of intraoperative ultrasound in 33 canine and five feline patients and its ability to localize and identify anatomical structures and pathological lesions in canines and felines undergoing intracranial surgery are described from a case series. All were client-owned referral patients admitted for neurologic evaluation, with an advanced imaging diagnosis of an intracranial lesion, and underwent surgical biopsy or surgical removal of the lesion. Medical records, retrieval and review of imaging reports, and characterization of findings for all canine and feline patients show that intraoperative ultrasound guidance was used in intracranial procedures during the period of 2012 and 2019. Twenty-nine of the canine patients had intracranial tumors. The remainder had various other conditions requiring intracranial intervention. Three of the feline patients had meningiomas, one had a depressed skull fracture, and one had an epidural hematoma. The tumors appeared hyperechoic on intraoperative ultrasound with the exception of cystic portions of the masses and correlated with the size and location seen on advanced imaging. Statistical comparison of the size of images seen on ultrasound and on MRI for 20 of the canine tumors revealed no statistical differences. Neuroanatomical structures, including vascular components, were easily identified, and tumor images correlated well with preoperative advanced imaging. The authors conclude that intraoperative ultrasound is a valuable asset in intracranial mass removals and can augment surgical guidance in a variety of intracranial disorders that require surgery. This is the first known publication in veterinary surgery of using intraoperative ultrasound as a tool in the operating theater to identify, localize, and monitor the removal/biopsy of intracranial lesions in small animals undergoing craniotomy/craniectomy.

Shared hotspot mutations in spontaneously arising cancers position dog as an unparalleled comparative model for precision therapeutics

Naturally occurring canine cancers have remarkable similarities to their human counterparts. In order to determine whether these similarities occur at the molecular level, we investigated hotspot mutations in a variety of spontaneously arising canine cancers and found high concordance in oncogenic drivers between cancers in both species. These findings suggest that canines may present a powerful and complementary model for preclinical investigations for targeted cancer therapeutics. Through analysis of 708 client-owned dogs from 96 breeds (plus mixed breeds) with 23 common tumor types, we discovered mutations in 50 well-established oncogenes and tumor suppressors, and compared them to those reported in human cancers. TP53 is the most commonly mutated gene, detected in 30.81% of canine tumors overall and >40% in hemangiosarcoma and osteosarcoma. Canine tumors share mutational hotspots with human tumors in oncogenes including PIK3CA, KRAS, NRAS, BRAF, KIT and EGFR. Hotspot mutations with significant (P<0.0001) association to tumor type include NRAS G61R and PIK3CA H1047R in hemangiosarcoma, ERBB2 V659E in pulmonary carcinoma, and BRAF V588E in urothelial carcinoma. This work positions canines as excellent spontaneous models of human cancers that can help to investigate a wide spectrum of targeted therapies.

Characterization of receptor tyrosine kinase activation and biological activity of toceranib phosphate in canine urothelial carcinoma cell lines

Background Urothelial carcinoma (UC) accounts for > 90% of canine tumors occurring in the urinary bladder. Toceranib phosphate (TOC) is a multi-target receptor tyrosine kinase (RTK) inhibitor that exhibits activity against members of the split kinase family of RTKs. The purpose of this study was to evaluate primary UC tumors and UC cell lines for the expression and activation of VEGFR2, PDGFRα, PDGFRβ, and KIT to assess whether dysregulation of these RTKs may contribute to the observed biological activity of TOC. Results Transcript for VEGFR2, PDGFRα, PDGFRβ, and KIT was detected in all UC tissue samples and UC cell lines. The Proteome Profiler™ Human Phospho-RTK Array Kit (R & D Systems) provided a platform to assess phosphorylation of 42 different RTKs in primary UC tumors and UC cell lines. Evidence of PDGFRα and PDGFRβ phosphorylation was present in only 11% or 33% of UC tumors, respectively, and 25% of UC cell lines. Treatment of UC cell lines with TOC had no significant impact on cell proliferation, including UC cell lines with evidence of PDGFRβ phosphorylation. Conclusions Phosphorylation of several key RTKs targeted by TOC is present in a small subset of primary UC tumors and UC cell lines, suggesting that these RTKs do not exist in a state of continuous activation. These data suggest that activation of RTKs targeted by TOC is present in a small subset of UC tumors and UC cell lines and that treatment with TOC at physiologically relevant concentrations has no direct anti-proliferative effect on UC cells.

An Update on Molecular Pathways Regulating Vasculogenic Mimicry in Human Osteosarcoma and Their Role in Canine Oncology

Canine tumors are valuable comparative models for human counterparts, especially to explore novel biomarkers and to understand pathways and processes involved in metastasis. Vasculogenic mimicry (VM) is a unique property of malignant cancer cells which promote metastasis. Thus, it represents an opportunity to investigate both the molecular mechanisms and the therapeutic targets of a crucial phenotypic malignant switch. Although this biological process has been largely investigated in different human cancer types, including osteosarcoma, it is still largely unknown in veterinary pathology, where it has been mainly explored in canine mammary tumors. The presence of VM in human osteosarcoma is associated with poor clinical outcome, reduced patient survival, and increased risk of metastasis and it shares the main pathways involved in other type of human tumors. This review illustrates the main findings concerning the VM process in human osteosarcoma, search for the related current knowledge in canine pathology and oncology, and potential involvement of multiple pathways in VM formation, in order to provide a basis for future investigations on VM in canine tumors.

Mismatch repair deficiency in canine neoplasms

The DNA mismatch repair (MMR) system preserves genomic stability by identifying and repairing mismatched nucleotides in the DNA replication process. The dysfunction of the MMR system, also known as mismatch repair deficiency (dMMR), is implicated as a predictive biomarker for the efficacy of immune checkpoint blockade therapy regardless of the tumor type in humans. This study aimed to evaluate the immunolabeling of MMR proteins in canine tumors and to identify the types of tumors having dMMR. First, we performed immunohistochemistry in 8 different canine tumors (oral malignant melanoma, high-to-intermediate grade lymphoma, mast cell tumor, malignant mammary gland tumor, urothelial carcinoma, hepatocellular carcinoma, osteosarcoma, and hemangiosarcoma) with 15 samples each to analyze the immunolabeling of canine mismatch repair proteins (MSH2, MSH6, and MLH1) using anti-human monoclonal antibodies. We found that more than half of canine oral malignant melanoma (60%) and hepatocellular carcinoma (53%) samples and fewer of the other canine tumors had loss of immunolabeling in ≥1 MMR protein (ie, evidence of defective MMR proteins, based on the definition of dMMR in the humans). Antibodies against human MSH2, MSH6, and MLH1 were cross-reactive with the corresponding canine protein as confirmed using MMR gene knockout canine cell lines. Further studies are required to investigate the clinical outcomes in canine spontaneous tumors with dMMR to determine the potential for immune checkpoint blockade therapy for these tumor types.

A potential bat adenovirus-based oncolytic virus targeting canine cancers

Although a canine adenovirus (CAdV)-based oncolytic virus (OV) candidate targeting canine tumors has been reported, its oncolytic effect could be attenuated by CAdV vaccine-induced neutralizing antibodies in dog patients. To circumvent this issue, we focused on the bat adenovirus (BtAdV) strain, which was previously isolated from healthy microbats. We previously showed that this virus replicated efficiently in canine cell lines and did not serologically cross-react with CAdVs, suggesting that it may offer the possibility of an OV candidate for canine tumors. Here, we tested the growth properties and cytotoxicity of the BtAdV Mm32 strain in a panel of canine tumor cells and found that its characteristics were equivalent to those of CAdVs. To produce an Mm32 construct with enhanced tumor specificity, we established a novel reverse genetics system for BtAdV based on bacterial artificial chromosomes, and generated a recombinant virus, Mm32-E1Ap + cTERTp, by inserting a tumor-specific canine telomerase reverse transcriptase promoter into its E1A regulatory region. The growth and cytotoxicity of this recombinant were superior to those of wild-type Mm32 in canine tumor cells, unlike in normal canine cells. These data suggest that Mm32-E1Ap + cTERTp could be a promising OV for alternative canine cancer therapies.

MicroRNA Dysregulation in Canine Meningioma: RT-qPCR Analysis of Formalin-Fixed Paraffin-Embedded Samples

MicroRNAs (miRNAs) are small non-coding RNAs that play key roles in tumorigenesis as modulators of cell signaling pathways. miRNA expression has been found to be dysregulated in several human and canine tumors, but data are not yet available on canine meningioma. In this study, we analyzed the expression of 12 miRNAs (i.e. miR-335, miR-200a, miR-98, miR-96, miR-190a, miR-29c, miR-219-5p, miR-155, miR-146a, miR-145, miR-136, miR-451) by RT-qPCR in a series of 41 formalin-fixed, paraffin-embedded canine meningiomas, and normal arachnoid samples. We identified 8 dysregulated miRNAs that might be involved in canine meningioma pathogenesis. Five miRNAs (i.e. miR-96, miR-145, miR-335, miR-200a, miR-29c), were downregulated in tumor samples and 3 (i.e. miR-136, miR-155, miR-146a) were upregulated. Moreover, miR-200a was overexpressed in grade III compared to grade I and grade II meningiomas, suggesting that it might have a dual role in tumor initiation and progression. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses suggest that dysregulated miRNAs might influence cellular processes and pathways mainly involved in tumor cell migration, extracellular matrix interactions, cell proliferation, and inflammatory responses. The characterization of miRNA functions in canine meningiomas is needed to assess their potential clinical utility, also in view of the relevance of the dog as a potential spontaneous animal model of human disease.

Pharmacological Regulation of Tumor Hypoxia in Model Murine Tumors and Spontaneous Canine Tumors

Background: Hypoxia is found in many solid tumors and is associated with increased disease aggressiveness and resistance to therapy. Reducing oxygen demand by targeting mitochondrial oxidative metabolism is an emerging concept in translational cancer research aimed at reducing hypoxia. We have shown that the U.S. Food and Drug Administration (FDA)-approved drug papaverine and its novel derivative SMV-32 are potent mitochondrial complex I inhibitors. Methods: We used a dynamic in vivo luciferase reporter system, pODD-Luc, to evaluate the impact of pharmacological manipulation of mitochondrial metabolism on the levels of tumor hypoxia in transplanted mouse tumors. We also imaged canine patients with blood oxygen level-dependent (BOLD) MRI at baseline and one hour after a dose of 1 or 2 mg/kg papaverine. Results: We showed that the pharmacological suppression of mitochondrial oxygen consumption (OCR) in tumor-bearing mice increases tumor oxygenation, while the stimulation of mitochondrial OCR decreases tumor oxygenation. In parallel experiments in a small series of spontaneous canine sarcomas treated at The Ohio State University (OSU) Veterinary Medical Center, we observed a significant increase in BOLD signals indicative of an increase in tumor oxygenation of up to 10–50 mm HgO2. Conclusion: In both transplanted murine tumors and spontaneous canine tumors we found that decreasing mitochondrial metabolism can decrease tumor hypoxia, potentially offering a therapeutic advantage.