ACVR and ECVDI consensus statement: Reporting elements for toxicity criteria of the veterinary radiation therapy oncology group v2.0.

The toxicity criteria of the veterinary radiation therapy oncology group (VRTOG) version 2 guidelines are a substantial update to reflect significant advances in radiation oncology over the last three decades. Radiation therapy techniques provide precise and spatially accurate radiation delivery, which facilitates treating tumors in more anatomic locations and incorporating hypofractionated protocols. The purpose of this update is to aid radiation oncology teams in capturing and grading clinically relevant data that impacts the decision-making process in everyday practice and the assessment of clinical trials involving radiation therapy. A dedicated committee initially updated the criteria to include more anatomical sites and grades to characterize a broad spectrum of possible radiation-induced acute and late tissue changes. Through the revision process, which solicited and incorporated feedback from all radiation oncologists within the American College of Veterinary Radiology (ACVR) and specialists outside the ACVR, the authors endeavored to create a grading structure reflective of clinical decision-making in daily radiation oncology. The updated VRTOG v2 toxicity criteria guideline complements the updated Veterinary Cooperative Oncology Group-Common Terminology Criteria for Adverse Events (VCOG-CTCAE v2) guidelines. Because radiation oncology continues to progress rapidly, the VRTOG toxicity criteria should be regularly updated as adverse event data that will be collected following this update further informs the practice of radiation oncology.

First, do no harm. Ethical and legal issues of artificial intelligence and machine learning in veterinary radiology and radiation oncology.

Artificial Intelligence and machine learning are novel technologies that will change the way veterinary medicine is practiced. Exactly how this change will occur is yet to be determined, and, as is the nature with disruptive technologies, will be difficult to predict. Ushering in this new tool in a conscientious way will require knowledge of the terminology and types of AI as well as forward thinking regarding the ethical and legal implications within the profession. Developers as well as end users will need to consider the ethical and legal components alongside functional creation of algorithms in order to foster acceptance and adoption, and most importantly to prevent patient harm. There are key differences in deployment of these technologies in veterinary medicine relative to human healthcare, namely our ability to perform euthanasia, and the lack of regulatory validation to bring these technologies to market. These differences along with others create a much different landscape than AI use in human medicine, and necessitate proactive planning in order to prevent catastrophic outcomes, encourage development and adoption, and protect the profession from unnecessary liability. The authors offer that deploying these technologies prior to considering the larger ethical and legal implications and without stringent validation is putting the AI cart before the horse, and risks putting patients and the profession in harm's way.

A NOVEL, REMOVABLE, CERROBEND, BEAM-BLOCKING DEVICE FOR RADIATION THERAPY OF THE CANINE HEAD AND NECK: PILOT STUDY.

Radiation therapy of the head and neck can result in mucositis and other acute affects in the oral cavity. This prospective pilot study evaluated a novel, intraoral, beam-blocking device for use during imaging and therapeutic procedures. The beam-blocking device was made from a metal alloy inserted into a coated frozen dessert mold (Popsicle® Mold, Cost Plus World Market, Oakland, CA). The device was designed so that it could be inserted into an outer shell, which in turn allowed it to be placed or removed depending on the need due to beam configuration. A Farmer type ionization chamber and virtual water phantom were used to assess effects of field size on transmission. Six large breed cadaver dogs, donated by the owner after death, were recruited for the study. Delivered dose at the dorsal and ventral surfaces of the device, with and without the alloy block in place, were measured using radiochromic film. It was determined that transmission was field size dependent with larger field sizes leading to decreased attenuation of the beam, likely secondary to scatter. The mean and median transmission on the ventral surface without the beam-blocking device was 0.94 [range 0.94-0.96]. The mean and median transmission with the beam-blocking device was 0.52 [range 0.50-0.57]. The mean and median increase in dose due to backscatter on the dorsal surface of the beam-blocking device was 0.04 [range 0.02-0.04]. Findings indicated that this novel device can help attenuate radiation dose ventral to the block in dogs, with minimal backscatter.

Tolerability of oral sorafenib in pet dogs with a diagnosis of cancer.

Sorafenib is a multi-target small molecule inhibitor of the RAF kinase family and VEGFR-2/PDGFR. The US Food and Drug Administration approved sorafenib in human patients with liver, thyroid, or renal carcinoma. The aim of this study was to help guide future pharmacokinetic (PK) studies of sorafenib in dogs with a cancer diagnosis. Client-owned dogs were eligible if they had a cytologic or histologic diagnosis of cancer. Patients were enrolled at escalating doses of sorafenib. Patients were evaluable for the study if they received at least one dose of sorafenib and presented 1 week later for a follow-up examination, blood work, and assessment of drug tolerability. The goal of this study was not to define a maximum tolerated dose as may be reasonable in conventional cytotoxic chemotherapy drugs, but rather to describe the tolerability of this drug in dogs with a cancer diagnosis, as a prequel to future sorafenib PK studies. No patients in the study had any evidence of adverse events that were attributable to sorafenib. Doses of 3 mg/kg were well tolerated and associated with a suggestion of clinical activity, supportive of future PK, and pharmacodynamic analysis. Such future studies are recommended at this dose to define the associated exposure achieved and determine a reasonable schedule for sorafenib administration.

Protein Phosphatase 2A Inhibition with LB100 Enhances Radiation-Induced Mitotic Catastrophe and Tumor Growth Delay in Glioblastoma.

Protein phosphatase 2A (PP2A) is a tumor suppressor whose function is lost in many cancers. An emerging, though counterintuitive, therapeutic approach is inhibition of PP2A to drive damaged cells through the cell cycle, sensitizing them to radiotherapy. We investigated the effects of PP2A inhibition on U251 glioblastoma cells following radiation treatment in vitro and in a xenograft mouse model in vivo. Radiotherapy alone augmented PP2A activity, though this was significantly attenuated with combination LB100 treatment. LB100 treatment yielded a radiation dose enhancement factor of 1.45 and increased the rate of postradiation mitotic catastrophe at 72 and 96 hours. Glioblastoma cells treated with combination LB100 and radiotherapy maintained increased γ-H2AX expression at 24 hours, diminishing cellular repair of radiation-induced DNA double-strand breaks. Combination therapy significantly enhanced tumor growth delay and mouse survival and decreased p53 expression 3.68-fold, compared with radiotherapy alone. LB100 treatment effectively inhibited PP2A activity and enhanced U251 glioblastoma radiosensitivity in vitro and in vivo. Combination treatment with LB100 and radiation significantly delayed tumor growth, prolonging survival. The mechanism of radiosensitization appears to be related to increased mitotic catastrophe, decreased capacity for repair of DNA double-strand breaks, and diminished p53 DNA-damage response pathway activity.

Ionizing radiation and glioblastoma exosomes: implications in tumor biology and cell migration.

Exosomes are nanometer-sized lipid vesicles released ubiquitously by cells, which have been shown to have a normal physiological role, as well as influence the tumor microenvironment and aid metastasis. Recent studies highlight the ability of exosomes to convey tumor-suppressive and oncogenic mRNAs, microRNAs, and proteins to a receiving cell, subsequently activating downstream signaling pathways and influencing cellular phenotype. Here, we show that radiation increases the abundance of exosomes released by glioblastoma cells and normal astrocytes. Exosomes derived from irradiated cells enhanced the migration of recipient cells, and their molecular profiling revealed an abundance of molecules related to signaling pathways important for cell migration. In particular, connective tissue growth factor (CTGF) mRNA and insulin-like growth factor binding protein 2 (IGFBP2) protein levels were elevated, and coculture of nonirradiated cells with exosomes isolated from irradiated cells increased CTGF protein expression in the recipient cells. Additionally, these exosomes enhanced the activation of neurotrophic tyrosine kinase receptor type 1 (TrkA), focal adhesion kinase, Paxillin, and proto-oncogene tyrosine-protein kinase Src (Src) in recipient cells, molecules involved in cell migration. Collectively, our data suggest that radiation influences exosome abundance, specifically alters their molecular composition, and on uptake, promotes a migratory phenotype.

Radiosensitization by the novel DNA intercalating agent vosaroxin.

PURPOSE: Vosaroxin is a first in class naphthyridine analog structurally related to quinolone antibacterials, that intercalates DNA and inhibits topoisomerase II. Vosaroxin is not a P-glycoprotein receptor substrate and its activity is independent of p53, thus evading common drug resistance mechanisms. To evaluate vosaroxin as a clinically applicable radiation sensitizer, we investigated its effects on tumor cell radiosensitivity in vitro and in vivo. METHODS: Vosaroxin's effect on post-irradiation sensitivity of U251, DU145, and MiaPaca-2 cells was assessed by clonogenic assay. Subsequent mechanistic and in vivo studies were performed with U251 cells. Cell cycle distribution and G2 checkpoint integrity was analyzed by flow cytometry. DNA damage and repair was evaluated by a high throughput gamma-H2AX assay. Apoptosis was assessed by flow cytometry. Mitotic catastrophe was assessed by microscopic evidence of fragmented nuclei by immunofluorescence. In vivo radiosensitization was measured by subcutaneous tumor growth delay. RESULTS: 50-100 nmol/L treatment with vosaroxin resulted in radiosensitization of all 3 cell lines tested with a dose enhancement factor of 1.20 to 1.51 measured at a surviving fraction of 0.1. The maximal dose enhancement was seen in U251 cells treated with 75 nmol/L vosaroxin (DEF 1.51). Vosaroxin exposure did not change cell cycle distribution prior to irradiation nor alter G2 checkpoint integrity after irradiation. No difference was seen in the apoptotic fraction regardless of drug or radiation treatment. The number of cells in mitotic catastrophe was significantly greater in irradiated cells treated with vosaroxin than cells receiving radiation only at 72 hr (p = 0.009). Vosaroxin alone did not significantly increase mitotic catastrophe over control (p = 0.53). Cells treated with vosaroxin and radiation maintained significantly higher gamma-H2AX levels than cells treated with vehicle control (p = 0.014), vosaroxin (p = 0.042), or radiation alone (p = 0.039) after 24 hr. In vivo tumor growth delay was 1.5 days for vosaroxin alone (IV 10 mg/kg), 1.0 days for radiation (3 Gy) alone, and 8.6 days for the group treated with vosaroxin 4 hours prior to radiation. CONCLUSIONS: Vosaroxin enhanced tumor cell radiosensitivity in vitro and in vivo. The mechanism appears to be related to inhibition of DNA repair and increased mitotic catastrophe.

Development and characterization of 5 canine B-cell lymphoma cell lines.

Canine and human lymphoma share similar characteristics in disease development and response to therapy. Translational research can be furthered using tools such as canine cell lines to model therapeutic compounds and strategies. We developed 5 B-cell lymphoma cell lines from dogs with confirmed large B-cell lymphoma. These cell lines were CD3, CD18, CD20, and CD90 positive with variable CD79a, CD1c and CD34 expression. All cell lines were tumorigenic in Nu/nu mice and were wild type for p53. Canine lymphoma cell lines serve as an important resource for translational lymphoma research.

Modeling opportunities in comparative oncology for drug development.

Successful development of novel cancer drugs depends on well-reasoned scientific drug discovery, rigorous preclinical development, and carefully conceived clinical trials. Failure in any of these steps contributes to poor rates of approval for new drugs to treat cancer. As technological and scientific advances have opened the door to a variety of novel approaches to cancer drug discovery and development, preclinical models that can answer questions about the activity and safety of novel therapies are increasingly necessary. The advance of a drug to clinical trials based on information from preclinical models presupposes that the models convey informative data for future use in human patients with cancer. The study of novel cancer drugs using in vitro models is highly controllable, reproducible, relatively inexpensive, and linked to high throughput. However, these models fail to reproduce many of the complex features of human cancer. Mouse models address some of these limitations but have important biological differences from human cancer. The integration of studies using pet dogs with spontaneously occurring tumors as models in the development path can answer questions not adequately addressed in conventional models and is therefore gaining attention and interest in drug development communities. The study of novel cancer drugs in dogs with naturally occurring tumors allows drug assessment in a cancer that shares many fundamental features with the human cancer condition, and thus provides an opportunity to answer questions that inform the cancer drug development path in ways not possible in more conventional models.

Spontaneous hemoperitoneum in cats: 65 cases (1994-2006).

OBJECTIVE: To describe the clinical signs, physical examination findings, clinical laboratory abnormalities, etiology, and outcome in cats with spontaneous hemoperitoneum. DESIGN: Retrospective case series. ANIMALS: 65 client-owned cats with spontaneous hemoperitoneum. PROCEDURES: Medical records of cats with spontaneous hemoperitoneum at 7 large referral clinics were reviewed. Cats were included if a definitive diagnosis of spontaneous hemoperitoneum could be obtained from review of the medical records. RESULTS: 65 cats met inclusion criteria. The most common historical findings were lethargy, anorexia, and vomiting. Common findings on physical examination included inadequate hydration status and hypothermia. The most common clinicopathologic abnormalities were high serum AST activity, anemia, prolonged prothrombin time, and prolonged partial thromboplastin time. Forty-six percent (30/65) of cats had abdominal neoplasia, and 54% (35/65) had nonneoplastic conditions. Hemangiosarcoma was the most often diagnosed neoplasm (18/30; 60%), and the spleen was the most common location for neoplasia (11/30; 37%). Eight cats survived to be discharged from the hospital. Cats with neoplasia were significantly older and had significantly lower PCVs than cats with non-neoplastic disease. CONCLUSIONS AND CLINICAL RELEVANCE: Spontaneous hemoperitoneum in cats often results in debilitating clinical consequences. In contrast to dogs with hemoperitoneum, the cause of hemoperitoneum in cats is approximately evenly distributed between neoplastic and nonneoplastic diseases. Although only a few cats were treated in this study, the prognosis appears poor.

Assessment of the accuracy and precision of a patient immobilization device for radiation therapy in canine head and neck tumors.

The positioning accuracy and precision of a head and neck immobilization device for radiation therapy of tumors in the canine skull was evaluated. Nineteen dogs with a spontaneous tumor of the head were enrolled including 12 with an intracranial mass and seven with an intranasal or maxillary tumor. Three hundred thirty-three pairs of orthogonal digital portal radiographs were analyzed to assess patient displacement in the cranial-caudal, lateral, and dorso-ventral directions. The mean systematic displacement was 0.8, 1, and 0.9 mm. The mean random displacement was 1.9, 1.6, and 1.5 mm. These values resulted in an overall displacement of 2.1 mm in the cranial-caudal direction, 1.8mm in the lateral direction, and 1.7 mm in the dorsal-ventral direction. The mean displacement value of the three dimensional (3D) vector was 2.4 mm with a standard deviation of 2.1. Ninety-five percent of all vectors were < 6.4 mm. This study quantifies the precision and accuracy of this particular positioning device. Knowing the limitations and setup variability of the system being used to set patients up for daily radiotherapy is paramount to planning and delivering appropriate radiation doses, especially as more complex treatment methods are used.

Temporal changes in characteristics of injection-site sarcomas in cats: 392 cases (1990-2006).

OBJECTIVE: To evaluate changes in characteristics of feline injection-site sarcomas (ISSs) from 1990 through 2006. DESIGN: Retrospective case series. ANIMALS: 392 cats with a histologic diagnosis of soft tissue sarcoma, osteosarcoma, or chondrosarcoma at potential injection sites. PROCEDURES: Classification and anatomic location of tumors and signalment of affected cats were compared between ISSs diagnosed before and after publication of the Vaccine Associated Feline Sarcoma Task Force vaccination recommendations in 1996. RESULTS: From before to after publication of the vaccination recommendations, proportions of ISSs significantly decreased in the interscapular (53.4% to 39.5%) and right and left thoracic (10.2% to 3.6% and 9.1% to 1.3%, respectively) regions. On the other hand, proportions of ISSs significantly increased in the right thoracic limb (1.1% to 9.5%) and the combined regions of the right pelvic limb with right lateral aspect of the abdomen (12.5% to 25.0%) and the left pelvic limb with left lateral aspect of the abdomen (11.4% to 13.8%). Patterns of tumor classification and signalment did not change. CONCLUSIONS AND CLINICAL RELEVANCE: Despite publication of the vaccination recommendations, a high proportion of tumors still developed in the interscapular region. There was also an increase in lateral abdominal ISSs, which are more difficult to treat and are likely attributable to aberrant placement of injections intended for the pelvic limbs. Veterinarians are complying with vaccination recommendations to some extent, but they need to focus on administering vaccines as distally as possible on a limb to allow for complete surgical margins if amputation of a limb is required.

Evaluation of the mammalian target of rapamycin pathway and the effect of rapamycin on target expression and cellular proliferation in osteosarcoma cells from dogs.

OBJECTIVE: To investigate activation of the mammalian target of rapamycin (mTOR) pathway and the antitumor effect of rapamycin in canine osteosarcoma cells. SAMPLE POPULATION: 3 established primary canine osteosarcoma cell lines generated from naturally developing tumors. PROCEDURES: Expression of total and phosphorylated mTOR and p70S6 kinase was assessed by use of western blot analysis in canine osteosarcoma cells with and without the addition of rapamycin. A clonogenic assay was performed to determine the surviving fraction of osteosarcoma cells at various concentrations of rapamycin. RESULTS: Total and phosphorylated mTOR and p70S6 kinase expression was evident in all 3 cell lines evaluated, which was indicative of activation of this pathway. Treatment with rapamycin resulted in a time-dependent decrease in phosphorylated mTOR expression and a lack of detectable phosphorylated p70S6 kinase. No detectable change in expression of total mTOR and total p70S6 kinase was identified after rapamycin treatment. The clonogenic assay revealed a significant dose-dependent decrease in the surviving fraction for all 3 cell lines when treated with rapamycin. CONCLUSIONS AND CLINICAL RELEVANCE: These data indicated that mTOR and its downstream product are present and active in canine osteosarcoma cells. The pathway can be inhibited by rapamycin, and treatment of cells with rapamycin decreased the surviving tumor cell fraction. These data support the molecular basis for further investigation into the use of mTOR inhibitors as an antineoplastic approach for dogs with osteosarcoma.

Evaluation of strontium Sr 90 for the treatment of superficial squamous cell carcinoma of the nasal planum in cats: 49 cases (1990-2006).

OBJECTIVE: To determine progression-free and overall survival times of cats with squamous cell carcinoma (SCC) of the nasal planum following treatment with a single fraction of strontium Sr 90 ((90)Sr). DESIGN: Retrospective case series. ANIMALS: 49 cats with SCC of the nasal planum. PROCEDURES: Information including FIV infection status, diagnosis of SCC vs SCC in situ (ie, evidence that the tumor did or did not penetrate the epidermal basement membrane, respectively), (90)Sr dose and number of probe applications, treatment-related response and complications, and recurrence of SCC and new lesion development was obtained from medical records. The relationships of these variables with calculated progression-free and overall survival times were assessed. RESULTS: Of 49 cats that underwent (90)Sr plesiotherapy (median dose, 128 Gy), 48 (98%) had a response to treatment and 43 (88%) had a complete response. Median progression-free and overall survival times were 1,710 and 3,076 days, respectively. Treatment complications were infrequent (4 [8%] cats) and mild. Following treatment, the SCC recurrence rate was 20% (10/49 cats); 16 (33%) cats developed new lesions in other locations. Overall survival time was significantly longer for cats with a complete response to treatment than for those with a partial response. None of the other variables evaluated had a significant effect on progression-free or overall survival time. CONCLUSIONS AND CLINICAL RELEVANCE: Treatment of cats with SCC of the nasal planum with a single fraction of (90)Sr appeared to be effective and well tolerated. Initial response to treatment was predictive of overall survival time.