Design of a randomized, placebo-controlled study evaluating efficacy and safety of a cancer preventative vaccine in dogs.

Preventative anti-cancer vaccination strategies have long been hampered by the challenge of targeting the diverse array of potential tumor antigens, with successes to date limited to cancers with viral etiologies. Identification and vaccination against frameshift neoantigens conserved across multiple species and tumor histologies is a potential cancer preventative strategy currently being investigated. Companion dogs spontaneously develop cancers at a similar incidence to those in people and are a complementary comparative patient population for the development of novel anti-cancer therapeutics. In addition to an intact immune system with tumors that arise in an autochthonous tumor microenvironment, dogs also have a shorter lifespan and temporally compressed tumor natural history as compared to humans, which allows for more rapid evaluation of safety, immunogenicity, and efficacy of cancer vaccination strategies. Here we describe the study protocol for the Vaccination Against Canine Cancer Study (VACCS), the largest interventional cancer clinical trial conducted in companion dogs to date. In addition to safety and immunogenicity, the primary endpoint of VACCS is the cumulative incidence (CI) of dogs developing malignant neoplasia of any type at the end of the study period. Secondary endpoints include changes in incidence of specific tumor types, survival times following neoplasia diagnosis, and all-cause mortality.

Cohort profile: The Golden Retriever Lifetime Study (GRLS).

The aim of this article is to provide a detailed description of the Golden Retriever Lifetime Study (GRLS), a prospective cohort study investigating nutritional, environmental, lifestyle, and genetic risk factors for cancer and other common diseases in dogs. Primary outcomes of interest include hemangiosarcoma, lymphoma, osteosarcoma, and high-grade mast cell tumors. Secondary outcomes of interest include other cancers, hypothyroidism, epilepsy, atopy, otitis externa, hip dysplasia, heart failure, and renal failure. A total of 3,044 United States Golden Retrievers aged 6 months to 2 years completed baseline enrollment from June 2012 to April 2015. As of May 31, 2021, 2,251 dogs remain engaged in the study, 352 have died, and 441 are lost to follow-up. Extensive annual questionnaires completed by owners and veterinarians gather information about lifestyle, environmental exposures, physical activity, reproductive history, behavior, diet, medications, and diagnoses. Dogs also have annual veterinary examinations and biospecimen collection (blood, serum, hair, nails, feces, urine) for biobanking. Additional reporting, including histology and tumor biobanking, is conducted for any malignancies or deaths. When an animal dies, full medical records are obtained, and necropsies are requested at owner discretion. Full or partial necropsies have been performed on 218 dogs. Questionnaire data are freely available to researchers with approved credentials who agree to a data use agreement. In addition, researchers can submit proposals to utilize biospecimens or obtain additional data.

Characterizing the molecular and immune landscape of canine bladder cancer.

Transitional cell carcinoma (TCC), also known as urothelial carcinoma, is the most common bladder cancer in humans and dogs. Approximately one-quarter of human TCCs are muscle-invasive and associated with a high risk of death from metastasis. Canine TCC (cTCC) tumours are typically high-grade and muscle-invasive. Shared similarities in risk factors, histopathology, and clinical presentation suggest that cTCC may serve as a model for the assessment of novel therapeutics that may inform therapies for human muscle-invasive TCC. The goal of this study was to characterize cTCC at the molecular level to identify drivers of oncogenesis and druggable targets. We performed whole exome sequencing (WES) of 11 cTCC tumours and three matched normal samples, identifying 583 variants in protein-coding genes. The most common variant was a V-to-E missense mutation in BRAF, identified in 4 out of 11 samples (36%) via WES. Sanger sequencing identified BRAF variants in 8 out of the same 11 cTCC samples, as well as in 22 out of 32 formalin-fixed paraffin embedded (FFPE) cTCC samples, suggesting an overall prevalence of 70%. RNA-Seq was performed to compare the gene expression profiles of cTCC tumours to normal bladder tissue. cTCC tumours exhibited up-regulation of genes involved in the cell cycle, DNA repair, and antiviral immunity. We also analysed the immune landscape of cTCC using immune gene signatures and immunohistochemical analysis. A subset of tumours had characteristics of a hot tumour microenvironment and exhibited high expression of signatures associated with complete response to PD-1/PD-L1 blockade in human bladder cancer.

The Golden Retriever Lifetime Study: Assessing factors associated with owner compliance after the first year of enrollment.

BACKGROUND: The Golden Retriever Lifetime Study (GRLS) is one of the largest canine cohort studies undertaken in the United States to date. This study design allows for evaluation of multiple exposures and outcomes throughout the lifetime of each dog, but relies on participants to comply with study requirements over a long period of time. Failure to do so can lead to biased reporting of results. OBJECTIVES: To examine factors associated with dog owner compliance for GRLS. ANIMALS: Golden Retrievers (n = 3044) whose owners elected to participate in GRLS. METHODS: Prospective, cohort study. A logistic regression model was constructed to examine associations between data collected at the time of initial enrollment in GRLS and the outcome of failure to fulfill all study obligations at the end of the first year after enrollment in GRLS. RESULTS: There were 192 (6.3%) owners who did not comply with study requirements 1 year after enrollment. Owners of dogs without a record of vaccination had nearly 4 times higher odds (adjusted OR: 3.7, 95% CI: 1.5, 9.2) of being noncompliant than owners of vaccinated dogs and owners of dogs that slept in the garage had nearly 6 times higher odds (adjusted OR: 5.7, 95% CI: 1.9, 17.0) of being noncompliant than owners of dogs that slept in their bedroom. CONCLUSIONS AND CLINICAL IMPORTANCE: Survey questions about a dog's sleeping location at night and vaccination status are important indicators of an owner's odds of compliance in a prospective study. Use of similar questions during enrollment in cohort studies might help to predict owner compliance that can aid in subject selection.

FasTag: Automatic text classification of unstructured medical narratives.

Unstructured clinical narratives are continuously being recorded as part of delivery of care in electronic health records, and dedicated tagging staff spend considerable effort manually assigning clinical codes for billing purposes. Despite these efforts, however, label availability and accuracy are both suboptimal. In this retrospective study, we aimed to automate the assignment of top-level International Classification of Diseases version 9 (ICD-9) codes to clinical records from human and veterinary data stores using minimal manual labor and feature curation. Automating top-level annotations could in turn enable rapid cohort identification, especially in a veterinary setting. To this end, we trained long short-term memory (LSTM) recurrent neural networks (RNNs) on 52,722 human and 89,591 veterinary records. We investigated the accuracy of both separate-domain and combined-domain models and probed model portability. We established relevant baseline classification performances by training Decision Trees (DT) and Random Forests (RF). We also investigated whether transforming the data using MetaMap Lite, a clinical natural language processing tool, affected classification performance. We showed that the LSTM-RNNs accurately classify veterinary and human text narratives into top-level categories with an average weighted macro F1 score of 0.74 and 0.68 respectively. In the "neoplasia" category, the model trained on veterinary data had a high validation accuracy in veterinary data and moderate accuracy in human data, with F1 scores of 0.91 and 0.70 respectively. Our LSTM method scored slightly higher than that of the DT and RF models. The use of LSTM-RNN models represents a scalable structure that could prove useful in cohort identification for comparative oncology studies. Digitization of human and veterinary health information will continue to be a reality, particularly in the form of unstructured narratives. Our approach is a step forward for these two domains to learn from and inform one another.

VetTag: improving automated veterinary diagnosis coding via large-scale language modeling.

Unlike human medical records, most of the veterinary records are free text without standard diagnosis coding. The lack of systematic coding is a major barrier to the growing interest in leveraging veterinary records for public health and translational research. Recent machine learning effort is limited to predicting 42 top-level diagnosis categories from veterinary notes. Here we develop a large-scale algorithm to automatically predict all 4577 standard veterinary diagnosis codes from free text. We train our algorithm on a curated dataset of over 100 K expert labeled veterinary notes and over one million unlabeled notes. Our algorithm is based on the adapted Transformer architecture and we demonstrate that large-scale language modeling on the unlabeled notes via pretraining and as an auxiliary objective during supervised learning greatly improves performance. We systematically evaluate the performance of the model and several baselines in challenging settings where algorithms trained on one hospital are evaluated in a different hospital with substantial domain shift. In addition, we show that hierarchical training can address severe data imbalances for fine-grained diagnosis with a few training cases, and we provide interpretation for what is learned by the deep network. Our algorithm addresses an important challenge in veterinary medicine, and our model and experiments add insights into the power of unsupervised learning for clinical natural language processing.

Age at gonadectomy and risk of overweight/obesity and orthopedic injury in a cohort of Golden Retrievers.

INTRODUCTION: In the United States, gonadectomy is common and widely promoted as a component of responsible pet ownership. The recent publication of several studies examining the effect of gonadectomy on future health has challenged long-held assumptions and recommendations for gonadectomy in companion animals. The purpose of this study was to characterize the associations between gonadectomy and two outcomes: overweight/obesity and orthopedic injuries, in a large prospective study of Golden Retrievers. METHODS: Age at gonadectomy was divided into four categories: intact (reference), ≤ 6 months, > 6 months ‒ ≤ 12 months, and > 12 months. Dogs with a Purina Body Condition Score of 7 or greater were classified as overweight or obese. Orthopedic injuries considered were the first instance of veterinary-reported cranial cruciate ligament injury and clinically evident osteoarthritis. We performed survival analyses on a cohort of Golden Retrievers to estimate the associations of interest using proportional hazards. We adjusted for age at study enrollment, owner-reported activity level, and dog's sex. RESULTS: Compared to intact dogs, all gonadectomy age categories showed increased risk for the development of overweight/obesity. (≤ 6 months, HR: 1.81, 95% CI: 1.36-2.40), p-value: <0.0001; 6 months to ≤ 12 months, HR: 2.21, 95% CI: 1.77-2.73, p-value: < 0.0001; > 12 months, HR: 1.56, 95% CI: 1.24-1.96, p-value: 0.0001). Compared to intact dogs, dogs who were ≤ 6 months at gonadectomy had increased risk for orthopedic injury (HR: 4.06, 95% CI: 2.15-7.67, p-value: <0.00001). DISCUSSION: This study presents prospectively acquired data demonstrating that gonadectomy is a risk factor for both overweight/obesity and chronic non-traumatic orthopedic injuries in a prospective cohort of Golden Retrievers. Our data suggest that gonadectomy at any age is a risk factor for overweight or obesity, but delaying gonadectomy until dogs are at least 6-12 months of age may help to decrease the risk for orthopedic injury.

Inbreeding depression causes reduced fecundity in Golden Retrievers.

Inbreeding depression has been demonstrated to impact vital rates, productivity, and performance in human populations, wild and endangered species, and in recent years, the domestic species. In all cases, standardized, high-quality phenotype data on all individuals are invaluable for longitudinal analyses such as those required to evaluate vital rates of a study cohort. Further, many investigators agree upon the preference for and utility of genomic measures of inbreeding in lieu of pedigree-based estimates of inbreeding. We evaluated the association of measures of reproductive fitness in 93 Golden Retrievers enrolled in the Golden Retriever Lifetime Study with a genomic measurement of inbreeding, FROH. We demonstrate a statistically significant negative correlation between fecundity and FROH. This work sets the stage for larger scale analyses to investigate genomic regions associated with fecundity and other measures of fitness.

DeepTag: inferring diagnoses from veterinary clinical notes.

Large scale veterinary clinical records can become a powerful resource for patient care and research. However, clinicians lack the time and resource to annotate patient records with standard medical diagnostic codes and most veterinary visits are captured in free-text notes. The lack of standard coding makes it challenging to use the clinical data to improve patient care. It is also a major impediment to cross-species translational research, which relies on the ability to accurately identify patient cohorts with specific diagnostic criteria in humans and animals. In order to reduce the coding burden for veterinary clinical practice and aid translational research, we have developed a deep learning algorithm, DeepTag, which automatically infers diagnostic codes from veterinary free-text notes. DeepTag is trained on a newly curated dataset of 112,558 veterinary notes manually annotated by experts. DeepTag extends multitask LSTM with an improved hierarchical objective that captures the semantic structures between diseases. To foster human-machine collaboration, DeepTag also learns to abstain in examples when it is uncertain and defers them to human experts, resulting in improved performance. DeepTag accurately infers disease codes from free-text even in challenging cross-hospital settings where the text comes from different clinical settings than the ones used for training. It enables automated disease annotation across a broad range of clinical diagnoses with minimal preprocessing. The technical framework in this work can be applied in other medical domains that currently lack medical coding resources.

The Golden Retriever Lifetime Study: establishing an observational cohort study with translational relevance for human health.

The Golden Retriever Lifetime Study (GRLS) is the first prospective longitudinal study attempted in veterinary medicine to identify the major dietary, genetic and environmental risk factors for cancer and other important diseases in dogs. The GRLS is an observational study that will follow a cohort of 3000 purebred Golden Retrievers throughout their lives via annual online questionnaires from the dog owner and annual physical examinations and collection of biological samples by the primary care veterinarian. The field of comparative medicine investigating naturally occurring disorders in pets is specifically relevant to the many diseases that have a genetic basis for disease in both animals and humans, including cancer, blindness, metabolic and behavioural disorders and some neurodegenerative disorders. The opportunity for the GRLS to provide high-quality data for translational comparative medical initiatives in several disease categories is great. In particular, the opportunity to develop a lifetime dataset of lifestyle and activity, environmental exposure and diet history combined with simultaneous annual biological sample sets and detailed health outcomes will provide disease incidence data for this cohort of geographically dispersed dogs and associations with a wide variety of potential risk factors. The GRLS will provide a lifetime historical context, repeated biological sample sets and outcomes necessary to interrogate complex associations between genes and environmental influences and cancer.

Multiresidue analysis of pesticides in urine of healthy adult companion dogs.

The objective of this study was to determine the background exposures to pesticides as detected in urine from 21 healthy companion dogs in Northern Colorado. A panel of 301 pesticides was used to screen urine samples collected from dogs using an established ultraperformance liquid chromatography-mass spectrometry (UPLC-MS/MS) platform. Canine food intakes were controlled for one month on diets that were also screened for pesticide contents. Fifteen distinct pesticides were detected in urine. The most frequently detected compounds in canine urine samples collected over a 1 month period were atrazine, fuberidazole, imidacloprid, terbumeton, and clopyralid. Fuberidazole was the only pesticide detected in both the diets and urine. Companion dogs develop many similar chronic diseases as humans and represent a relevant model for biomonitoring combinations of environmental pesticide exposures, as well as for evaluating the potential relationships between environmental exposures and disease risk.

Evaluation of anti-human leukocyte antigen-DR monoclonal antibody therapy in spontaneous canine lymphoma.

A pilot study of anti-human leukocyte antigen (HLA)-DR monoclonal antibody (mAb) in dogs with lymphoma was undertaken to verify the suitability of a canine model to address therapeutically relevant endpoints prior to a full trial in dogs, and ultimately human investigation. In vitro studies demonstrated that L243, a murine IgG1 anti-HLA-DR, binds to normal and malignant canine lymphocytes and induces apoptosis in canine lymphoma cells. Moreover, L243 was administered safely to normal dogs and dogs with lymphoma, and bound to malignant cells in nodal tissue. Preliminary evidence of transient disease stabilization was observed in a subset of dogs with advanced-stage lymphoma following L243 immunotherapy. hL243γ4P (IMMU-114), a humanized IgG4 anti-HLA-DR, currently under evaluation preclinically for human trials, was also shown to bind malignant canine lymphocytes, and safety and pharmacokinetic data from the administration of IMMU-114 to normal dogs indicate similar behavior to L243 in these assessments. These findings provide a rationale for the use of dogs with lymphoma in safety and efficacy evaluations of anti-HLA-DR mAbs for both veterinary and human applications.

5,10-Methenyltetrahydrofolate synthetase activity is increased in tumors and modifies the efficacy of antipurine LY309887.

Methenyltetrahydrofolate synthetase (MTHFS) expression enhances folate-dependent de novo purine biosynthesis. In this study, the effect of increased MTHFS expression on the efficacy of the glycinamide ribonucleotide formyltransferase (GARFT) inhibitor LY309887 was investigated in SH-SY5Y neuroblastoma. GARFT catalyzes the incorporation of formate, in the form of 10-formyltetrahydrofolate, into the C8 position of the purine ring during de novo purine biosynthesis. SH-SY5Y neuroblastoma with increased MTHFS expression displayed a 4-fold resistance to the GARFT inhibitor LY309887, but did not exhibit resistance to the thymidylate synthase inhibitor Pemetrexed. This finding supports a mechanism whereby MTHFS increases the availability of 10-formyltetrahydrofolate for GARFT. MTHFS expression is elevated in animal tumor tissues compared to surrounding normal tissue, consistent with the dependence of transformed cells on de novo purine biosynthesis. The level of MTHFS expression in tumors may predict the efficacy of antipurine agents that target GARFT.

In vitro and in vivo evaluation of combined calcitriol and cisplatin in dogs with spontaneously occurring tumors.

PURPOSE: Calcitriol potentiates cisplatin-mediated activity in a variety of tumor models. We examine here, the effect of calcitriol and cisplatin pre-clinically and clinically in canine spontaneous tumors through in vitro studies on tumor cells and through a phase I study of calcitriol and cisplatin to identify the maximum-tolerated dosage (MTD) of this combination in dogs with cancer and to characterize the pharmacokinetic disposition of calcitriol in dogs. METHODS: Canine tumor cells were investigated for calcitriol/cisplatin interactions on proliferation using an MTT assay in a median-dose effect analysis; data were used to derive a combination index (CI). Cisplatin was given at a fixed dosage of 60 mg/m2. Calcitriol was given i.v. and the dosage was escalated in cohorts of three dogs until the MTD was defined. Serum calcitriol concentrations were quantified by radioimmunoassay. RESULTS: In vitro, CIs < 1.0 were obtained for all combinations of calcitriol/cisplatin examined. The MTD was 3.75 microg/kg calcitriol in combination with cisplatin, and hypercalcemia was the dose-limiting toxicosis. The relationship between calcitriol dosage and either Cmax or AUC was linear. Calcitriol dosages >1.5 microg/kg achieved Cmax > or = 9.8 ng/mL and dosages >1.0 microg/kg achieved AUC > or = 45 h ng/mL. CONCLUSIONS: Calcitriol and cisplatin have synergistic antiproliferative effects on multiple canine tumor cells and high-dosages of i.v. calcitriol in combination with cisplatin can be safely administered to dogs. Cmax and AUC at the MTD 3.75 microg/kg calcitriol exceed concentrations associated with antitumor activity in a murine model, indicating this combination might have significant clinical utility in dogs.

Efficacy of camptothecin and polymer-conjugated camptothecin in tumor spheroids and solid tumors.

Camptothecin (CPT) is an anti-cancer drug with low solubility in aqueous solutions, which limits its efficacy during chemotherapy. To bypass this problem, CPT was conjugated to poly(ethylene glycol) (PEG) to make CPT more hydrophilic: CM-PEG-CPT (carboxylmethylpoly(ethlyene glycol)-camptothecin), CM-PEG-GLY-CPT (carboxylmethyl-poly(ethlyene glycol)-glycine-camptothecin) and CM-PEG-SAR-CPT (carboxylmethyl-poly(ethlyene glycol)-sarcosine camptothecin) were synthesized. These conjugates differed in the amino-acid linker, which altered the hydrolysis rate of CPT from CPT-PEG. We tested the hypothesis that CPT conjugates were more effective than unconjugated CPT in effectiveness upon direct delivery to solid tumors using two systems: in vitro tumor spheroids suspended in collagen gels and in vivo solid tumors in rats. CPT was effective in spheroids, but not in flank tumors. However, when CPT was conjugated, there was improvement in the treatment of spheroids and, to a lesser extent, tumors in rats. There was no difference in therapeutic effects among the various conjugates. We conclude that conjugation of CPT to PEG enhances CPT solubility and improves effectiveness of delivery to tumors.

Sodium iodide I 131 treatment of dogs with nonresectable thyroid tumors: 39 cases (1990-2003).

OBJECTIVE: To determine outcome for dogs with nonresectable thyroid carcinomas treated with sodium iodide I 131 and identify factors associated with outcome. DESIGN: Retrospective case series. Animals-39 dogs. PROCEDURES: A definitive or presumptive diagnosis of thyroid tumor was made on the basis of cytologic or histologic examination, abnormal accumulation of sodium pertechnetate Tc 99m during scintigraphy, or both, and dogs were treated with sodium iodide I 131. Dogs with cervical thyroid tumors were evaluated 3 to 6 weeks after 131I therapy, and residual tumor was resected when feasible. RESULTS: Prior to 131I therapy, 32 dogs had a solitary mass and 7 had metastases; 21 were hyperthyroid, 16 were euthyroid, and 2 were hypothyroid. Median survival time for dogs with local or regional tumors (ie, stage II or III) was significantly longer (839 days) than median survival time for dogs with metastasis (366 days). Tumor site (cervical vs ectopic), dose of sodium iodide I 131, age, body weight, treatment (131I therapy alone vs 131I therapy followed by surgery), and serum T4 concentration prior to 131I therapy were not significantly associated with survival time. Three dogs died of radioiodine-associated myelosuppression within 3 months after treatment, but no specific factor associated with development of toxicosis was identified. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that 131I therapy may result in prolonged survival times in dogs with nonresectable thyroid tumors, regardless of serum thyroxine concentration prior to treatment. Dogs undergoing 131I therapy should be monitored for signs of bone marrow suppression.

Phase I and pharmacokinetic evaluation of the combination of orally administered docetaxel and cyclosporin A in tumor-bearing dogs.

OBJECTIVE: To determine the maximum tolerated dose and characterize the pharmacokinetic disposition of an orally administered combination of docetaxel and cyclosporin A (CSA) in dogs with tumors. ANIMALS: 16 client-owned dogs with metastatic or advanced-stage refractory tumors. PROCEDURES: An open-label, dose-escalation, single-dose, phase I study of docetaxel administered in combination with a fixed dose of CSA was conducted. Docetaxel (at doses of 1.5, 1.625, or 1.75 mg/kg) and CSA (5 mg/kg) were administered concurrently via gavage twice during a 3-week period. Plasma docetaxel concentrations were quantified by use of high-performance liquid chromatography, and pharmacokinetic disposition was characterized by use of noncompartmental analysis. Dogs' clinical signs and results of hematologic and biochemical analyses were monitored for evidence of toxicosis. RESULTS: No acute hypersensitivity reactions were observed after oral administration of docetaxel. Disposition of docetaxel was dose independent over the range evaluated, and pharmacokinetic variables were similar to those reported in previous studies involving healthy dogs, with the exception that values for clearance were significantly higher in the dogs reported here. The maximum tolerated dose of docetaxel was 1.625 mg/kg. Gastrointestinal signs of toxicosis were dose limiting. CONCLUSIONS AND CLINICAL RELEVANCE: The absence of myelosuppression suggested that the docetaxel-CSA combination may be administered more frequently than the schedule used. Further studies are warranted to evaluate combination treatment administered on a biweekly schedule in dogs with epithelial tumors.

Evaluation of strontium 90 irradiation in treatment of cutaneous mast cell tumors in cats: 35 cases (1992-2002).

OBJECTIVE: To determine the efficacy of strontium 90 beta irradiation in the management of cutaneous mast cell tumors (CMCTs) in cats. STUDY DESIGN: Retrospective case series. ANIMALS: 35 client-owned cats with CMCTs. PROCEDURE: Medical records of cats with CMCTs in which tumors were radiated by use of a strontium 90 ophthalmic applicator from 1992 to 2002 were reviewed. Cats were included if CMCT was diagnosed, there were no other sites of MCT involvement at the time of treatment, and records contained adequate follow-up information to permit retrospective assessment of local tumor control. RESULTS: 54 tumors in 35 cats were treated with a median dose of 135 Gy of strontium 90 beta irradiation, resulting in local tumor control in 53 of 54 (98%) tumors with a median follow-up time of 783 days after treatment. Median survival time was 1,075 days. Adverse effects of treatment appeared to be infrequent and of mild severity. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that strontium 90 beta irradiation resulted in long-term tumor control and should be considered an effective alternative to surgical resection in management of CMCTs in cats.